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Jahrestagung der Deutschen, Österreichischen und Schweizerischen Gesellschaften für Hämatologie und Medizinische Onkologie Basel, 9.-13. Oktober 2015: Abstracts

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Oncol Res Treat 2015;38(suppl 5):1-288



Primäre, chronische Immunthrombozytopenie

V6 - Primary, chronic immunethrombocytopenia

Imbach P.1

1Universität Basel, Uettligen, Switzerland

Introduction: Within the last 3 decades the knowledge of the history, demographics, immune pathology, clinical presentation and management dramatically have been expanded.

Methods: Prospecive registries, guidelines, the characteristics of the disturbed immune response by laboratory and clinical studies and new modalities of management are the main factors of improved knowledges.

Results: The prospective registries of immune thrombocytopenia led to new terminologies. Since only a part of patients with chronic ITP has symptoms of bleeding thrombocytopenic “Purpura” is no longer in use. The pathogenetic alterations of the innate and the adaptive immune system and the immune modulatory effects of new treatment modalities are the reasons to adapt the “i” from idiopathic to immune or autoimmune thrombocytopenia.In addition studies of the amount of platlet counts between 100-150 K showed that less than 7 percents of individuals developed ITP during further observation times. Follow up data of prospective registries with high numbers of patients revealed spontaneous recovery of up to 25% of patients between 6-12 months after diagnosis. Thus the terme “chronic ITP “ has been postponed to 12 months after diagnosis.

It was recognized that patients with no or mild bleeding management often do not need active treatment, but these patients have to be strictly followed for improvement of their quality of life.

Patients with acute and/or severe bleeding need intravenous immunoglobulins IVIG, high dose steroids and rarely platelet transfusions.

The guidelines of definitions and managements resulted in more individualized management including the quality of life of the patient.

Special considerations of management are needed in hyperactive children, in elderly persons with higher rates of comorbities, in pregnant womem, especially during labor an delivery, and in patients, who need preventive treatment (i.e befory surgery, acive sports etc.).

The management may be observation only in patient with no or mild bleeding. Active treatment modalities are now turning from the classic forms with high risk of adverse effects to acceptable side efffects (i.e. rituximab) and treatments with low adverse effects and higher rates of responses (i.e.in patients with bleeding. IVIG, in patients at risk of bleeding: thrombopoetin receptor agonists).

Conclusions: Registries may help to define special risk groups of patients with chronic ITP, which then should be evaluated by controlled studies.

Disclosure: No conflict of interest disclosed.

Wissenschaftliches Symposium


V13 - Clonal evolution in acute lymphoblastic leukemia - new insights via high-throughput sequencing and MRD analyses

Brüggemann M.1

1Universitätsklinikum Schleswig-Holstein, Campus Kiel, Medizinische Klinik II, Kiel, Germany

Acute lymphoblastic leukemia (ALL) is regarded as monoclonal disease deriving from an uncontrolled proliferation of a transformed lymphoid precursor cell. However, already early studies on immunoglobuline (IG) and T-cell receptor (TR) gene rearrangement patterns in ALL for MRD analyses challenged the concept of monoclonality as recurrently the number of detectable clonal rearrangements exceeds the maximum number of rearrangements within one clone and differences in rearrangements patterns are detected between initial diagnosis and relapse. Conventional treatment concepts are generally not affected by subclone detection because the diagnostic workup focuses on cytogenetic, molecular and immunophenotypic characteristics of the leukemic bulk with these factors being partially used for risk stratification of ALL. However, our increased understanding of molecular mechanisms of cancer and availability of drugs targeting them is changing the meaning of subclonal aberrations and their evolution in the diagnostics, treatment and follow-up of ALL. Next generation sequencing (NGS) of amplified IG/TR gene rearrangements allows sensitive MRD assessement of leukemia associated IG/TR gene rearrangements and gives insights into composition of additional low frequency subclones. Published studies identified up to thousands of different subclones per patients using this method. Also within the GMALL a series of diagnostic and relapse ALL samples were investigated using IG/TR NGS demonstrating high levels of oligoclonality. However, within this limited series none of the minor subclones at diagnosis became dominant at relapse so that the meaning of these subclones remains to be elucidated. Molecular analyses of other recurrent genetic alterations like intrageneic deletions of the BTG1, IKZF1 and ERG confirm ALL clonal variegation also for these aberrations. If molecular aberrations serve as treatment targets monitoring of these targets can identify clonal selection processes. If e. g. NUP214-ABL1 positive T-ALL is treated with tyrosine kinase inhibitors selection of NUB214-ABL1 negative subclones is a possible mechanism of resistance. Also low-level resistance mutations like BCR-ABL tyrosine kinase domain mutations in newly diagnosed or recurrent BCR-ABL positive ALL may play a future role for choice of tyrosine kinase inhibitors. Therefore, sensitive mutation detection strategies to monitor driver mutations or druggable targets may complement the analysis of bulk MRD.

Disclosure: No conflict of interest disclosed.


Infektionen in der Onkologie und Hämatologie

V16 - Diagnosis of fungal infections - current guideline 2015

Schwartz S.1

1Charité Campus Benjamin Franklin, Medizinische Klinik mit Schwerpunkt Hämatologie und Onkologie, Berlin, Germany

Invasive fungal infections (IFI) remain a diagnostic challenge, although progress with development of new tools has been made. To establish a definite diagnosis of an IFI, growth of the infecting fungus from an otherwise sterile site or histopathological or microscopic demonstration of the fungus with evidence of tissue invasion or damage or microscopy with characteristic morphological findings is usually required. This is of particular importance in growing numbers of patients with infections caused by rare, emerging fungi. However, invasive procedures are not always feasible due to frequent coagulation impairment in immunocompromised patients. Imaging (eg, high-resolution CT-scan) and serological techniques (eg, galactomannan) are well established, but, with few exceptions (eg, cryptococcosis), findings support a probable diagnosis and are not sufficient to establish a definite diagnosis. Fungus-specific PCR-techniques, either targeting pan-fungal or genus-specific gene sequences, could detect presence of fungal nucleic acids in blood specimens or bronchoalveolar lavage (BAL). This technique displays an enhanced sensitivity and specificity, particularly when BAL samples are analysed. Combination of results from PCR and serological studies could further enhance the diagnostic sensitivity and specificity. However, various PCR techniques exist and there are no commonly accepted protocols, which currently precludes incorporation of this technique as mandatory into diagnostic algorithms.

A new, innovative diagnostic tool, which detects volatile, fungus-specific compounds in breath samples from patients with pulmonary aspergillosis, has been recently developed. This technique not only has the potential to be used as a rapid bedside test, but to distinguish infections with different fungi. Another novel approach uses detection of fungus-specific, CD4-positive T-cells in blood samples. The sensitivity of this test is enhanced by brief in vitro stimulation with fungal antigens and subsequent detection of CD154 up-regulation by flow cytometry. This diagnostic approach is somewhat limited by large volumes of diagnostic blood specimens, but could differentiate between infections with various types of fungi.

In conclusion, the updated guideline about the diagnosis of fungal infections will not only gather new data about established diagnostic tools, but incorporate new, innovative diagnostic tools.

Disclosure: Stefan Schwartz: Advisory Role: Pfizer Pharma GmbH, BTG International Ltd, AMGEN; Financing of Scientific Research: MSD Sharp & Dohme, Pfizer Pharma GmbH, Gilead Sciences GmbH, Astellas GmbH, AMGEN.

V18 - Emergence of multi-resistant bacteria in hematological and oncological settings

Biehl L.M.1,2

1Klinik I für Innere Medizin, Uniklinik, Köln, Germany, 2Deutsches Zentrum für Infektionsforschung (DZIF), Bonn-Köln, Germany

The prevalence of multi-resistant bacteria (MRB), in particular gram negative bacteria, is increasing worldwide. While these bacteria are most frequently found colonizing the intestinal tract, they can cause invasive infections especially in immunocompromised patients. Due to the limited therapeutic options and the frequent delay in adequate treatment, infections are associated with an increased mortality as compared to non multi-resistant pathogens.

The epidemiology of extended-spectrum ß-lactamase producing Enterobacteriaceae (ESBL-E) and carbapenem-resistant gram negative bacteria is highly depending on regional factors and charateristics of the population under scrutiny. So far, carbapenem-resistance is still infrequent in Germany and Northern Europe. Certain risk factors including previous antibiotic treatment and medical care in high-risk countries have been identified. In settings with hematological and oncological patients, a screening strategy based on the local epidemiology should be considered. Furthermore, individualized adaption of empirical antibiotic treatment in febrile neutropenia depending on colonization status may be beneficial. However, this needs to be implemented with caution to avoid overuse of broad-spectrum antibiotcs and further selection pressure.

Disclosure: No conflict of interest disclosed.


Palliativmedizin interprofessionelle und interdisziplinäre Sichtweise

V22 - The view on palliative medicine from different perspectives: Palliative care, medical oncology, internal medicine

Lordick F.1

1Universitäres Krebszentrum Leipzig (UCCL), Leipzig, Germany

6 years ago ASCO released a statement that was adopted by the Arbeitsgemeinschaft Palliativmedizin of the German Cancer Society: “Palliative cancer care is the integration into cancer care of therapies that address the multiple issues that cause suffering for patients and their families and impact their life quality. Effective provision of palliative cancer care requires an interdisciplinary team that can provide care in all patient settings, including outpatient clinics, acute and long-term care facilities, and private homes. Changes in current policy, drug availability, and education are necessary for the integration of palliative care throughout the experience of cancer, for the achievement of quality improvement initiatives, and for effective palliative cancer care research. The need for palliative cancer care is greater than ever notwithstanding the strides made over the last decade. Further efforts are needed to realize the integration of palliative care in the model and vision of comprehensive cancer care by 2020.”

The publication of the German multidisciplinary and multiprofessional evidence-based and expert-consented (S3) “Guideline for Palliative Care of Patients with Incurable Cancer” is an important step into this direction. Another milestone shall be the planned law for improving hospice and palliative care in Germany (Hospiz- und Palliativgesetz HPG) which is in preparation.

In the past, research data in the field of palliative care were scarce. Actions were taken on a more intuitive and empirical basis. Meanwhile, important research results have been published elucidating for instance the role of early integration of palliative care into routine oncology. This raises the question who should deliver palliative care to cancer patients. First and foremost, we need to better understand the real burden on our patients, their preferences and care needs during the course of an incurable disease. This is why we started a multidisciplinary research program in the scientific working group (Sektion B) of the German Cancer Society. Eventually, we need to tailor “who shall provide” and “how to deliver” optimized palliative care. The current understanding is that every physician who cares for patients with severe and incurable diseases should have been trained and should be able to deliver general aspects of symptom relieve and palliative care. For more complicated and extended palliative and “end-of-life” care, specialized teams are required.

Disclosure: No conflict of interest disclosed.

Freier Vortrag

Pankreaskarzinom / Hepatobiliäres Karzinom / Melanom

V23 - The pancreatic carcinoma - treatment research and treatment reality in oncology practices

Weber K.1, Schlichting A.2, Tessen H.-W.3,4

1Onkologische Praxis Hellersdorf, Berlin, Germany, 2rgb Onkologisches Management GmbH, Sarstedt, Germany, 3Onkologische Kooperation Harz, Goslar, Germany, 4Projektgruppe Internistische Onkologie (PIO), Goslar, Germany

Introduction: According to the 2013-S3 guideline “Exocrine pancreatic carcinoma“, surgery is the only curative treatment option for pancreatic carcinoma. Gemcitabine, gemcitabine/ erlotinib, FOLFIRINOX are recommended as adjuvant or palliative 1st-line therapy, 5-FU/oxaliplatin as 2nd-line therapy.

Methods: Data related to the treatment of pancreatic cancer have been analysed within the national scientific register ONCOReg since March 2009. The register contains the records of a total of 30,186 patients from 355 practices in 15 federal states, including 1,722 pancreatic carcinomas from 66 practices (2,542 therapies). 1,522 disease histories are available for evaluation so far.


Patient characteristics:

Gender: 821 (53.9%) male, 701 (46.1%) female

Age at initial diagnosis: 69 (35-95) year; 22% older than 75 years

UICC stages: 43 (2.8%) I; 540 (35.5%) II; 154 (10.1%) III; 698 (45.9%) IV, 88 (5.8%) n.k.

634 (41.7%) patients were operated. An R0 resection was achieved in 491 cases (77.4%), an R1 resection in 109 (17.2%) patients.

Adjuvant therapies: 420 (66.2%) patients, R1-additive therapies: 90 (14.2%); 490 (96.1%) gemcitabine monotherapy. Duration of treatment at 5.1 resp. 4.7 months.

Survival rates of the adjuvant resp. R1-additive therapy: DFS 14.1 resp. 9.1 months; OS 33.6 resp. 18.8 months.

Palliative therapies: 1,268 patients received 1,978 palliative therapies: 726 (57.3%) gemcitabine; 371 (29.3%) gemcitabine/erlotinib; 138 (10.9%) FOLFIRINOX; 130 (10.3%) OFF; 88 (7.0%) FUFOX, 68 (5.4%) gemcitabine/albumin-bound paclitaxel; 54 (4.3%) gemcitabine/oxaliplatin. The median duration of treatment was at 78 days (91 days for 1st-line therapies).

1,268 patients received a 1st-line therapy; 519 (41.0%) a 2nd-line; 143 (11.3%) a 3rd-line therapy.

The responses were assessed in 1,846 therapies, and the objective response (CR/PR) was at 9.9%. In 34.0% of the cases a halt of the progression of the disease was achieved. A progression had to be observed in 36.3%, with the PD rate increasing with increasing therapy line (1st-line 32.6%; 2nd-line 43.3%; 3rd-line 50.0%).

Survival data:

PFS: 1st-line 4.9 mths.; 2nd-line 3.3 mths.

OS: 1st-line 9.0 mths.; 2nd-line 6.2 mths.

Conclusions: Data collection and analysis is an integral part of the routine in oncology practices.Data on the duration of therapy, the response and survival of selected first- and second-line therapies will be presented.

Disclosure: No conflict of interest disclosed.

V24 - Transduction with C-C-chemokine receptor type 4 (CCR4) enhances tumor-specific migration of adoptively transferred T cells in a model of pancreatic cancer

Rapp M.1, Grassmann S.1, Endres S.1, Anz D.1, Kobold S.1

1Ludwig-Maximilians-Universität, Abteilung für Klinische Pharmakologie, München, Germany

Introduction: Regulatory T cells selectively express C-C-chemokine receptor type 4 (CCR4) and are attracted by intratumoral CCL22 while CCR4 is absent from most cytotoxic T cells. We hypothesized that inducing forced expression of CCR4 on adoptively transferred cytotoxic T cells could enhance therapeutic efficacy.

Methods: CCR4 and a new non-functional CCR4-deletion mutant were cloned into the retroviral vector pMP71 and transduced into murine ovalbumin-specific T cells (OT-1). T cell migration and function in vitro were assessed using transwell assays. In vivo efficacy was assessed in a murine model induced with the syngeneic pancreatic cell line Panc02-OVA.

Results: In vitro, CCR4- but not CCR4del-transduced OT-1 T cells specifically migrated towards recombinant CCL22 and tumor-induced CCL22 (25-fold increase). In vivo, CCR4-transduced OT-1 T cells cured 50% of mice bearing established Panc-OVA tumors compared to 12% in the CCR4del- or untransduced OT-1 T cell- treated mice (n = 8 per group, p < 0.0001). Cured mice were protected from rechallenge, indicating the induction of anti-tumor immunity. Enhanced numbers of CCR4-transduced OT-1 T cells compared to CCR4del-transduced cells were found in the tumor, suggesting enhanced homing to the tumor site as one mode of action of CCR4-transduced T cells.

Conclusions: The transduction of cytotoxic T cells with CCR4 represents a new therapeutic approach for effectively guiding adoptively transferred T cells into pancreatic tumors and thereby inducing a potent antitumor immune response.

Disclosure: No conflict of interest disclosed.

V25 - Evaluation of systemic inflammatory response (SIR) markers in pancreatic ductal adenocarcinoma (PDAC)

Markus M.1, Kasper S.1, Noureddine R.1, Abramczyk M.1, Paul A.2, Gerken G.3, Schmid K.W.4, Markus P.5, Schumacher B.6, Meiler J.1, Wiesweg M.1, Kaiser G.2, Dechêne A.3, Trarbach T.7, Schuler M.1, Abendroth A.1

1Universitätsklinikum Essen, Westdeutsches Tumorzentrum, Innere Klinik (Tumorforschung), Essen, Germany, 2Universitätsklinikum Essen, Westdeutsches Tumorzentrum, Klinik für Allgemein-, Viszeral- und Transplantationschirurgie, Essen, Germany, 3Universitätsklinikum Essen, Westdeutsches Tumorzentrum, Klinik für Gastroenterologie und Hepatologie, Essen, Germany, 4Universitätsklinikum Essen, Westdeutsches Tumorzentrum, Institut für Pathologie, Essen, Germany, 5Elisabeth Krankenhaus Essen, Klinik für Allgemein-, Viszeral- und Unfallchirurgie, Essen, Germany, 6Elisabeth Krankenhaus Essen, Klinik für Gastroenterologie, Essen, Germany, 7iOMEDICO AG, Freiburg, Germany

Introduction: Intratumoral inflammation is a known hallmark of cancer and can promote tumorigenesis especially in PDAC. Recently, anti-inflammatory agents like JAK2- or CCR2-inhibitors showed promising results in early clinical trials in patients with metastatic PDAC. So far, valid biomarkers for the response prediction to these novel anti-inflammatory drugs are missing but essential, as only a subset of patients seem to respond. We retrospectively evaluated the prognostic and predictive value of routinely assessed inflammatory serum response (SIR) biomarkers in patients with advanced PDAC treated with systemic chemotherapy at the West German Cancer Center (WTZ) in Essen, one of the 13 Oncology Centers of Excellence in Germany.

Methods: A total of 245 PDAC patients treated at the WTZ from 2005-2012 were retrospectively evaluated for the SIR markers: Glasgow Prognostic Score (GPS, mGPS), C-reactive-Protein (CRP), Neutrophile-/Lymphocyte Ratio (NLR), Lymphocyte-Monocyte Ratio (LMR) and Platelet-/Lymphocyte Ratio (PLR). SIR markers were correlated with clinico-pathological findings, response to chemotherapy and overall survival (OS) using Kaplan-Meier curves, univariate and multivariate Cox proportional models.

Results: In metastatic PDAC median OS was 8.5 months (95% CI 6.95-10.35). All evaluated SIR markers except of PLR were significantly associated with reduced OS (Figure 1). The HR and (95% CI) for PLR>200 were: 0.829 (0.512-1.309, p = 0.403), for NLR< 5: 1.899 (1.175-3.069, p = 0.009), for LMR< 2.8: 1.926 (1.111-3.338, p = 0.019), for CRP>normal: 2.536 (1.568-4.101, p < 0.001), for GPS>2: 3.266 (1.269-8.407, p = 0.014) and for mGPS: 1.993 (1.010-3.930, p = 0.047). In patients receiving gemcitabine monotherapy (48.8%) the evaluated SIR markers had no impact on progression free survival (PFS), but in patients receiving combination chemotherapy (51,2%, mostly gemcitabine/cisplatin or oxaliplatin) most evaluated SIR markers correlated with reduced PFS.

Conclusion: SIR biomarkers, routinely assessed in clinical settings can be easily used as prognostic and predictive markers to optimize therapeutic strategies in PDAC.


Disclosure: No conflict of interest disclosed.

V26 - Progression-free and overall survival of patients with advanced or metastatic pancreatic cancer in German outpatient centers - first outcome data from the clinical TPK Registry

Hegewisch-Becker S.1, Hofheinz R.2, Wolf T.3, Aldaoud A.4, Harde J.5, Kopfmann S.5, Marschner N.6

1Onkologische Schwerpunktpraxis, Hamburg, Germany, 2Universitätsklinikum, Mannheim, Germany, 3Gemeinschaftspraxis Hämatologie - Onkologie, Dresden, Germany, 4Dr. Aldaoud - Dr. Schwarzer Forschungsgesellschaft mbH, Leipzig, Germany, 5iOMEDICO, Freiburg i.Br., Germany, 6Praxis für Interdisziplinäre Onkologie & Hämatologie, Freiburg i.Br., Germany

Introduction: Pancreatic cancer is the fourth leading cause of cancer death in Europe. About 80% of patients (pts) are diagnosed with locally advanced or metastatic disease and with a 5-year survival rate of less than 2%. Thus, pancreatic cancer remains a disease with dismal prognosis. Within the last decade new treatment options have become available. Clinical registries provide insight into treatment reality and real-life effectiveness outside of clinical trials.

Methods: The Tumor Registry Pancreatic Cancer (TPK) is a prospective, multicenter, observational study of pts with advanced or metastatic pancreatic cancer receiving systemic antineoplastic palliative therapy. Since February 2014, 99 study sites in Germany have recruited more than 400 pts at the start of their first-line treatment. A broad set of data regarding pts and tumor characteristics, comorbidities, factors affecting therapeutic decision making, previous and ongoing systemic treatments as well as outcome data such as response, progression free (PFS) and overall survival data are recorded. Here, characteristics and first-line treatment of pts with advanced or metastatic pancreatic cancer are reported for the first 187 pts recruited into the registry. In addition, outcome data such as first-line PFS and six-month survival rate are presented.

Results: At the start of palliative treatment pts mean age was 69 years (yrs), 55% of pts were male. 73% of pts had metastatic disease. Palliative 1st-line treatment was generally based on gemcitabine in combination with nab-paclitaxel (GEM+Nab-Pac, 42%), gemcitabine monotherapy (GEM mono, 27%) and less frequently on FOLFIRINOX (18%). Pts receiving GEM mono are older and have more comorbidities than pts receiving GEM+Nab-Pac or FOLFIRINOX (mean age 74 yrs vs. 69 and 61 yrs; mean Charlson comorbidity score 1.1 vs. 0.4 and 0.3). After a median follow-up of 7.6 months, median PFS of all pts is 5.3 months (95% CI: 4.7-6.2). Overall survival rate at 6 months is 62%.

Conclusion: In routine practice, pts with metastatic or advanced pancreatic cancer are treated according to age and comorbidities. Although pts in routine practice are markedly older than pts in clinical trials, median PFS is promising. Further analysis will investigate prognosis factors and the effectiveness of different treatment options.

Disclosure: No conflict of interest disclosed.

V27 - Evaluation of systemic inflammatory response (SIR) markers in advanced biliary tract cancer (ABTC)

Abendroth A.1, Markus M.1, Abramczyk M.1, Noureddine R.1, Paul A.2, Gerken G.3, Schmid K.W.4, Markus P.5, Schumacher B.6, Meiler J.1, Wiesweg M.1, Kaiser G.2, Dechêne A.3, Trarbach T.7, Schuler M.1, Kasper S.1

1Universitätsklinikum Essen, Westdeutsches Tumorzentrum, Innere Klinik (Tumorforschung), Essen, Germany, 2Universitätsklinikum Essen, Westdeutsches Tumorzentrum, Klinik für Allgemein,- Viszeral- und Transplantationschirurgie, Essen, Germany, 3Universitätsklinikum Essen, Westdeutsches Tumorzentrum, Klinik für Gastroenterologie und Hepatologie, Essen, Germany, 4Universitätsklinikum Essen, Westdeutsches Tumorzentrum, Institut für Pathologie, Essen, Germany, 5Elisabeth Krankenhaus Essen, Klinik für Allgemein,- Viszeral- und Unfallchirurgie, Essen, Germany, 6Elisabeth Krankenhaus Essen, Klinik für Gastroenterologie, Essen, Germany, 7iOMEDICO AG, Freiburg, Germany

Introduction: Systemic inflammatory response (SIR) markers are prognostic in several cancers but have not been evaluated systematically in large ABTC series. We evaluated the prognostic and predictive value of the modified Glasgow Prognostic Score (mGPS), C-reactive protein (CRP), Neutrophile-/Lymphocyte Ratio (NLR), Lymphocyte-Monocyte Ratio (LMR) and Platelet-/Lymphocyte Ratio (PLR) in patients with ABTC treated with systemic chemotherapy at the West German Cancer Center (WTZ) in Essen, one of the 13 Oncology Centers of Excellence in Germany.

Methods: SIR markers were retrospectively evaluated in 147 patients (pts) with ABTC treated with palliative systemic chemotherapy at the WTZ from 2005-2012. Cutoff values for PLR, NLR and LMR defining high vs. low scores were selected by rounding the mean values for the entire cohort, which were 218, 4.6 and 2.6, respectively. For CRP, values above 10mg/dl were selected. The mGPS is based on CRP and albumin. Pts who had both elevated CRP (>10mg/l) and hypoalbuminaemia (< 35g/l) had a score of 2. SIR markers were correlated with clinico-pathological findings, response to chemotherapy, progression free (PFS) and overall survival (OS) using Kaplan-Meier curves and Cox proportional hazard models.

Results: In ABTC median OS was 13.9 months (95% CI 10.5-17.4). All evaluated SIR markers were significantly associated with reduced OS (table 1). Interestingly, in pts receiving combination chemotherapies (78.8%, mostly gemcitabine/oxaliplatin or cisplatin) none of the evaluated SIR markers had an impact on PFS or OS. In contrast pts receiving monochemotherapy (21.2%, mostly gemcitabine) SIR markers strongly correlated with reduced PFS and OS.

Conclusion: SIR biomarkers, routinely assessed in clinical settings can be easily used as prognostic and predictive markers to optimize therapeutic strategies in ABTC. To overcome the negative prognostic impact of high SIR combination chemotherapies should be recommended.


Disclosure: No conflict of interest disclosed.

V28 - H3K9-active demethylases disable oncogene-induced senescence, drive melanomagenesis, and are promising therapeutic targets

Yu Y.1, Yue B.1, Ji S.2,3, Lohneis P.4, Werner-Klein M.5, Hummel M.4, Dörken B.1,2, Lee S.1,2, Schmitt C.1,2

1Max-Delbrück-Center for Moleculare Medicine, Berlin, Germany, 2Charité - Universitätsmedizin Berlin, Medical Department of Hematology, Oncology and Tumor Immunology and Molekulares Krebsforschungszentrum, Berlin, Germany, 3Chinese Academy of Medical Sciences, Peking Union Medical College, Peking, China, 4Charité - Universitätsmedizin Berlin, Department of Pathology, Berlin, Germany, 5University Regensburg, Institute of Immunology, Regensburg, Germany

Introduction: Oncogene-induced senescence (OIS) is a failsafe program against imminent transformation and further propagation of (pre-)malignant cells. Paradigmatic are Ras- or BRAF-driven nevus cell nevi, whose senescent state serves as a barrier to melanoma formation. Mechanistically, histone H3 lysine 9-trimethylation (H3K9me3)-governed silencing of S-phase-promoting E2F target genes is essential for the sustained cell-cycle arrest. Given the importance of H3K9me3 and the frequent overexpression of H3K9-active demethylases in various cancer types, we hypothesized that candidate demethylases such as LSD1 and JMJD2C might exert their pro-tumorigenic potential via blocking OIS.

Methods: Human and murine fibroblasts, primary human melanocytes, nevus and melanoma biopsies, melanoma cell lines and primary single disseminated melanoma cells (DMC) were subjected to functional analyses (expression, growth, senescence, transformation) in culture or as xenotransplants in nude mice after stable gene transfer or knockdown of relevant moieties, including LSD1, JMJD2C, oncogenic Ras and BRAF. In addition, pharmacological inhibition of LSD1 and JMJD2C was exploited in vitro and in vivo.

Results: Ras- or BRAF-expressing mouse or human fibroblasts, as well as primary human melanocytes bypassed OIS if co-expressing LSD1 or JMJDC2. Moreover, stable expression of LSD1 or JMJD2C transformed Ras-driven mouse embryo fibroblasts. Importantly, activation of H3K9 demethylases in senescent cells enforced cell-cycle re-entry, i.e. licensed reversal of OIS (“escape”). On the contrary, inhibition of LSD1 or JMJD2C in demethylase- and Ras-overexpressing fibroblasts by RNA interference or chemical inhibitors restored senescence. We detected LSD1 and JMJD2C expression in melanoma biopsies, but not premalignant nevi samples, and found high-level H3K9-active demethylase expression as a prominent feature of melanoma cell lines. Inhibiting LSD1 or JMJD2C in melanoma cell lines or DMC induced cellular senescence, and controlled xenograft growth in nude mice. Remarkably, targeting LSD1 and JMJD2C even blocked melanoma progression of BRAF inhibitor (i.e. Vemurafenib)-resistant cells in vitro and in vivo.

Conclusions: Our data unveil the essential but highly dynamic role of the H3K9me3 mark in OIS, the - in principle - intrinsic reversibility of this otherwise very stable condition, and suggest H3K9-active demethylases as promising targets for novel anti-cancer treatment strategies.

Disclosure: No conflict of interest disclosed.


Ovarial- und Uteruskarzinom

V31 - Immunotherapy of ovarian cancer

Weiss L.1, Mlineritsch B.1, Greil R.1

1Paracelsus Medizinische Universität Salzburg, Universitätsklinik für Innere Medizin III, Salzburg, Austria

Ovarian cancer is the second most common gynecologic malignancy and with only 45% of patients surviving 5 years after diagnosis, new therapeutic strategies are urgently needed. A vast body of evidence suggests that in general ovarian cancer cells can be recognized by the immune system and the presence of tumor infiltrating T-cells is associated with a significantly better clinical outcome. Despite this immunogenicity overt disease can evolve and eventually patients may succumb to their disease - possibly due to immune evasion strategies exerted by the tumor. As one of these mechanisms, the Programmed Death-Ligand 1 (PD-L1) can be expressed by tumor cells and by binding to its receptor - the Programmed Death Protein 1 (PD-1) - it can down-regulate the function of T-cells. High expression of PD-L1 in ovarian cancer has been shown to be associated with significantly shorter survival. Earlier immunotherapy trials using intraperitoneal interleukin 2 and/or the adoptive transfer of tumor-infiltrating T-cells provided first evidence of clinical efficacy of immunotherapy in ovarian cancer. Also several vaccination studies could demonstrate an increase in tumor-specific T-cells which in some cases even conferred clinical benefit. Various tumor-associated antigens such as NY-ESO-1 or individual patients' tumor cell lysate have been used for vaccination strategies. So called immune checkpoint inhibitors have shown unprecedented responses in notoriously difficult-to-treat cancers such as melanoma and lung cancer. The anti-PD-1 antibodies nivolumab and pembrolizumab, as well as the anti-PD-L1 antibodies BMS-936559 and avelumab are the first representatives of this new class of drugs having shown highly promising results in ovarian cancer, with some patients even achieving complete and also durable remissions. Combining different immunotherapeutic approaches might further increase the clinical efficacy in the future.

Disclosure: Lukas Weiss: Other Financial Relationships: Reisekostenerstattung (Merck) Richard Greil: No conflict of interest disclosed.

Freier Vortrag

AML experimentell I

V32 - DNMT3A mutations (DNMT3Amut) in acute myeloid leukemia (AML): monitoring of minimal residual disease (MRD). A study of the AML Study Group (AMLSG)

Gaidzik V.I.1, Weber D.1, Paschka P.1, Krieger S.1, Kaumanns A.1, Krönke J.1, Kapp-Schwörer S.1, Köhne C.-H.2, Horst H.-A.3, Schmidt-Wolf I.G.H.4, Held G.5, Kündgen A.6, Ringhoffer M.7, Götze K.8, Kindler T.9, Fiedler W.10, Wattad M.11, Corbacioglu A.1, Bullinger L.1, Schlegelberger B.12, Thol F.12, Heuser M.12, Ganser A.12, Schlenk R.F.1, Döhner H.1, Döhner K.1

1Uniklinik, Ulm, Germany, 2Klinikum, Oldenburg, Germany, 3Universitätsklinikum Schleswig-Holstein-Campus, Kiel, Germany, 4Universitätsklinikum, Bonn, Germany, 5Universitätsklinikum des Saarlandes, Homburg, Germany, 6Universitätsklinikum, Düsseldorf, Germany, 7Städtisches Klinikum Karlsruhe GmbH, Karlsruhe, Germany, 8Klinikum rechts der Isar der Technischen Universität, München, Germany, 9Universitätsmedizin, Mainz, Germany, 10Universitätsklinikum Hamburg-Eppendorf, Hamburg, Germany, 11Kliniken Essen Süd, Ev. Krankenhaus Essen-Werden gGmbH, Essen, Germany, 12Medizinische Hochschule Hannover, Hannover, Germany

Introduction: The awareness of DNMT3Amut as a pre-leukemic lesion and founder mutation in AML as well as its presence in apparently healthy elderly brought new light into the disease. Within our treatment trials we sought to analyze if MRD monitoring in DNMT3Amut patients (pts) is clinically relevant and delivers further information for clonal hematopoiesis.

Methods: We analyzed the DNMT3Amut R882H (n = 111) and R882C (n = 48) in AML pts entered on three AMLSG treatment trials [AML HD98A (n = 14; NCT00146120), AMLSG 07-04 (n = 87; NCT00151242), AMLSG 09-09 (n = 58; NCT00893399)] using a cDNA-based RQ-PCR-assay by TaqMan technology (sensitivity between 10-3 and 10-4).

Results: In total, 1,168 samples [bone marrow (BM), n = 615; peripheral blood (PB), n = 553] from 159 DNMT3Amut pts were analysed [diagnosis, n = 256; during therapy, n = 719; follow-up, n = 193]. Median BM DNMT3Amut transcript levels (TL) at diagnosis were 12690 (range, 0-54280); TL were not associated with known clinical characteristics, mutations in NPM1 or FLT3 or impact on OS and RFS. DNMT3Amut TL during therapy were significantly higher in BM than in PB after induction I, consolidation I and II (p = 0.01; p = 0.0003; p = 0.01). After double induction (DI) there was no difference in the median TL between 102 pts in complete remission (CR) and 12 pts not in CR. BM DNMT3Amut TL as log 10 transformed continuous variable during therapy had no influence on the endpoints death or relapse. The greatest TL reduction was seen after induction I (one log); after DI and end of therapy (ET), only 7/75 and 4/63 BM samples became MRD negative. At these two time-points MRD positivity did not impact OS (p = 0.89; p = 0.73) and remission duration (RD; p = 0.61; p = 0.30). Next we investigated the MRD DNMT3Amut log10-reduction (compared to level at diagnosis) with the median as a cut-off. There was no significant correlation for pts with a higher compared to a lower TL reduction for OS and RD after DI and ET (p = 0.87; p = 0.38; p = 0.67; p = 0.57). The BM DNMT3Amut TL as 4 increasing equally sized intervals according to the quartiles of the distribution revealed no prognostic impact after DI and ET on OS and RD (p = 0.24; p = 0.85; p = 0.38; p = 0.44).

Conclusions: In our study neither DNMT3Amut TL nor its kinetics had a prognostic impact. Most pts showed persistent DNMT3Amut TL supporting the role of clonally derived hematopoiesis. By serial investigation of DNMT3Amut and its co-mutations the role of clonal evolution of AML will be further elucidated.

Disclosure: No conflict of interest disclosed.

V33 - SH-2251 as a new possible treatment for AML

Lams R.F.1, Botezatu L.1, Hönes J.1, Michel L.1, Köster R.1, Al-Matary Y.1, Dührsen U.1, Khandanpour C.1

1University Hospital Essen, Department of Hematology, Essen, Germany

Acute myeloid leukemia (AML) is a malignant neoplasia of the myelopoiesis. Despite intensive therapy the overall five year survival rate is under 30%, warranting new approaches. Oncorequisite factors like Growth factor independence 1 (Gfi-1) are promising new approaches to treat AML.

Gfi-1 is a transcriptional repressor, which bind to and restricts p53 activity by modifying p53. In addition to this, it binds to the regulatory elements of pro-apoptotic genes such as Puma, Noxa (Pmaip1) and Bax (Bbc3). We could previously show that genetically inhibiting Gfi1 in leukemic cells leads to massive apoptosis in leukemic cells without side effects on normal cells. However no drugs are available to inhibit Gfi1 expression in patients. In this study, we examined the suitability of the novel thioamide-related compound SH-2251, which represses Gfi1 expression, to cure AML. To address this, we performed in vitro and in vivo studies using SH-2251, compound, which dramatically decreases Gfi1 mRNA expression.

We treated different Gfi1 expressing human and murine AML cell lines such as Kasumi1, KG1, SKNO, HL-60 and C1498 by various concentrations of SH-2251. To rule out unspecific toxic effects we also testes non Gfi1 expressing cell lines. The results revealed that SH-2251 mediated inhibition of Gfi1 leads to an increased level of apoptosis and decreased cell numbers in the different Gfi1 expressing cell lines. Moreover, SH-2251 did not influence non-Gfi1 expressing tumor cell-lines, thus highlighting the specificity of this compound.

To confirm our findings in vivo, we next investigated the effects of oral administration of SH-2251 (50mg/kg) in an AML mouse model. We transplanted mice with murine leukemic cells expressing the human oncofusion protein AML1/ETO9a, which can be recurrently found in AML patient cohorts. We observed that SH-2251 treatment inhibited AML development when compared with vehicle-administated control group.

In summary, SH-2251 could be a new promising approach to treat Gfi1 expressing AML independent of their leukemia driving mutations, potentially opening a novel treatment approach for these patients.

Disclosure: No conflict of interest disclosed.

V34 - Gfi1b plays an important role in initiation and progression of murine and human acute myeloid leukemia (AML) by regulating key oncogenic pathways

Thivakaran A.1, Botezatu L.1, Hönes J.M.1, Zeller A.1, Michel L.1, Görgens A.2, Lennartz K.3, Köster R.1, Opalka B.1, Giebel B.2, Dührsen U.1, Khandanpour C.1

1University Hospital Essen, Department of Hematology, Essen, Germany, 2University Hospital Essen, Institute for Transfusion Medicine, Essen, Germany, 3University Hospital Essen, Institute for Cell Biology, Essen, Germany

Gfi1b (growth factor independence 1b) is a transcription factor important for the development of erythroid and megakaryocytic lineage. It also regulates quiescence and cell cycle progression of hematopoietic stem cells (HSCs) and early progenitor cells. We explored how different levels of Gfi1b influence the onset and progression of acute myeloid leukemia (AML). Published and own patient's data indicated that Gfi1b is expressed at a lower level in leukemic blasts and leukemic stem cells (LSC) compared to non-malignant cells. To investigate the role of Gfi1b in initiation and progression of AML we used the Nup98/HoxD13 MDS mouse model that recapitulates key features of MDS including transformation to AML.

To study the influence of different Gfi1b level on leukemia development, we have used two different mouse strains: Gfi1bGFP/WT mice, in which one allele of Gfi1b is replaced by GFP and a conditional mouse strain (Mx Cre tg Gfi1b fl/fl) in which the expression of Gfi1b can be abrogated after pIpC injection. We crossed these mice with Nup98HoxD13 MDS mice and observed that Gfi1b heterozygosity accelerated AML development (p < 0.0001) compared to wt mice. Complete loss of Gfi1b resulted in a significantly earlier onset of AML (p = <0.0001). Loss of Gfi1b alone was not lethal for more than the 6 months. While all Gfi1b WT AML arising in the Nup98HoxD13 tg mice could be clearly attributed to the myeloid lineage, Gfi1b-deficient leukemic cells did not express any lineage marker but expressed very high levels of c-Kit, indicating that KO of Gfi1b is associated with immature leukemia. Gfi1b induces demethylation of H3K4 and deacetylation of H3K9. We found that Gfi1b deficient cells had a genomewide upregulation of H3K9 acetylation and methylation and that this was associated with pathways in stem cell and integrin regulation. Gfi1b regulates ITGB3 (integrin beta 3), which is required to promote growth and expansion of LSC. Conditional deletion of Gfi1b increased expression ITGB3 in bone marrow leukemic. On a molecular level, we found that loss of Gfi1b leads to increase levels of ROS (Reactive oxygen species). Excessive amounts of ROS causes the peroxidation of nucleic acids that could lead to carcinogenesis. In Gfi1b-KO leukemic cells gene expression arrays revealed upregulation of LSC genes and signaling prooncogenic pathways such as Wnt and hedgehog.

Collectively, these data suggest that Gfi1b is a novel prognostic marker and tumor suppressor in AML.

Disclosure: No conflict of interest disclosed.

V35 - Targeting aberrant NCAM (neural cell adhesion molecule; CD56) expression in acute myeloid leukemia

Sasca D.1, Schüler A.1, Kriege O.1, Kunz K.1, Szybinski J.1, Fehr E.-M.1, Haehnel P.S.1, Gebhardt W.H.2, Reid G.2, Theobald M.1, Bullinger L.3, Kindler T.1

1University Medical Center of Mainz, III. Medical Department, Hematology, Oncology, Pneumology, Mainz, Germany, 2Institute of Molecular Biology, Mainz, Germany, 3University Hospital of Ulm, Department of Internal Medicine III, Ulm, Germany

Introduction: Acute myeloid leukemia (AML) is a heterogeneous disease of the hematopoietic progenitor cell driven by the subsequent acquisition of genetic alterations. Approximately 20% of AML patients show strong expression of CD56 (neural cell adhesion molecule; NCAM), but the functional consequences have not been investigated to date. This study aims to examine the biological role of NCAM in AML and explore whether NCAM represents a potential therapeutic target.

Methods: Leukemic cells were analyzed for proliferation, cell death, engraftment potential in xenografts and downstream signaling pathways after knock-down or overexpression of NCAM. Upstream regulatory mechanisms were investigated by DNaseI-hypersensitivity assays. To analyze NCAM effects at the stem cell level we used a well-established murine MLL-AF9 (MA9) leukemia model using C57BL/6 wild-type or NCAM-/- mice and performed serial replating and bone marrow (BM) transplantation experiments.

Results: NCAM was highly expressed in several cell lines and correlated with MA9+ leukemia. Knockdown of NCAM caused diminished proliferation, G1-arrest and finally cell death. Suppression of NCAM sensitized leukemic blasts to genotoxic stress in vitro and prolonged survival of xenografts, whereas exogenous overexpression in NCAM-negative cell lines caused an increased resistance to treatment. Applying DNaseI-hypersensitivity assays we demonstrate accessible binding sites for the transcription factors MEIS1, MEF2c and STAT1. shRNA-mediated knockdown of MEIS1, MEF2c and MLL-AF9 resulted in downregulation of NCAM expression, suggesting an upstream regulatory role for MLL-AF9. Analysis of downstream signaling pathways upon knockdown of NCAM revealed reduced expression levels of beta-Catenin and its downstream products but also activation of SMAD2/3, upregulation of p21 and downregulation of BCL2. To analyze NCAM expression in leukemia-initiating cells (LICs), we transduced wild-type and NCAM-/- derived BM cells with MA9 expressing retrovirus and transplanted syngeneic recipient mice. LICs (Lin-c-Kit+CD34+FcgR+) showed strong surface expression of NCAM, whereas normal HSCs (Lin-c-Kit+Sca1+) were NCAM-negative. Recipients of NCAM-/-MA9_cells developed leukemia with prolonged latency, and in colony assays replating was diminished.

Conclusions: Our data suggest that NCAM is involved in self-renewal and stress resistance of leukemic cells. Targeting of NCAM could become a promising therapeutic strategy in AML.

Disclosure: No conflict of interest disclosed.

V36 - Targeting of acute myeloid leukemia initiating cells with Fc optimized CD96 antibodies

Staudinger M.1, Peipp M.1, Kellner C.1, Bulduk M.1, Humpe A.1, Gramatzki M.1

1Universität Kiel, Sektion f. Stammzell- & Immuntherapie, 2. Med. Klinik, Kiel, Germany

Introduction: Acute myeloid leukemia (AML) is maintained by a population of leukemic stem cells (LSC). LSC residing in patients after intensive chemotherapy account for the high rate of relapse observed in AML patients. The development of novel approaches for efficient elimination of LSC is a prerequisite to improve AML therapy. Previously we demonstrated with our antibody TH-111 that CD96, a surface antigen identified on AML-LSC, represents a suitable target for ex vivo purging of autologous stem cell grafts. Since certain situations would require antibody application in the patient, here, a novel Fc optimized antibody against CD96 was designed to allow targeting of AML-LSC in vivo.

Methods: CD96 antibodies with Fc variants optimized for antibody-dependent cell-mediated cytotoxicity (ADCC) or complement-dependent cytotoxicity (CDC) were generated. Effector mechanisms triggered by the antibody variants were analyzed. NK cells from healthy donors were used as effector cells. Plasma served as source of complement. ADCC and CDC activity against cell lines and freshly isolated tumor cells was analyzed by flow cytometry or 51Cr assays. Colony forming assays were performed to analyze the cytotoxic potential of the novel CD96 antibody variants against hematopoietic progenitor cells (HPC).

Results: The variable regions of an affinity maturated scFv were used for generating full length CD96 antibodies with engineered IgG1-based Fc variants optimized for ADCC or CDC activity. ADCC was identified as the major effector mechanism. The ADCC-optimized CD96 antibody efficiently recruited NK cells to lyse KG1a myeloid blasts as well as primary AML tumor cells. The cytotoxic activity was compared to a similarly designed antibody directed against CD34, to secure that this ADCC-optimized CD96 antibody discriminates LSC from healthy HPC. Both, CD34+ HPC from healthy donors and CD34+/CD96+ KG1a cells were lysed by the CD34 antibody. In contrast, our CD96 antibody did not kill CD34+ HPC but efficiently eliminated KG1a. The lack of stem cell toxicity was supported by colony forming assays demonstrating that the capacity of HPC to proliferate and differentiate was not affected.

Conclusion: The particular features of Fc-optimized CD96 antibodies lacking stem cell cytotoxicity but eliminating AML-LSC may provide an additional therapeutic option for AML patients in certain clinical situations for instance in the setting of allogeneic stem cell transplantation.

Disclosure: No conflict of interest disclosed.

V37 - Targeted resequencing of MLL-PTD positive AML patients reveals a high prevalence of co-occurring mutations in epigenetic regulator genes

Herold S.1,2, Stange T.3, Kuhn M.3, Platzbecker U.1, Roeder I.3, Röllig C.1, Serve H.4, Berdel W.E.5, Bornhäuser M.1, Ehninger G.1, Thiede C.1

1Universitätsklinikum Dresden, Medizinische Klinik I, Dresden, Germany, 2Deutsches Krebsforschungszentrum, Deutsches Konsortium für Translationale Krebsforschung DKTK Dresden, Heidelberg, Germany, 3TU Dresden, Institut für Medizinische Informatik und Biometrie, Dresden, Germany, 4Universitätsklinikum Frankfurt, Medizinische Klinik II, Frankfurt, Germany, 5Medizinische Klinik A, Medizinische Fakultät Münster, Münster, Germany

Introduction: Partial tandem duplications within the MLL gene (MLL-PTD) are a recurrent molecular alteration in AML (up to 10%) and are associated with a poor prognosis. Little is known on the molecular profile and cooperating mutations in patients with MLL-PTD. In order to identify accompanying mutations, we performed whole exome sequencing (WES) of 8 MLL-PTD patients. Based on the observed alterations we then resequenced 118 MLL-PTD patients using either a custom targeted panel or a comprehensive commercial amplicon panel.

Methods: Patients included in this analysis were treated in prospective protocols of the Study Alliance Leukemia (SAL). We performed WES of FACS-sorted leukemic blasts and T-cells. For targeted resequencing, in total 1407 amplicons (custom panel) and 568 amplicons (TruSight Myeloid panel) were sequenced.

Results: WES of 8 MLL-PTD patients revealed a total number of 490 SNVs. Most frequently mutated genes were the epigenetic regulators DNMT3A, IDH1/2 and TET2. Confirmed somatic mutations were also found in genes less known to be mutated in AML, such as ATM, GNAS, TET1 and EP300. The targeted resequencing of 118 patients revealed a total number of 249 individual mutations, 16 genes were not found to be mutated, and in 8 patients, no co-occurring mutations were identified. Most frequently mutated genes in the entire MLL-PTD cohort were DNMT3A (33.9%), FLT3 (33.1%; 27.9% ITD; 5.0% TKD), IDH1 (22.9%), IDH2 (28.8%) and TET2 (19.5%) (Fig.1), whereas typical NPM1 mutations were rarely seen (2.4%). Especially mutations in IDH1, IDH2, and TET2 were found very frequently and about twice as common as in the general AML patient population.

Conclusions: Our analysis did not reveal any new and specific individual aberration present in patients with MLL-PTD mutations. However, our data indicate a very high prevalence of alterations in epigenetic regulator genes (close to 80%), which strongly argues for a particular disease biology in MLL-PTD patients and might also implicate that treatment based on demethylating agents or histone-deacetylase inhibitors might be especially attractive for patients with MLL-PTD.


Disclosure: Sylvia Herold: Expert Testimony: Illumina Christian Thiede: Stock Ownership: Ownership Agendix; Expert Testimony: Illumina.

Freier Vortrag

Multiples Myelom klinisch

V38 - Single versus tandem high-dose melphalan followed by autologous blood stem cell transplantation in newly-diagnosed multiple myeloma: final results from the GMMG-HD2 trial

Mai E.K.1, Benner A.2, Bertsch U.1, Schmidt-Wolf I.G.H.3, Hänel A.4, Kunzmann V.5, Naumann R.6, Neben K.7, Egerer G.1, Hillengass J.1, Neubauer A.8, Peyn A.9, Ko Y.-D.10, Peter N.11, Salwender H.J.12, Scheid C.13, Goldschmidt H.1,14, German-speaking Myeloma Multicenter Group (GMMG)

1University Clinic Heidelberg, Department of Internal Medicine V, Heidelberg, Germany, 2German Cancer Research Center (DKFZ), Division of Biostatistics, Heidelberg, Germany, 3University Clinic Bonn, Center for Integrated Oncology, Medizinische Klinik und Poliklinik III, Bonn, Germany, 4Klinikum Chemnitz gGmbH, Department of Internal Medicine III, Chemnitz, Germany, 5University Clinic Würzburg, Department of Internal Medicine II, Würzburg, Germany, 6Gemeinschaftsklinikum Mittelrhein, Department of Hematology and Oncology, Koblenz, Germany, 7Klinikum Baden Baden, Department of Hematology and Oncology, Baden Baden, Germany, 8University Clinic Giessen/Marburg, Department of Hematology and Oncology, Marburg, Germany, 9Klinikum Bremen Mitte, Department of Internal Medicine I, Bremen, Germany, 10Evangelische Kliniken Bonn, Department of Hematology and Oncology, Bonn, Germany, 11Carl-Thiem Klinikum Cottbus, Department of Internal Medicine II, Cottbus, Germany, 12Asklepios Hospital Hamburg Altona, Department of Hematology and Oncology, Hamburg, Germany, 13University Hospital Köln, Department of Internal Medicine I, Köln, Germany, 14Nationales Centrum für Tumorerkrankungen (NCT), Heidelberg, Germany

Introduction: The role of upfront single versus tandem high-dose melphalan (HDM) followed by autologous blood stem cell transplantation (ASCT) in multiple myeloma (MM) remains controversial. The open-label, randomized, multicenter phase III trial HD2 is the only prospective trial that applied the recommended HDM of 200mg/m2 prior to ASCT in the single and tandem group.

Methods: Between 12/1998 and 07/2002, 385 patients were randomized to receive either single (arm A) or tandem (arm B) HDM+ASCT after VAD/VID/VCAP-like induction therapy (IT). Main inclusion criteria were Salmon/Durie stage II/III, age ≥18 or <66 years (at initial diagnosis) and at least stable disease (SD) after IT. The primary analysis of the study compared event-free survival (EFS) two years after randomization.

Results: The intention-to-treat (ITT) population consisted of 177/181 (arm A/B) patients, respectively. In arm B, 47 patients refused second HDM+ASCT due to non-medical reasons. Non-inferiority of single HDM+ASCT versus tandem HDM+ASCT regarding 2-year EFS was demonstrated (ITT, HR = 0.93 with a one-sided confidence limit of 0.77). A per-protocol (PP) analysis including patients who received the assigned treatment (single/tandem HDM+ASCT, n = 156/n = 93 patients) revealed a non-significant trend towards prolonged EFS in the tandem HDM+ASCT group (p = 0.12). The CR rates before/after first HDM+ASCT were comparable in both groups. In arm B, CR rates significantly improved after second HDM+ASCT (ITT, after first/second HDM+ASCT: 15.1% vs. 19.4%, p = 0.04). Achieving a CR before and after first/second HDM+ASCT significantly improved EFS (p = 0.02) and OS (p = 0.03) in the whole study population. On long-term follow-up no OS differences were observed (ITT/PP: p = 0.33/0.90).

Conclusions: We observed a trend towards a prolonged EFS in arm B in the PP population, which did not translate into the ITT population. With a median follow-up of more than 11 years, no OS differences were observed. The high number of patients refusing second HDM+ASCT confound these findings. Tandem HDM+ASCT significantly increased CR rates, and achieving a CR remained an important prognostic factor to prolong EFS and OS in the whole study population. Therefore, tandem HDM+ASCT can constitute an important therapeutic approach to improve CR rates. Subgroups that benefit from tandem HDM+ASCT and the role of single versus tandem HDM+ASCT remain to be defined in the era of novel agents.

Disclosure: Elias Mai: Other Financial Relationships: Travel grants from Janssen-Cilag, Celgene, Onyx, and Mundipharma, outside the submitted work Hartmut Goldschmidt: Advisory Role: Personal fees and other support from Janssen-Cilag, Celgene, Novartis, Onyx, Millennium, Chugai, outside the submitted work; Financing of Scientific Research: Personal fees and other support from Janssen-Cilag, Celgene, Novartis, Onyx, Millennium, Chugai, outside the submitted work; Expert Testimony: Grants from Janssen-Cilag, Celgene, Novartis, Chugai, outside the submitted work.

V39 - IKZF1 expression is associated with outcome in lenalidomide treated newly diagnosed multiple myeloma

Kuchenbauer F.1, Krönke J.1, Kull M.1, Teleanu V.1, Bullinger L.1, Straka C.2, Mügge L.-O.3, Bassermann F.4, Engelhardt M.5, Bargou R.6, Einsele H.6, Knop S.6, Langer C.1, Deutsche Studiengruppe Multiples Myelom (DSMM)

1Universitätsklinik Ulm, Innere Medizin III, Ulm, Germany, 2Hematology and Oncology, Schön Klinik Starnberger See, Berg, Germany, 3Friedrich Schiller University, Internal Medicine II, Jena, Germany, 4Klinikum Rechts der Isar, Department of Medicine III, München, Germany, 5University of Freiburg, Hematology and Oncology, Freiburg, Germany, 6University Hospital of Würzburg, Department of Internal Medicine II, Würzburg, Germany

Introduction: Lenalidomide is an immunomodulatory drug (IMiD) with high activity in multiple myeloma (MM). IMiD binding to the CRBN E3 ubiquitin ligase results in targeted ubiquitination and degradation of the lymphoid transcription factors IKZF1 and IKZF3 and constitutes a unique mechanism of action for a class of drugs.

Methods: We analyzed mRNA expression levels of IKZF1, IKZF3, and CRBN by real-time quantitative PCR (RQ-PCR) in CD138-selected pre-treatment samples from 60 pts with newly diagnosed MM uniformly treated within a phase II clinical trial of the German Myeloma Study Group (DSMM). Additionally, all pts were characterized for the presence of cytogenetic abnormalities using a comprehensive set of FISH probes. All patients received induction consisting of four cycles of lenalidomide, adriamycin and dexamethasone followed by high-dose melphalan (Mel-200) with autologous stem cell transplantation (aSCT). Genetically defined low- and intermediate risk patients received a second aSCT, while those pts with high-risk cytogenetics (presence of a del17p13, t(4;14) or t(14;16)) underwent allogeneic stem cell transplantation if a donor was available. All patients received lenalidomide maintenance for 12 months.

Results: Median expression levels normalized to plasma cells from healthy volunteers of CRBN, IKZF1, and IKZF3 were 0.66 (range, 0.12-2.86), 0.66 (0.16-2.92), and 0.52 (0.19-5.13), respectively. Expression levels of CRBN, IKZF1, and IKZF3 were not affected by the presence of chromosomal aberrations (del13q14, del17p13, del9q34, +1q21, t(4;14), t(11;14), t(14;16)).

Patients achieving a complete response (CR) or very good partial response (VGPR) had a trend towards a lower IKZF1 expression than patients achieving a partial response (PR) (median IKZF1 expression: 0.62 vs 0.84, p = 0.07). Consistently, lower IKZF1 expression levels were associated with a better outcome: when segregating IKZF1 expression levels into quartiles, pts with the lowest (Q1) quartile of IKZF1 expression had a 3 year progression-free survival (PFS) of 86% compared to 51% in patients of the remaining quartiles (Q2-Q4)(Log rank test, p = 0.01). This translated into a better overall survival (OS) (IKZF1 Q1:100% vs IKZF1 Q2-4:74% after 3 years, log rank p = 0.03). In contrast, CRBN and IKZF3 expression levels had no impact on response, PFS, or OS.

Disclosure: Florian Kuchenbauer: No conflict of interest disclosed. Christian Langer: Advisory Role: Celgene, Janssen-Cilag, Bristol-Myers Squibb.

V40 - Comparative assessment of myeloma response to induction treatment in the GMMG MM5 study using IMWG criteria and hevylite assay

Scheid C.1, Hose D.2, Bertsch U.3, Hielscher T.4, Kunz C.4, Salwender H.5, Haenel M.6, Merz M.2, Mai E.K.2, Schurich B.2, Munder M.7, Schmidt-Wolf I.8, Gerecke C.9, Lindemann W.10, Zeis M.11, Weisel K.12, Dürig J.13, Jauch A.14, Peters-Regehr T.15, Zorn M.16, Goldschmidt H.3, GMMG

1Klinik I für Innere Medizin, Uniklinik, Köln, Germany, 2Medizinische Klinik V, Universitätsklinikum, Heidelberg, Germany, 3Sektion Multiples Myelom der Medizinischen Klinik V und des Nationalen Centrums für Tumorerkrankungen (NCT), Universität Heidelberg, Heidelberg, Germany, 4Biostatistics, German Cancer Research Center, Heidelberg, Germany, 5Asklepios Hospital Altona, Hamburg, Germany, 6Klinikum Chemnitz gGmbH, Chemnitz, Germany, 7University Medical Center, Mainz, Germany, 8Center for Integrated Oncology (CIO) and Medizinische Klinik und Poliklinik III, University of Bonn, Bonn, Germany, 9Hematology and Oncology, Helios-Hospital Berlin Buch, Berlin, Germany, 10Kath. Krankenhaus, Hagen, Germany, 11Department of Hematology and Stem Cell Transplantation, Asklepios Hospital St. Georg, Hamburg, Germany, 12Dept. of Hematology, Oncology and Immunology, University of Tuebingen, Tübingen, Germany, 13University Hospital, Essen, Germany, 14Institute of Human Genetics, University Hospital, Heidelberg, Germany, 15The binding site GmbH, Schwetzingen, Germany, 16Sektion Klinische Chemie, Universitätsklinikum, Heidelberg, Germany

Introduction: Treatment of multiple myeloma has substantially improved over the last decade. However response assessment still relies on parameters such as serum protein electropheresis or immunofixation. A new response category of stringent CR was introduced using information on serum free light chains and bone marrow clonality.

Recently the hevylite assay measuring kappa and lambda-restricted IgG and IgA was introduced. The objective of this study was to analyse the value of using the hevylite assay for assessing ealry MM response.

Methods: The MM5 study of the German GMMG randomised 604 newly diagnosed myeloma-patients to receive bortezomib, dexamethasone and either adriamycin or cyclophosphamide as induction before high-dose chemotherapy. Response after 3 cycles of induction was assessed using IMWG criteria. Clonal IgG and IgA was measured by hevylite in frozen serum samples from study baseline and after induction in 154 patients.

Results: 109 patients had IgG-paraprotein with 84 (77%) kappa and 25 (23%) lambda while 45 patients had IgA with 34 (76%) kappa and 11 (24%) lambda. Response to induction according to IMWG criteria was CR in 8 (5.2%), nCR in 19 (12.3%), VGPR in 27 (17.5%), PR in 71 (46.1%), MR in 19 (12.3%), SD in 7 (4.5%) and missing in 3 patients (1.9%). No patient had sCR based on free-light chain ratios. The frequencies of normal hevylite kappa/lambda ratios in the different IMWG response categories were as follows:

IgG-myeloma: CR :1/2 (50%), nCR: 2/4 (33.3%), VGPR: 1/20 (4.8%), PR: 3/52 (5.5%), MR: 0/16, SD: 1/4 (20%), missing: 0/2.

IgA- myeloma: CR: 4/6 (66.7%), nCR: 2/13 (15.4%), VGPR: 2/6 (33.3%), PR: 3/15 (20%), MR: 0/3, SD: 0/2, missing: 0/0.

Persistent pair suppression of non-clonal Ig was found in 85/109 (78%) for IgG and 30/46 (65%) for IgA.

Conclusion: Triplet induction achieved a CR/nCR rate of nearly 20%, while hevylite kappa/lambda normalisation occurred in 7.5% (IgG) and 24% (IgA). There was little overlap to IMWG response. This may be related to the fact that hevylite response not merely mirrors paraprotein response, but also captures suppression of the non-involved pair. It seems warranted to validate hevylite response after induction treatment with progression-free and overall survival once further follow-up data becomes available.

Disclosure: Christof Scheid: Financing of Scientific Research: binding site, Janssen, Celgene, Amgen Hartmut Goldschmidt: Expert Testimony: binding site, Janssen, Celgene, Bristol Myers Squibb.

V41 - Molecular profiling in patients with plasma cell myeloma treated with novel agents

Medinger M.1, Jakab A.1, Halter J.1, Heim D.1, Buser A.1, Gerull S.1, Stern M.1, Passweg J.1

1Universitätsspital Basel, Hämatologie, Basel, Switzerland

Introduction: Angiogenesis plays an important role in the pathogenesis and in progression of plasma cell myeloma (PCM). Novel agents like thalidomide, lenalidomide or bortezomib have in part anti-angiogenic properties. In this study we examined gene expression of angiogenic molecules in patients with PCM and correlated these markers to treatment response to novel agents.

Methods: We included 103 PCM patients treated with novel agents IMiDs (thalidomide or lenalidomide)-based regimens, (bortezomib (V)-based regimens or a combination of IMiD and bortezomib-based regimen (IMiD/V). Total RNA from peripheral blood mononuclear cells was isolated before anti-myeloma therapy and after 3 cycles. The mRNA levels of angiogenic molecules were measured using the Human Angiogenesis RT2 Profiler PCR Array. Changes in gene expression between responders and non-responders were analyzed as a fold increase/decrease. The response evaluation was performed after 3 cycles.

Results: Of the 103 patients included 32 patients were treated with an IMiD-based regimen, 34 with a V-based regimen and 37 with the combination (IMiD/V). The CR + VGPR rate was 43% in the IMiD-based group, 51% in the V-based group and 55% in the combination group (IMiD/V). Baseline gene expression of 11 from 84 genes tested correlated with advanced disease stage (p < 0.005 in each case). Regarding all 103 patients, gene expression of 15 from 84 genes tested (pre and post-treatment and changes in levels pre-treatment/post-treatment) were significantly different in responders compared to non-responders. Responders had a decreased expression proangiogenic factors (e.g., IL-6, VEGFA, ANGPT2, NRP1, FGFR, ECGF, and MMP-9) and increased expression of antiangiogenic factors (e.g., TIMP-3, TGF-β, and CXCL10). In the different treatment groups, the angiogenic gene levels (14/84) were significantly different in responders compared to non-responders in the IMiD-based group and the combination group after 3 cycles of therapy but not in the V-based group.

Conclusions: In the IMiD-containing therapy groups we found significant changes of angiogenic gene expression in responders compared to non-responders, whereas in the bortezomib-based group the difference in angiogenic gene expression was not significant. Possibly, anti-angiogenic properties play a higher role in IMID based treatment as compared to proteasome inhibitor based treatment.

Disclosure: No conflict of interest disclosed.

V42 - Vorinostat (V), bortezomib (B), doxorubicin (Dox) and dexamethasone (Dex, VBDD) in relapsed or refractory multiple myeloma patients (pts): results of an open, non-comparative, phase I/II investigator initiated trial (IIT)

Keller A.1,2, Waldschmidt J.M.1,2, Wider D.1, Jakobs D.1, Möller M.1, Reinhardt H.1, Pantic M.1, Grishina O.1, Ihorst G.3, May A.4, Frey A.4, Kohlweyer U.1, Jung M.5, Duyster J.1,2, Wäsch R.1,2, Engelhardt M.1,2

1Freiburg University Medical Center, Department of Hematology, Oncology and Stem Cell Transplantation, Freiburg, Germany, 2CCCF, Freiburg, Germany, 3Freiburg University Medical Center, Early Clinical Trials Unit (ECTU), Freiburg, Germany, 4Freiburg University Medical Center, Department of Pathology, Freiburg, Germany, 5Freiburg University, Institute of Pharmaceutical Sciences, Freiburg, Germany

Introduction: Vorinostat (V) was chosen as an epigenetic agent within this VBDD-regimen, since it has shown promising anti-tumor effects as a histone deacetylase inhibitor (HDACi). V inhibits the enzyme activity of HDAC1, 2, 3 and 6, allowing the activation of tumor suppressor genes, thereby slowing down cancer cells. Synergistic effects of V and B have been shown. V antagonizes escape mechanisms secondary to B-based therapies. Dox and Dex were chosen in this VBDD-regimen, because they are active and approved in relapsed and refractory (RR) MM. Since MM relapse leading to RRMM is common, novel treatment combinations are vitally needed. The aim of this phase I/II IIT was to test the tolerability and activity of the VBDD schedule within an out-pt regimen for RRMM pts.

Methods: V was escalated from 100mg (dose level 0), to 200mg (+1) and 300mg (+2). Primary objectives (MTD; 3+3 dose escalation), secondary objectives (safety, IMWG/EBMT responses, PFS, OS) and correlative endpoints (organ function, prognostic factors, QoL, comorbidity and HDAC-activity in PBMNCs/BM) were assessed throughout the trial. Dose limiting toxicities (DLTs) were defined as any possibly drug-related adverse event (AEs) ≥grade 3 (CTCAE) during the 1st cycle.

Results: 33 RRMM pts have been enrolled (median age 63 (47-78), KPS 90% (70-100%). The number of prior therapy lines was substantial with a median of 3 (1-8; with prior B, SCT and IMiDs in 88%, 94% and 42%, respectively). No DLTs have been observed. SAEs amounted to 15 and occurred in 9/33 pts (27%): amongst them 2 nonfatal SAEs judged to be related to the investigational therapy (1 bacteraemia, 1 herpes zoster reactivation). The response was rewarding with best ORR (>PR) and clinical benefit rate (CBR; >SD) of 62% and 97%, and end of treatment (EoT) ORREoT (>PR) and CBREoT (>SD) of 45% and 83%, respectively. Preliminary data propose that the response was independent of unfavorable cytogenetics. Comorbidity assessment assured no decline in pts´ condition. Ongoing pharmacodynamic analyses show HDAC downregulation after the 2nd cycle.

Conclusions: VBDD is an effective and well-tolerated out-pt regimen with promising response rates in heavily pretreated RRMM pts. The employed VBDD-regimen, with a continuous, rather than pulsed V-schedule, constitutes a promising treatment option, expands current standard therapies and, similarly to other HDACi, suggests V as a valuable add-on within this combination schedule.

* equal contribution AK + JW.

Disclosure: Alexander Keller: No conflict of interest disclosed. Monika Engelhardt: Financing of Scientific Research: Ja; Immaterial Conflict of Interests: Educational grants by MSD, Janssen-Cilag, Comprehensive Cancer Center Freiburg (CCCF) and Biotera fundation (ME)

V43 - Successful treatment of patients with newly diagnosed/untreated light chain multiple myeloma with a combination of bendamustine; prednisone and bortezomib (BPV)

Pönisch W.1, Mrachacz H.2, Khoder N.3, Plötze M.3, Holzvogt B.3, Andrea M.3, Schliwa T.3, Heyn S.3, Pfrepper C.4, Franke G.N.3, Krahl R.3, Jentzsch M.3, Leiblein S.3, Schwind S.3, Vucinic V.3, Niederwieser D.3, OSHO, Ostdeutsche Studiengruppe für Hämatologie und Onkologie

1Universitätsklinikum Leipzig, Hämatologie, internistische Onkologie, Leipzig, Germany, 2Universitätsklinikum Leipzig, Hämatologie, Leipzig, Germany, 3Universitätsklinikum, Leipzig, Germany, 4Diakonissenkrankenhaus, Leipzig, Germany

Background: Patients with light chain myeloma have frequently a light chain tubular cast nephropathy, which can lead to severe renal impairment.

Objective: Both bortezomib and bendamustine have been identified as quickly acting, effective and well tolerated drugs and might therefore constitute an adequate combination regimen for patients with newly diagnosed/untreated light chain multiple myeloma.

Methods: Between September 2009 and March 2014, 20 patients with newly diagnosed/untreated light chain multiple myeloma were treated with bendamustine 60 mg/qm on days 1 and 2, bortezomib 1.3 mg/qm on days 1, 4, 8 and 11, and prednisone 100 mg on days 1, 2, 4, 8 and 11 once every 21 days. 5 patients (25%) had a moderate or severe renal dysfunction (eGFR 15-59 ml/min) and 9 patients (45%) a renal failure/dialysis (eGFR < 15 ml/min).

Results: The median number of the BPV-treatment was 2 (1-5) cycles. 19 patients (95%) responded after at least one cycle of chemotherapy with 3 sCR, 4 nCR, 5 VGPR, and 7 PR. The myeloma light chains decreased rapidly, reaching the best response after the first cycle in 8 and after the second cycle in additional 9 patients.

16 patients discontinued therapy after median 2 cycles of BPV treatment to receive autologous (n = 13) or autologous/allogeneic SCT (n = 3). All together 10/14 patients with at least moderate renal failure improved their renal function (4 CRrenal, 2 PRrenal, 4 MRrenal). 3 of the 6 dialysis-dependent patients became dialysis-independent.

With a median follow up of 23 months of the surviving patients, median PFS and OS for patients at 24 months were 90% and 95%, respectively. The most common severe side effect was grade 3-4 leukocytopenia in 25% of the patients. Grade 3-4 thrombocytopenia was observed in 15% of the patients. Moderate to severe infection were seen in 4 patients.

Conclusion: We conclude that BPV is effective and well tolerated in patients with newly diagnosed/untreated light chain multiple myeloma.

Disclosure: No conflict of interest disclosed.

Freier Vortrag

Lungenkarzinom I

V44 - Tumor-infiltrating B lymphocytes characterized by CD79a and MUM1 independently predict outcome in patients with non-small cell lung cancer

Fischer R.N.1, Scheel A.H.2, Rothschild S.I.1,3,4, Schlößer H.A.3,5, Wolf J.1, Büttner R.2, Ansén S.1, von Bergwelt-Baildon M.S.3

1Uniklinik Köln, Centrum für Integrierte Onkologie (CIO), Lung Cancer Group Cologne, Köln, Germany, 2Uniklinik Köln, Institut für Pathologie, Köln, Germany, 3Uniklinik Köln, Klinik I für Innere Medizin, Cologne Interventional Immunology, Köln, Germany, 4Universitätsspital Basel, Medizinische Onkologie, Basel, Switzerland, 5Uniklinik Köln, Klinik und Poliklinik für Allgemein-, Viszeral- und Tumorchirurgie, Köln, Germany

Introduction: Tumor-infiltrating lymphocytes play an important role in cell-mediated immune-destruction of cancer cells and tumor growth control. T-cell infiltrates are robust prognostic biomarkers in a variety of solid tumors. In non-small cell lung cancer (NSCLC) a prognostic role of T cell subtypes, natural killer cells and dendritic cells within the tumor stroma has been described. Here, we studied the role of tumor-infiltrating B cells characterized by CD79a (B-cell antigen receptor complex-associated protein alpha chain) and MUM1 surface expression (Multiple myeloma oncogene 1) in patients with NSCLC. To our knowledge, this study represents the so far largest cohort analyzing the prognostic impact of tumor-infiltrating B cells in NSCLC.

Methods: B cell infiltration was quantified using immunohistochemistry and antibodies to CD79a (Dako, clone JCB117) and MUM1 (Dako, clone MUM1p) on tissue microarrays of paraffin embedded tumor sections. Genetic driver mutations were identified by next-generation sequencing and FISH analysis.

Results: 478 tissue samples from NSCLC patients were available for final analysis. Median age of the patient population was 66 years with 65% male patients. 56% had adenocarcinoma and 39% squamous cell histology. 65% of patients were diagnosed with localized disease (stage I: 44%; stage II: 21%), 30% locally advanced disease (stage III) and 6% were diagnosed with stage IV. CD79a and MUM1 positive cells were detected in 40.8% (195/478) and 40.2% (192/478) of the analyzed NSCLC tissue samples, respectively. B cell infiltration was not associated with clinical or histo-pathological characteristics.

CD79a expression was associated with a trend towards a better outcome (median OS 49 vs. 40 months, p = 0.069). The expression of MUM1 showed a significantly prolonged overall survival (median OS 54 vs. 40 months, p = 0.025). In the multivariate analysis B cell infiltration characterized by positivity for CD79a and MUM1 was an independent prognostic marker for survival (p = 0.045) as was MUM1 expression alone (p = 0.031).

Conclusions: In this cohort of patients with localized and locally advanced stage NSCLC, B cell infiltration characterized by immunohistochemical positivity for CD79a and MUM1 represents an independent prognostic marker. This finding supports the hypothesis of a B cell-mediated anti-tumor immunity.

Disclosure: No conflict of interest disclosed.

V45 - Do we miss changes in quality of life? Data from the LuLife project in patients with non-small cell lung cancer in Germany

von Verschuer U.1, Sandner R.2, Däßler K.-U.3, Tessen H.W.4, Münz M.5, Spring L.5, Jänicke M.5, Marschner N.6

1Hämato-Onkologische Gemeinschaftspraxis, Essen, Germany, 2Passauer Onkologische Praxis Dres. Prenninger & Sandner, Passau, Germany, 3Onkologische Schwerpunktpraxis, Freital, Germany, 4Onkologische Schwerpunktpraxis, Goslar, Germany, 5iOMEDICO, Freiburg, Germany, 6Praxis für interdisziplinäre Onkologie & Hämatologie, Freiburg, Germany

Introduction: Most patients (pts) with lung cancer are diagnosed with advanced disease and receive palliative chemotherapy. Both tumor burden and chemotherapy can strongly affect quality of life (QoL) but little is known about how QoL changes over time in pts in routine care. The LuLife project prospectively investigated qol and other patient reported outcomes (PROs) in real-life pts.

Methods: LuLife was conducted with the tumor registry on lung cancer (TLK), which prospectively collects data on routine treatment of lung cancer as administered by office-based medical oncologists in Germany and was started in 2010. In LuLife the validated questionnaires EORTC QLQ-C30 (global QoL) and -LC13 (lung cancer module) were sent to pts with NSCLC at start of 1st-line treatment and then every eight weeks up to six times, and filled at home. Here, data on global as well as cancer- and symptom-related QoL over time are presented.

Results: 507 pts answered at least one questionnaire. They had a mean age of 66 years at the start of 1st-line treatment. ECOG status was 0/1/≥2 for 30/56/14% of pts. Most pts (>80%) received platin based chemotherapy. Questionnaires were answered by 485/424/353/290/240/194 pts at 0/8/16/24/32/40 weeks which represents a return rate of 96/87/81/76/73/65% of all pts alive at the respective time points. In total 38% of the pts returned the last questionnaire. Overall, the better the PRO values at baseline, the more questionnaires were returned. Analysis of mean difference to baseline revealed no mean change in global QoL, pain, fatigue and emotional function over time, but small to medium clinically relevant mean deterioration in social, physical, role and cognitive function. Alopecia strongly increased within the first two months, while dyspnoe (small change) and neuropathic symptoms (large change) continuously worsened over time. Despite little mean changes from baseline in the whole population, analysis of the time to deterioration revealed that, after 16 weeks 30% of all pts had reported clinically relevant decreased QoL (individual deterioration of at least 10% of the score) and worsened pain, but far more worsened fatigue (50%) or dyspnoe (40%).

Conclusion: Our data show that PROs of NSCLC pts change over time but results, especially on global QoLQoL, differ depending on the method of choice. It is crucial to select appropriate analyses to avoid missing changes in PROs, which could indicate a need to improve patient care.

Disclosure: No conflict of interest disclosed.

V46 - Randomized study of individualized, pharmacokinetically-guided dosing of paclitaxel combined with carboplatin in advanced non-small cell lung cancer patients

Joerger M.1, Salamone S.J.2, von Pawel J.3, Kraff S.4, Fischer J.R.5, Eberhardt W.E.6, Gauler T.C.7, Müller L.8, Reinmuth N.9, Reck M.9, Kimmich M.10, Mayer F.11, Kopp H.-G.11, Behringer D.M.12, Ko Y.-D.13, Früh M.1, Hilger R.A.7, Roessler M.14, Moritz B.14, Miller M.C.2, Jaehde U.4, CESAR Central European Society for Anticancer Drug Research-EWIV

1Cantonal Hospital, St. Gallen, Switzerland, 2Saladax Biomedical Inc., Bethlehem, United States, 3Asklepios Fachkliniken, Klinik für Pneumologie, Gauting, Germany, 4Institute of Pharmacy, Clinical Pharmacy, University, Bonn, Germany, 5Klinik, Löwenstein, Germany, 6West German Cancer Center, Ruhrlandklinik, Essen, Germany, 7Universitätsklinikum, Essen, Germany, 8Onkologische Schwerpunktpraxis, Leer, Germany, 9LungenClinic, Großhansdorf, Germany, 10Klinik Schillerhöhe, Gerlingen, Germany, 11Universitätsklinikum, Tübingen, Germany, 12Augusta-Kranken-Anstalt, Bochum, Germany, 13Johanniter Krankenhaus, Bonn, Germany, 14CESAR Central Office, Wien, Austria

Introduction: Variability of chemotherapy exposure may cause severe toxicity or lack of efficacy. Paclitaxel (PTX) exposure (time above a plasma concentration of 0.05 mM, Tc > 0.05) has been shown to predict toxicity. Whereas carboplatin dosing is adapted to kidney function, PTX dosing only accounts for body-surface area. We developed a PTX dosing algorithm for avoidance of supra- or subtherapeutic PTX exposure based on Tc>0.05 determined from a single blood sample drawn 18-30 hours after starting PTX infusion. This study was initiated to validate the use of a pharmacokinetically (PK)-guided dosing strategy for PTX to optimize exposure in patients with advanced non-small cell lung cancer (NSCLC).

Methods: 304 advanced NSCLC patients were randomly assigned to receive up to 6 cycles of first-line 3-weekly carboplatin AUC6 combined with PTX either at a standard dose of 200 mg/m2 (Arm A) or at a PK-guided dose (Arm B). Initial PTX dose in Arm B was between 150 to 200 mg/m2 based on age and sex. PTX exposure was determined from a single blood sample drawn 18-30 hr after infusion initiation using an HPLC-UV assay. PTX doses were adjusted according to the previous cycle exposure to target a Tc>0.05 between 26 and 31 hours using our dosing algorithm. Primary objective of the study was to detect an 11% reduction of grade 4 neutropenia with PK-guided PTX dosing. Secondary objectives included comparison of neuropathy rates, tumor response and progression-free survival (PFS) between the study arms.

Results: Compared to standard dosing, PK-guided dosing of PTX reduced the incidence of grade 4 neutropenia (measured on day 15 of each cycle) (21% vs. 15%, P = 0.029), grade ≥2 neuropathy (27% vs. 14%, P < 0.001), and grade ≥3 neuropathy (8% vs. 1%, P < 0.001). PK-guided dosing of PTX led to a dose reduction in 62% and a dose increase in 17% of the patients. The proportion above the target exposure was reduced from 41% in cycle 1 to 2% in cycle 6. Objective response rate between Arms A and B was similar (32% vs. 29%, P = 0.70), as was the median PFS (5.2 vs. 4.7 months, hazard ratio 1.1, 95% CI 0.8-1.4, P = 0.54).

Conclusion: PK-guided dosing of PTX improved the risk-benefit profile in patients with advanced NSCLC. The dosing algorithim resulted in reduction of grade 4 neutropenia and neuropathy ≥ grade 3, without reducing efficacy.

Disclosure: No conflict of interest disclosed.

V47 - P53 disruptive mutation is a negative predictive factor in EGFR M+ NSCLC treated with TKI

Lüers A.C.1,2, Neemann N.1, Prenzel R.3, Scriba D.C.4, Willborn K.C.5, Stropiep U.2, Falk M.6, Hallas C.6, Tiemann M.6, Griesinger F.1,2

1Pius-Hospital, University Department Internal Medicine-Oncology, Oldenburg, Germany, 2Pius-Hospital, Hematology and Oncology, Oldenburg, Germany, 3Pius-Hospital, Pneumology, Oldenburg, Germany, 4Pius-Hospital, Thoracic Surgery, Oldenburg, Germany, 5Pius-Hospital, Radiotherapy, Oldenburg, Germany, 6Hematopathology, Hamburg, Germany

Introduction: p53 mutations are common in lung cancer, and have also been described in EGFR mutated patients. The impact of p53 mutations in EGFR M+ patients is controversial, especially if classified as “disruptive” and “non-disruptive” according to their functional effect on the p53 protein as proposed by Poeta and colleagues. The aim of the study was therefore to systematically analyze EGFR and p53 mutations within a cohort of patients with lung cancer stage IV (UICC 7), to correlate alterations with clinical characteristics and to investigate a potential impact of p53 mutations on treatment outcome.

Methods: 267 patients from a single center diagnosed with lung cancer stage IV were studied for the presence of EGFR as well as inactivating p53 mutations. Methods for the detection of EGFR mutations included Sanger Sequencing and hybridization based COBAS testing. P53 mutations were detected by Sanger Sequencing. Clinical characteristics including smoking status were available for all patients.

Results: 267 consecutive patients at the lung cancer center of the Pius-Hospital Oldenburg were studied. The overall EGFR mutation rate was 19% (51/267) in all patients, 80% (41/51) showing common mutations of exon 19 or 21. P53 disruptive mutation showed in 16% (8/51) and p53 nondisruptive mutation occurred in 11% (22/51) whereas p53 WT was found in 47% (24/51). In 8/51 (16%) patients p53 analysis was not successful. OS was 37 months in p53 disruptive mutation and p53 WT patients compared to 19 months in p53 nondisruptive mutation (p < 0,05). PFS on 1st line TKI therapy was 18 months in p53 nondisruptive mutation and p53 WT patients and 6 months in p53 disruptive mutation (p < 0,024). Similar results could be shown in the EGFR common mutation subgroup but not in the uncommon mutation subgroup.

Conclusion: Significant differences in PFS and OS in EGFR M+ patients were observed depending on p53 mutation status. P53 mutational status is only predictive when disruptive and non-disruptive P53 mutations are differentiated. P53 should be tested prospectively in EGFR M+ patients as management of patients on 1st line TKI may be different.

Disclosure: Anne Lüers: Advisory Role: Boehringer Ingelheim; Expert Testimony: Astra Zeneca Frank Griesinger: Advisory Role: Boehringer Ingelheim, Roche, Pfizer, Astra, Novartis, Lilly, MSD, BMS, Clovis; Expert Testimony: Astra Zeneca.

V48 - Evaluation of disease-related symptoms in patients (pts) with advanced squamous (SQ) non-small cell lung cancer (NSCLC) treated with nivolumab (NIVO) or docetaxel (DOC)

Gralla R.J.1, Coon C.2, Taylor F.2, Penrod J.R.3, DeRosa M.2, Dastani H.3, Orsini L.3, Reck M.4

1Albert Einstein College of Medicine, Bronx, United States, 2Adelphi Values, Boston, United States, 3Bristol-Myers Squibb, Princeton, United States, 4Lungenklinik Großhansdorf, Airway Research Center North (ARCN), German Center for Lung Research (DZL), Großhansdorf, Germany

Background: CheckMate 017 (NCT01642004), a randomized, open-label, global phase 3 study, evaluated efficacy and safety of 2nd-line NIVO vs DOC in advanced SQ NSCLC pts. Overall survival was significantly superior and duration of treatment longer for NIVO vs DOC. Disease-related symptoms were also evaluated using the Lung Cancer Symptom Scale (LCSS), with scores reported from 0 (least severe) to 100 (most severe).

Methods: Assessment was performed every 4 weeks (Q4W) for NIVO and Q3W for DOC for the first 6 mos on treatment, then Q6W for the rest of the treatment period (both arms), and at follow-up visits 1 and 2 (post discontinuation). At each assessment, mean baseline (BL) and mean change from BL of the LCSS average symptom burden index (ASBI) were summarized by arm. Secondary endpoint was proportion of pts with clinically meaningful improvement in LCSS ASBI by wk 12 (≥10 point decrease, the minimally important difference [MID]) based on all randomized pts.

Results: Pt BL characteristics were generally balanced across treatment arms. LCSS completion rates at BL and ≥1 subsequent assessment were 68.9% for NIVO and 62.8% for DOC. Completion rates remained relatively consistent throughout assessments and by treatment arm. BL LCSS ASBI values were similar for NIVO (29.6; SD 16.4) and DOC (29.6; SD 14.7). By wk 12, 20.0% (27/135; 95% CI 13.6, 27.7) of NIVO vs 21.9% (30/137; 95% CI 15.3, 29.8) of DOC pts demonstrated clinically meaningful symptom improvement. The NIVO arm had statistically significant mean improvements from BL at each LCSS ASBI assessment from wks 12-54, after which sample sizes dropped to < 10 pts; mean improvement from wks 40-54 exceeded MID. DOC pts remaining on treatment had no statistically significant changes in LCSS ASBI through wk 18, after which the sample dropped to < 10 pts. At follow-up visits 1 and 2, mean LCSS ASBI for both NIVO and DOC pts indicated similar worsening of symptoms relative to BL (range, 5.5-9.5); for DOC pts, differences from BL were statistically significant.

Conclusion: The proportion of pts with meaningful symptom improvement by wk 12 was similar for both NIVO and DOC, but overall average symptom burden on NIVO improved from BL over most of the available follow-up period. Average symptom burden for DOC pts remained stable relative to BL during their shorter time on treatment. The results show statistically and clinically significant symptom reductions from BL for 2nd-line SQ NSCLC pts treated with NIVO.

Disclosure: Richard Gralla: Advisory Role: Bristol-Myers Squibb Martin Reck: Advisory Role: Hoffman La-Roche, Lilly, Merck Sharpe & Dohme, Bristol-Myers Squibb, AstraZeneca, Boehringer-Ingelheim, Pfizer, Novartis.

V49 - Clinical and genetic presentation of high-level cMET-amplified Non-small cell lung cancer (NSCLC) with adenocarcinoma histology

Eisert A.1, Scheffler M.1, Gogl L.1, Frank R.1, Scheel A.1, Merkelbach-Bruse S.1, Michels S.1, Fischer R.1, Büttner R.1, Wolf J.1

1Uniklinik, Köln, Germany

Introduction: cMET (MNNG HOS Transforming gene) is a proto-oncogene encoding the hepatocyte growth factor receptor (HGFR) with HGF as the only known ligand. Whereas the role of cMET mutations in NSCLC patients remains debated controversively, amplification of cMET is considered as one of the molecular resistance mechanisms in EGFR-mutated NSCLCs treated with 1st or 2nd generation EGFR-TKIs. Further, at least for high-level amplifications, these aberrations might be a possible target for cMET-targeted therapy also in EGFR-wt patients.

Patients and methods: Beside the screening of rebiopsied EGFR-TKI-resistant patients, cMET amplification status as assessed by fluorescence in-situ hybridization (FISH) was also analyzed in all presenting NSCLC patients with adenocarcinoma histology, regardless of therapy line or stage. A cohort of 1599 patients within a regional screening network was analyzed using FISH and next-generation sequencing (NGS). High-level amplifications were specified by the following criteria: cMET/CEN7 ratio of ≥ 2 or an average cMET gene copy number per cell of ≥ 6 or ≥ 10% of tumor cells containing ≥ cMET signals (according to Schildhaus et al., Clinical Cancer Research 2014).

Results: In a time period of one year 57 patients with high-level cMET amplification were identified: 35 patients (61.4%) were male and 22 patients (38.6%) were female. Of 41 patients analyzed for stage so far, 27 (66.9%) had metastatic disease (stage IV). cMET amplifications co-occurred with other driver mutations, especially EGFR (n = 7, 12.3%) and KRAS (n = 18, 31.6%). In 24 patients (42.1%) mutations within TP53 without further oncogene mutation could be detected. In 8 patients (14.0%) no other additional genetic aberrations were found.

Conclusion: High-level cMET amplification can occur with and without additional genetic lesions. About one third of high-level MET-amplified patients had a co-occurring KRAS mutation. In order to detect patients with high-level cMET amplification, it is not sufficient to concentrate on EGFR-mutated patients in the resistance setting. Further work is needed if these findings translate into clinical consequence.

Disclosure: Anna Eisert: No conflict of interest disclosed. Jürgen Wolf: Advisory Role: Advisory boards (compensated) and lecture fees: AstraZeneca, BMS, Boehringer-Ingelheim, Clovis, Novartis, Pfizer, Roche; Expert Testimony: Research support: Bayer, Boehringer-Ingelheim, Novartis, Pfizer, Roche.

Wissenschaftliches Symposium

Fortgeschrittenes Prostatakarzinom

V52 - Biomarker in mCRPC

Machtens S.1

1Marienkrankenhaus, Zentrum für Innere Medizin, Urologie und Kinderurologie, Leverkusen, Germany

Advances in the therapy of metastatic castration-refractory prostate cancer make an individual approach neccessary. This individualised decision can be supported by new biomarker. These biomarker can have an prognostic, predictive or monitoring character.

An overview is delivered in regard to current developments in the evaluation of new biomarkers and their possible influence on Treatment stratifications.

New aspects concerning the splice variants of the androgen receptor, gene fusions and others are highlighted.

Disclosure: Stefan Machtens: Financing of Scientific Research: Amgen, Sanofi Aventis, Novartis, Astellas, Janssen.


Supportivtherapie bei neuen systemischen Therapieformen

V66 - Endocrine side effects of anticancer drugs

Capraro J.1

1Kantonsspital Aarau, Endokrinologie, Diabetologie, Metabolismus, Aarau, Switzerland

Several of the newer anticancer drugs have been associated with distinct endocrine adverse effects. Regular screening and a high level of suspicion may help to prevent significant morbidity.

Immunotherapies against immune checkpoints that inhibit T cell activation (cytotoxic T

lymphocyte antigen 4 (CTLA-4)) and programmed cell death 1 (PD-1) have been found to adversely affect several endocrine tissues with hypopituitarism and adrenal failure being the most severe sequelae. Thyroid dysfunction may be another important associated condition. Central hypothyroidism is a well-recognized side effect of bexarotene. Older immunomodulating drugs, such as interferon-a and interleukin-2 are known to induce variably high incidence of autoimmune thyroid dysfunction. Tyrosine Kinase Inhibitors (TKI's) often induce thyroid disorders and regular testing of thyroid function is mandatory.

Therapy with mTOR inhibitors may result in severe dyslipidemia and diabetes mellitus. TKI's on the other hand may be associated with improved glucose metabolism in patients with diabetes, or with frank hypoglycemia. Abiraterone acetate, through its distinct effect on steroid metabolism, leads to adrenal mineralocorticoid excess which must be prevented by concomitant glucocorticoid administration.

Oncologists must be aware of these side effects in order to provide regular screening and initiate a specific diagnostic workup if suspicious clinical signs are present. If hormonal changes are detected early referral to an endocrinologist and multidisciplinary management are key to successful therapy.

Disclosure: No conflict of interest disclosed.


Nierenzellkarzinom - neue Aspekte und Standards

V67 - Management of side effects and adherence to oral cancer therapies in metastatic renal cell cancer

Rothermundt C.1

1Kantonsspital St. Gallen, Onkologie/Hämatologie, St. Gallen, Switzerland

Owing to targeted therapies and sequencing treatments, the life expectancy of patients with metastatic renal cell cancer (mRCC) has been extended from 13 months in the cytokine era (Motzer 2002) to over 30 months (Motzer 2014) today. All available drugs for mRCC are long-term therapies with continuous or intermittent dosing, which are given until disease progression or intolerable toxicity occurs. Thus patients spend long times “on treatment” (Negrier 2012). Retrospective data show that only 52%-79% of mRCC patients receive second-line treatment, depending on the choice of first-line treatment (Levy 2013), and 13% of patients receive third-line treatment (Iacovelli 2013). Tolerability of therapies and side effect management is therefore of importance in mRCC. Maintenance of an effective dose is a concern in patients treated with oral targeted therapies, in order to achieve optimal results. Decisions on dose reductions and interruptions should always be made knowing about potential concessions on efficacy.

There are several comprehensive reviews on side effects and side effect management in patients on targeted therapies for mRCC (Lacouture 2008, Kollmannsberger 2011, Eisen 2012, Mendez-Vidal 2012, Larkin 2014), concerning e.g. cardiovascular, cutaneous, constitutional, endocrine, gastrointestinal, laboratory and organ function toxicity.

Progress in cancer treatment has been accompanied by a paradigm shift in medication delivery: a shift in treatment away from the outpatient clinic and into the patient's home. There has been little emphasis on making sure patients take medications correctly once prescribed—the final step in the bench to patient pathway. However, the success of oncologic treatment depends on the patient's adherence to medication regimens (Gellad 2014). Factors associated with nonadherence include treatment regimen, the patient, social environment, and the clinician-patient interaction. Patient perceptions and motivations appear to be the most important determinants. Prediction of nonadherence is especially challenging. Interventions that have produced some improvement in adherence among oncologic patients include educational programs; behavioral modification techniques, and use of reminder systems and cues (Partridge 2002).

In Switzerland an educational program for oncology nurses and medical oncologists is underway to raise awareness for adherence, to establish adherence assessments and to plan interventions aimed at improving adherence.

Disclosure: Christian Rothermundt: Advisory Role: Pfizer; Financing of Scientific Research: GSK, Novartis.

V68 - Checkpoint inhibitors - a new road to salvation in mRCC?

Grünwald V.1

1MHH, Hannover, Germany

Checkpoint inhibitors have enriched the armentarium to treat cancer in recent years. Its implementation as the fifth cornerstone of cancer treatment is still novel and we will have to explore its role in the field in more detail in the upcoming years in order to understand its full scope. It is the start of a new era, with many research questions ahead.

Inhibitors of the programmed death receptor 1 (PD-1) or its ligand (PD-L1) have unleashed the T-cell response, through enhancing the cytotoxic T-cell response. Early clinical studies in metastatic renal cell carcinoma (mRCC) confirmed an objective response rate (ORR) of 20-22%, which warrants further drug development in this disease. More interestingly, PD-1 inhibition achieved a promising overall survival (OS) of 18-25 mo., which raises expectations in previously treated patients. Therefore a current phase III study tested PD-1 inhibition against everolimus in later lines of therapy, and results are awaited later this year.

CTLA4 is another T-cell checkpoint in control of the T-cell priming phase. The combination of CTLA4 and PD-1 inhibition is thought to enhance T-cell priming, while boosting cytotoxic T-cell response. Early clinical studies suggested an increase in ORR to 43-48% and render this approach a candidate for future drug development whether as adjuvant or aggressive palliative treatment with the aim to improve survival. However, much more checkpoints are known to control T-cell fate, and novel agents are being developed to target them. Results are eagerly awaited and may forge a complete new landscape in mRCC, were improvement in OS may be reached. It is a very encouraging time in oncology, with great expectations ahead of us.

Disclosure: Viktor Grünwald: Advisory Role: BMS, MSD, Pfizer, Novartis, GSK, Bayer; Financing of Scientific Research: BMS, Pfizer, Novartis, GSK.

Freier Vortrag

Lymphome aggressiv klinisch I

V71 - MRD eradication should be the therapeutic goal in mantle cell lymphoma and may enable tailored treatment approaches: Results of the intergroup trials of the European MCL Network

Pott C.1, Macintyre E.2, Delfau-Larue M.-H.3, Ribrag V.4, Unterhalt M.5, Kneba M.6, Hiddemann W.5, Hermine O.7, Dreyling M.5, Hoster E.5, EU-MCL study group

1University Hospital Schleswig-Holstein, Second Department of Medicine, Kiel, Germany, 2Laboratoire d'Hématologie, Hôpital Necker-Enfants-Malades, Paris, France, 3Institut Mondor de Récherche Biomédicale (IMRB), Département d'Immunologie, Dérmatologie et Oncologie, Créteil, France, 4Laboratoire de récherche translationelle, Institut Gustave Roussy, Villejuif, France, 5Department of Internal Medicine III, University of Munich, Großhadern, Germany, 6Second Medical Department, University Hospital Schleswig-Holstein, Campus Kiel, Kiel, Germany, 7Department of Clinical Hematology, University Paris Descartes, Necker Hospital, Paris, France

Introduction: Minimal residual disease (MRD) for response assessment allows prediction of the clinical course and prognosis prior to clinical study endpoints in B-cell malignancies. Based on the randomized intergroup trials of the EU-MCL Network for stage II-IV mantle cell lymphoma (MCL) investigating immuno-chemotherapy followed by autologous stem cell transplantation (ASCT) for patients <65 yrs and rituximab or interferon maintenance for patients >65 yrs, we prospectively monitored MRD.

Methods: MRD was analysed in peripheral blood (PB) after induction and in 3-monthly intervals after ASCT or 2-monthly during maintenance until progression. Patients in complete or partial remission 6 months after ASCT/end of induction (MCL Younger/MCL Elderly) were included. MRD status was defined as negative, if there were only negative MRD values, or positive in case of at least one positive MRD value during a 6-months period. Landmark analyses were performed for progression-free (PFS) and overall survival (OS) according to MRD status for each 6-months period until 4 years of follow-up.

Results: Among patients in remission 6 months after ASCT/end of induction, MRD results during follow-up were available in 406 patients (255 MCL Younger,151 MCL Elderly). The distribution into low, intermediate, and high risk MIPI of 44%, 34% and 22%, respectively, was similar to the total study population. During follow-up, about 20-25% of patients were MRD+ during the first 3 years, and less than 20% in the subsequent follow-up. In each landmark analysis, a positive MRD status was highly associated with a shorter PFS and OS. This association was independent of baseline MIPI score, treatment arm or protocol.

Thus, the prognostic value of MRD positivity was maintained in both trials MCL Younger and MCL Elderly. Remarkably, landmark analyses at all time points indicated a strong association also of low-level MRD with a shortened PFS. Finally, MRD analyses in 76 patients at relapse confirmed the close association of clinical relapse to MRD positivity, with only 10 patients (13%) being PB-negative. MRD reappearance preceeded clinical relapse in median 18 months.

Conclusion: Our data demonstrate that achievement and preservation of MRD response is the strongest independent predictor of prognosis in patients with MCL. Applying MRD status as tool for tailored treatment, therapeutic approaches should focus on a maximum MRD response to improve long term outcome.

Disclosure: No conflict of interest disclosed.

V72 - Acute hepatitis E infection in a young adult patient receiving rituximab for Burkitt lymphoma

Matschke J.1, Alashkar F.1, Gerken G.2, Dührsen U.1, Jochum C.2

1Department of Hematology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany, 2Department of Gastroenterology and Hepatology, University Hospital, University Duisburg-Essen, Essen, Germany

Introduction: Monoclonal antibodies like anti-CD20/rituximab have a high relevance for the therapy of B-cell-lymphoma. The risk for reactivation of hepatitis B was often described. Hepatitis E virus infections are rare and mostly self-limiting. Acute hepatitis E infections during a rituximab containing chemotherapy are not well described so far.

Case presentation: A 21 year old patient was diagnosed in November 2013 with Burkitt lymphoma. The chemotherapy regimen was chosen according to the GMALL-B-ALL/NHL 2002 study of the German Multicenter Study Group for Adult Acute Lymphoblastic Leukemia (GMALL) and was administered between November 2013 and June 2014. Three different courses (A,B,C) of chemotherapy were given in a 3-week interval and each was once repeated. Rituximab was given with the standard dose of 375 mg/m2 at the beginning of each course and with two additive cycles at the end of the six courses for a total of 8 administrations. The chemotherapy was well tolerated without major complications. After the fifth course of chemotherapy increasing transaminases were noticed (AST 109 U/L, ALT 123 U/L), which increased in the following two months (AST 414 U/L, ALT 739 U/L). Bilirubin concentrations were normal. A duplex sonography of the liver was without pathologic findings. Serological diagnostics excluded hepatitis A-C,E and cytomegalovirus. Two months after first observation of elevated transaminases and after the 8th administration of Rituximab a HEV viral load was detected using PCR. The patient developed a chronic hepatitis E during the following months. Seven months after the last course of rituximab B-cells reconstituted (CD19+-cells: 150/nl). Two weeks after reconstitution of B-cells hepatitis E PCR in blood and stool was negative and transaminases were normalized.

Conclusion: Patients with unknown acute or chronic hepatitis should be tested for hepatitis E. Acute hepatitis E infections could become chronically during immunosuppression, especially after administration of rituximab. B-cell reconstitution is important for curing hepatitis E.

Disclosure: No conflict of interest disclosed.

V73 - Long-term safety and efficacy of single-agent Ibrutinib in patients with relapsed or refractory mantle cell lymphoma: Updated results of an international, multicenter, open-label phase 2 study

Dreyling M.1, Wang M.L.2, Rule S.3, Martin P.4, Goy A.5, Auer R.6, Kahl B.S.7, Jurczak W.8, Advani R.H.9, Romaguera J.E.2, Williams M.E.10, Barrientos J.C.11, Chmielowska E.12, Radford J.13, Stilgenbauer S.14, Jedrzejczak W.W.15, Johnson P.16, Spurgeon S.E.17, Zhang L.2, Baher L.18, Cheng M.18, Beaupre D.M.18, Blum K.A.19

1University Hospital LMU, Munich, Germany, 2The University of Texas MD Anderson Cancer Center, Houston, United States, 3Plymouth University Peninsula Schools of Medicine and Dentistry, Plymouth, United Kingdom, 4Weill Cornell Medical College, New York, United States, 5John Theurer Cancer Center at Hackensack University Medical Center, Hackensack, United States, 6Barts Health National Health Service (NHS) Trust, London, United Kingdom, 7University of Wisconsin School of Medicine and Public Health, Madison, United States, 8Jagiellonian University, Krakow, Poland, 9Stanford University School of Medical College, New York, United States, 10University of Virginia School of Medicine, Charlottesville, United States, 11Hofstra North Shore-Long Island Jewish School of Medicine, Hempstead, United States, 12Oddzial Kliniczny Onkologii, Centrum Onkologii, Bydgoszcz, Poland, 13The University of Manchester and the Christie NHS Foundation Trust, Manchester, United Kingdom, 14Universitatsklinikum Ulm, Klinik fur Innere Medizin II, Ulm, Germany, 15Medical University of Warsaw, Warsaw, Poland, 16Cancer Research UK Clinical Centre, Southampton General Hospital, Southampton, United Kingdom, 17Oregon Health and Science University, Portland, United States, 18Pharmacyclics, Inc., Sunnyvale, United States, 19Ohio State University Comprehensive Cancer Center, Columbus, United States

Introduction: Ibrutinib is a first-in-class, oral inhibitor of Bruton's tyrosine kinase approved for MCL patients who have received at least 1 prior therapy. Previous results of the international, multicenter, open-label phase 2 trial demonstrated the promising activity and acceptable safety profile of ibrutinib in relapsed or refractory (R/R) MCL (Wang et al., N Engl J Med. 2013). Here, we present the updated safety and efficacy results of this trial at a median 26.7-month follow-up.

Methods: Patients (N = 111) received oral ibrutinib 560 mg once daily, and those with stable disease or better could enter a long-term extension study. The primary end point was investigator-assessed overall response rate (ORR) per the 2007 revised IWG criteria. Adverse events (AEs) were characterized by preferred terms using MedDRA version 16.1 and were evaluated over 6-month time intervals (1-6, 7-12, 13-18, 19-24, >24 months).

Results: The median patient age was 68 years (range, 40-84), with median 3 prior therapies (range, 1-5) and prior bortezomib in 43%. The median treatment duration was 8.3 months; 46% of patients were treated for >1 year, and 22% were treated for ≥2 years. The ORR was 67% (23% complete response) with a median duration of response of 17.5 months. The 24-month PFS and OS rates were 31% (95% CI: 22.3-40.4) and 47% (95% CI: 37.1-56.9), respectively. The most common AEs in >30% patients included diarrhea (54% all grade, 5% grade ≥3), fatigue (50% all grade, 5% grade ≥3), nausea (33% all grade, 1% grade ≥3), and dyspnea (32% all grade, 5% grade ≥3). All-grade infections were reported in 78% of patients, with the most frequent grade ≥3 infections being pneumonia (8%), urinary tract infection (4%), and cellulitis (3%). Any-grade bleeding was experienced by 50% of the patients, and grade ≥3 bleeding events in ≥2% patients were hematuria (2%) and subdural hematoma (2%). Common all-grade hematologic AEs were thrombocytopenia (22%), neutropenia (19%), and anemia (18%). Treatment discontinuation due to AEs was reported in 11% of patients. The prevalence of infection, diarrhea, and bleeding was highest for the first 6 months of therapy and less thereafter.

Conclusions: Ibrutinib continues to demonstrate a favorable benefit/risk profile in R/R MCL. Approximately one-third of patients remain progression-free at 2 years. The additional follow-up further supports the safety and durability of response with ibrutinib in MCL and does not reveal an increase in unforeseen AEs.

Disclosure: Martin Dreyling: Advisory Role: Pharmacyclics; Financing of Scientific Research: Pharmacyclics Kristie Blum: Expert Testimony: Pharmacyclics/Janssen.

V74 - Analysis of clinical characteristics and prognosis of patients with previously untreated diffuse large B-cell lymphoma of the gastrointestinal tract

Lehners N.1, Krämer I.1, Koschny R.2, Herfarth K.3, Egerer G.1, Ho A.D.1, Witzens-Harig M.1

1Uniklinik Heidelberg, Medizinische Klinik V, Heidelberg, Germany, 2Uniklinik Heidelberg, Medizinische Klinik IV, Heidelberg, Germany, 3Uniklinik Heidelberg, Radiologische Klinik, Heidelberg, Germany

Introduction: The gastrointestinal tract is one of the more frequent sites of extranodal involvement of diffuse large B-cell lymphoma (DLBCL). However, gastrointestinal DLBCL represents a very heterogeneous disease encompassing various biological entities. Therefore, optimal therapeutic strategies remain a matter of debate.

Methods: Patients with newly diagnosed DLBCL and gastrointestinal involvement presenting to the University Hospital of Heidelberg between 01/2000 and 12/2011 were included in this analysis. Progression-free survival (PFS) and overall survival (OS) were estimated using the Kaplan-Meier method. The impact of variables on PFS and OS was evaluated by univariate log-rank tests as well as by multivariate Cox regression.

Results: 82 patients were identified, 47 were male, 35 female. Median age was 60.5 years [range 19-86 years]. Site of involvement was stomach in 37 (45%), duodenum in 4 (5%), small intestine in 19 (23%), colon in 8 (10%) and multiple gastrointestinal sites in 14 (17%). 24 patients (29%) had IPI 0-1, 27 (33%) IPI 2-3, and 13 (16%) IPI 4-5; 18 patients had no evaluable IPI score. 5-year PFS was 65%, 5-year OS was 79%. There was no significant difference in regard to survival between the different sites of gastrointestinal involvement both in uni- and multivariate analysis. While neither age, sex, stage, and IPI score had a significant impact on survival in multivariate analysis, presence of B-symptoms was significantly associated with inferior PFS (p = 0.02) yet not with OS.

In regard to therapy, the addition of rituximab did not show a significant impact in regard to survival both in uni- and multivariate analysis. However, escalation of chemotherapy from CHOP to a more aggressive regimen (in our cohort mainly CHOEP) had significant positive impact on PFS (p = 0.004) and OS (p = 0.04) in univariate analysis; in multivariate analysis, it was significantly associated with prolonged PFS (p = 0.03) and showed a trend towards prolonged OS (p = 0.09). Patients receiving CHOEP had an excellent outcome with 5-year PFS 90% and 5-year OS 100%. While consolidative radiotherapy did not have an impact in univariate analysis, there was a trend towards beneficial impact on PFS (p = 0.09) and OS (p = 0.06) in multivariate analysis.

Conclusions: In our cohort of patients with DLBCL and gastrointestinal involvement escalation of chemotherapy to a more aggressive regimen than CHOP as well as consolidative radiotherapy seem to be beneficial therapeutic strategies.

Disclosure: No conflict of interest disclosed.

V75 - 1st-line treatment of patients with high-grade non-Hodgkin's lymphoma: Similar effectiveness with R-CHOP-14 or R-CHOP-21 - Data from the German prospective TLN Registry

Knauf W.1, Abenhardt W.2, Mohm J.3, Rauh J.4, Grugel R.5, Harde J.5, Jänicke M.5, Marschner N.6, TLN Registergruppe

1Onkologische Gemeinschaftspraxis, Frankfurt a.M., Germany, 2Münchner Onkologische Praxis Elisenhof, München, Germany, 3Onkologische Gemeinschaftspraxis, Dresden, Germany, 4Fachinternistische Gemeinschaftspraxis u. Therapiezentrum, Witten, Germany, 5iOMEDICO, Freiburg i. Br., Germany, 6Praxis für interdisziplinäre Onkologie & Hämatologie, Freiburg i. Br., Germany

Introduction: R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, prednisone) is the standard of care for patients (pts) with previously untreated high-grade (aggressive) non-Hodgkin's lymphoma (aNHL). Dose intensification of CHOP has shown ambiguous results. Whether the dose-dense two-weekly schedule (R-CHOP-14) is superior to the three-weekly schedule (R-CHOP-21) is a matter of debate. While clinical trials are restricted to highly selected pts, clinical registries offer insight into effectiveness of real-life treatment.

Methods: The open prospective clinical registry on lymphoid neoplasms (TLN Registry, ClinicalTrial.gov NCT00889798) documents the treatment of pts with lymphoid B-cell neoplasms by German office-based haematologists. Pts are followed for 5 years (yrs). Data regarding pts and tumour characteristics, co-morbidities, all systemic treatments and outcome are recorded. Between May 2009 and August 2014, a total of 3,798 pts have been recruited by 122 study sites.

Results: Of 565 pts with aNHL recruited at the start of 1st-line therapy with R-CHOP, 41% were treated with R-CHOP-14 and 59% received R-CHOP-21, with a tendency in favour of R-CHOP-21 over time.

Pts were 68 yrs old, 46% female, 25% presented with tumour stage IV (Ann Arbor) and 64% with at least one co-morbidity. Pts characteristics showed no considerable differences between the two schedules.

CHOP was applied for 6 cycles (median), rituximab for 8 cycles in R-CHOP-14 and 6 cycles in R-CHOP-21 (median). G-CSF was given in 98% (R-CHOP-14) and 63% (R-CHOP-21) of pts.

Objective response rate (ORR) as assessed by the local site was 96% (R-CHOP-14) and 93% (R-CHOP-21); the clinical (unconfirmed) complete remission rate was 65% (R-CHOP-14) and 66% (R-CHOP-21).

With a median follow-up of 25 months, 2yr progression-free survival rate (PFSREG) is 77% (R-CHOP-14) and 86% (R-CHOP-21). 2yr overall survival rate (OS) is 87% (R-CHOP-14) and 86% (R-CHOP-21). 10% of pts have received 2nd-line therapy. Overall, 10% of pts have been lost to follow-up.

At this point, the high rate of pts alive without progression (>80%) precludes multivariate regression analyses regarding factors affecting PFS or OS.

Conclusion: Our data show that in routine practice in Germany, pts with previously untreated high-grade NHL and treated with R-CHOP are more likely to receive the three-weekly schedule R-CHOP-21. First outcome data show that the effectiveness (ORR, PFSREG and OS) of both schedules is similar.

Disclosure: No conflict of interest disclosed.


Gesundheitspersonal neue Rollen, neue Aufgaben

V76 - Shaping careers of the future

Baumgartner U.1

1Kalaidos Fachhochschule, Departement Gesundheit, Zürich, Switzerland

Introduction: The demand for health care services is continuously increasing, boosted by demographic, epidemiological, economic and social changes (EVD, 2010). Nursing graduates from vocational and higher education levels are required to meet this challenge, and to provide sustainable care services. In 2014, 1,718 nurses graduated with a diploma in nursing (“Höhere Fachschule”) (Swiss Red Cross, 2015). Part of them are well prepared to advance their scope of practice with a BScN degree at a University of Applied Science (UAS) and foster innovative patient care. Some even pursue their career with an MScN, focusing either on practice development and project management, or on advanced practice roles for particularly complex patient situations, needed namely in geriatric and home care settings.

Methods: We aimed to identify the progress in clinical practice from a nursing diploma to a BScN degree. Firstly, we conducted an analysis of the clinical structures of our students. Secondly, we analyzed students' transfer reports written at the end of the BScN program. The CanMEDS model was used as reference for analysis (Canadian model for medical students' core competencies). It had also been used as framework for competencies in health care degree programs at Swiss UAS. The seven roles are: experts in care, communicator, team worker, manager, health advocate, scholar, professional (Ledergerber et al., 2009, p. 17).

Results: 10 reports from a BScN class were analysed using a coding scheme. Students demonstrate their systematic identification of patients' needs by applying clinical assessment skills. They are able to evaluate deteriorations more precisely and faster (expert). They communicate appreciatively and professionally (communicator). They extend their view from individuals to families and understand them as co-producers (health advocate). Students control the nursing process in challenging patient situations. They plan evidence-based interventions and reflect patient outcomes (professional). Students are able to support colleagues from other professions, including other students (scholar), and they are accountable within interprofessional teams (team worker).

Conclusion: Pursuing a BScN program after an associate degree, and studying within a didactics of research curriculum, enables and empowers graduates to meet new challenges and work in innovative care models.

Disclosure: Ursina Baumgartner: Employment or Leadership Position: Führungsposition.

Freier Vortrag


V80 - A new monoclonal antibody (CAL2) detects all types of CALRETICULIN mutations in formalin-fixed and paraffin embedded bone marrow biopsies

Stein H.1, Bob R.1, Dürkop H.1, Erck C.2, Kämpfe D.3, Kvasnicka H.-M.4, Martens H.5, Roth A.6, Streubel A.6

1Pathodiagnostik Berlin, Referenzzentrum Berlin für Hämatopathologie, Berlin, Germany, 2Helmholtz-Zentrum für Infektionsforschung GmbH, Braunschweig, Germany, 3Praxis für Onkologie, Lüdenscheid, Germany, 4Senckenbergisches Institut für Pathologie, Johann Wolfgang Goethe-Universität, Frankfurt am Main, Germany, 5Synaptic Systems GmbH, Göttingen, Germany, 6Medizinisches Versorgungszentrum am Helios Klinikum Emil von Behring, Labor für molekulare Diagnostik und Mikrobiologie, Berlin, Germany

Introduction: Recent advances in myeloproliferative neoplasms discovered CALRETICULIN (CALR) mutations as a major actor in these disorders. In contrast to JAK2 mutations being mainly associated with Polycythaemia vera, CALR mutations are only associated with primary myelofibrosis (PMF) and essential thrombocythaemia (ET) CALR mutations are present in the majority of primary myelofibrosis and essential thrombocythaemia patients lacking JAK2 and MPL mutations. Since these mutations are absent from reactive bone marrow (BM) lesions their presence indicates ET or PMF. So far these mutations are detectable only by molecular assays. Their molecular detection is cumbersome because of the great CALR-mutation heterogeneity. Therefore, the availability of a simple assay would be of great help. All CALR mutations reported lead to a frameshift generating a new 36 amino acid C-terminus.

Methods and results: We generated a monoclonal antibody (CAL2) to the C-neoterminus by immunizing mice with a representative peptide and compared its performance with Sanger sequencing data in 173 MPNs and other BM diseases. There was 100% correlation between the molecular and the immunhistological CALR-mutation-detection.

Conclusion: Thus, the immunodetection of CALR-mutations by the CAL2 antibody is a specific, sensitive, rapid, simple and low-cost method. The prevalent immunostaining of megakaryocytes confirms the strongest expression of mutated CALRETICULIN in megakaryocytes.

Disclosure: Harald Stein: Employment or Leadership Position: Geschäftsführer der Optistain GmbH; Stock Ownership: Anteilseigner an der Optistain GmbH; Honoraria: Anmeldung eingereicht; Expert Testimony: Eigenfinazierung.

V81 - Analyses of 357 patients with PMF, pET-MF and pPV-MF - a retrospective field study

Junghanß C.1, Springer G.2, Jentsch-Ullrich K.3, Balser C.4, Hauch U.5, Kisro J.K.6, Dengler J.7, Kiewe P.8, Königsmann M.9, Köchling G.10, Ostermann G.11, Kragl B.1

1Universitätsmedizin Rostock, Zentrum für Innere Medizin, Klinik III - Hämatologie, Onkologie und Palliativmedizin, Rostock, Germany, 2Hämato-Onkologische Schwerpunktpraxis Stuttgart am Hauptbahnhof, Stuttgart, Germany, 3Schwerpunktpraxis für Hämatologie und Onkologie, Magdeburg, Germany, 4Praxis für Innere Medizin, Hämatologie & internistische Onkologie, Marburg, Germany, 5Onkologische Praxis, Erfurt, Germany, 6Ambulantes Tumorzentrum, Neustadt, Germany, 7Schwerpunktpraxis Onkologie, Heilbronn, Germany, 8MVZ Onkologischer Schwerpunkt am Oskar-Helene-Heim, Berlin, Germany, 9Onkologisches Ambulanzzentrum am Diakoniekrankenhaus Henriettenstiftung gGmbH, Hannover, Germany, 10Onkologische Schwerpunktpraxis, Villingen, Germany, 11Novartis Pharma GmbH, Business Unit Oncology, Nürnberg, Germany

Introduction: Primary Myelofibrosis (PMF), post essential thrombocythemia (pET) and post polycythemia vera (pPV) MF are rare diseases. Therapeutical options have significantly improved by the development of novel drugs, improvement of allogeneic stem cell transplantation (SCT) and supportive care. Whereas PMF, pET-MF and pPV-MF clinical trial participants are well characterized regarding personal, disease and treatment issues this is not the case for the general MF population. Here, we performed a questionnaire poll in multiple institutions gathering data on 357 patients.

Methods: A two sided questionnaire asking for general patient and disease specific information as symptoms, splenomegaly, prognostic factors, past and current treatment and blood values, degree of MF in bone marrow and transfusion frequency was developed. Participating centers (n = 15) were mostly private offices in Germany. Data of 357 patients was documented by the participants and analyzed centrally. A median of 20 pts (range 10-60 pts) per center were documented in 2013/14.

Results: Gender distribution was: 48% female, 52% male. Age at first diagnosis was: < 50y (10%), 51-60y (21%), 61-70y (33%), >70y (36%). Most pts had PMF (69%), pPV-MF (13%), pET-MF (18%). Major disease durations: 1-5y (42%) and >5y (45%). Thrombocythopenia: <50GPT/l: 6.3%, 50 to <100GPT/l: 14.4% and thrombocytosis ≥450GPT/l: 20.7%. Hemoglobin [g/dl]: ≥10 (60%), 8-10.0 (28%), <8 (12%). 16% of all pts had WBC >25.000/µl and 13% showed circulation blasts in the peripheral blood, respectively. Further DIPPS characteristics were: >65y (72%) and constitutional symptoms 24%. Common symptoms included splenomegaly (66%), decreased fitness (41%) and weight loss (23%). Pruritus was present in 5% and night sweats in 13% of all pts. Concomitant diseases were: cardiac 57%, GI 12%, diabetes 10%, secondary neoplasia 9%. Non-medical treatments were rare: SCT 4.2%, splenectomy 2.8%, spleen irradiation 3.6%. Medical treatment included cytostatics (37%), anticoagulation (30%), JAK-inhibitors (27%), none (20%). Only 36% of all pts received any RBC transfusion, 12% received >50 units.

Conclusion: Daily practice MF pts share several characteristics with the COMFORT trial cohort. As expected the diseases were not as progressed as in the trials. Interestingly male gender predominance was not as pronounced in our study. SCT is a rarely used treatment whereas JAK2 inhibitors are frequently used.

Disclosure: No conflict of interest disclosed.

V82 - Interferon gamma secreted by activated NK cells has a negative impact on the treatment with tyrosine kinase inhibitors (TKI) in CML cells

Held S.A.E.1, Heine A.1, Schönberg K.1, Riethausen K.1, Daecke S.N.1, Wolf D.1, Brossart P.1

1Universitätsklinikum Bonn, Medizinische Klinik III, Bonn, Germany

Introduction: Chronic myeloid leukemia (CML) is a myeloproliferative disease which resolves in the constitutive activity of the BCR-ABL protein. Since the introduction of imatinib as first line therapy for CML, the overall survival of patients could be significantly extended. However, mutations of the BCR-ABL protein and environmental effects mediated by adhesion molecules, cytokines and growth factors were shown to induce resistance to TKIs. Therefore, we determined the role of activated NK cells and analyzed the effects of released IFNγ on CML cells.

Methods: CML cell lines and primary cells from patients with CML were treated with imatinib or nilotinib and incubated with IL12/IL18 activated or naïve NK cells in transwell experiments. Induction of apoptosis was analyzed by flow cytometry. In addition, IFNγ was used together with TKIs and apoptotic cell death as well as MHC class I expression was determined. Furthermore, effects of IFNα in contrast to IFNγ on apoptosis induction and MHC class I expression were examined.

Immunoblotting was performed to analyze the involved pathways.

Results: Incubation of CML cells in transwell assays with activated NK cells resulted in significant inhibition of the apoptosis induction by imatinib and nilotinib. To further understand the mechanisms mediating these effects, CML cells were treated with IFNγ in addition to the TKIs. In line with the results from previous experiments, IFNγ reduced the rate of cell death mediated by imatinib and nilotinib. Interestingly, in contrast to IFNα, IFNγ inhibited the TKI-induced downregulation of MHC-class I molecules on CML cells. In line with these results, Western blot analysis of K-562 and Kyo-1 cells incubated with imatinib or nilotinib in combination with IFNγ showed an increased activation of Jak2 in contrast to TKI treatment alone. To complete this, the activation of Jak2 by IFNγ resulted in an elevated nuclear expression of the transcription factor Runx1 which is supposed to play an important role in CML disease persistence.

Conclusion: Our results demonstrate that IFNγ released by activated NK cells can interfere with the action of TKIs in CML cells by reducing the TKI mediated apoptosis induction followed by an increased activation of Jak2 and expression of Runx1. These results might have important implications on the therapy in CML as well as in the development of immunotherapeutic approaches.

Disclosure: No conflict of interest disclosed.

V83 - The observed incidence of cardiovascular ischemic events in dasatinib-treated patients in clinical trials compared with the expected incidence based on general external populations

Saglio G.1, le Coutre P.2, Cortes J.3, Mayer J.4, Rowlings P.5, Mahon F.-X.6, Kroog G.7, Gooden K.7, Subar M.7, Shah N.P.8

1University of Torino, Ospedale San Luigi Gonzaga, Orbassano-Torino, Italy, 2Charité, Campus Virchow Klinikum, Universitätsmedizin Berlin, Berlin, Germany, 3The University of Texas MD Anderson Cancer Center, Houston, United States, 4University Hospital Brno, Brno, Czech Republic, 5Calvary Mater Newcastle Hospital, University of Newcastle, Newcastle, Australia, 6Centre Hospitalier Universitaire de Bordeaux and Institut Bergonié, Bordeaux, France, 7Bristol-Myers Squibb, Princeton, United States, 8UCSF School of Medicine, San Francisco, United States

Introduction: Arterial ischemic events have been observed with some BCR-ABL1-targeted tyrosine kinase inhibitors (TKIs); therefore, the incidence of cardiovascular (CV) ischemic events in dasatinib-treated patients was assessed.

Methods: CV ischemic events were assessed in a pooled population of patients with Ph+ leukemia across 11 clinical trials (N = 2712) and the two Phase 3 studies DASISION (N = 519, newly diagnosed patients with CML) and READY (N = 1518, patients with prostate cancer). Standardized incidence rates (SIRs) were calculated to determine if the number of observed events was different than expected compared with reference populations (general, N = 116,000,000; males, N = 56,000,000; CML patients, N = 16,000; prostate cancer patients, N = 530,000) from Truven's MarketScan Commercial Claims database, 2008-2013, narrowed to mimic trial eligibility. SIRs were calculated by dividing the observed number of events in dasatinib-treated patients by the expected number of events, based on dasatinib exposure and reference population event rates.

Results: Within the pooled population, 96 patients (4%) had CV ischemic events. In DASISION, 10 dasatinib- and 4 imatinib-treated patients had any grade CV ischemic events. In READY, 18 patients in the dasatinib arm and 9 patients in the placebo arm had any grade CV ischemic events. The majority of patients with an event had history and/or risk factors for atherosclerosis (77/96 in the pooled population; 8/10 with dasatinib in DASISION). Most cardiovascular ischemic events occurred within 1 year of dasatinib initiation (69/96 patients in the pooled population and 7/10 patients in DASISION). Based on SIRs, the observed number of cardiovascular ischemic events in dasatinib-treated patients was not higher than expected (95% confidence intervals were less than or included 1). SIRs should be interpreted with caution due to the limitations of the comparison between clinical and claims data (e.g. coding differences and surveillance bias).

Conclusions: The incidence of CV ischemic events in dasatinib-treated patients was low in this analysis. The majority of patients with an event had history and/or risk factors with most events occurring early and largely restricted to 1 year after initiating therapy. SIRs suggest the number of CV ischemic events among dasatinib-treated patients was not higher than expected.

Disclosure: Giuseppe Saglio: Financing of Scientific Research: Novartis, Bristol-Myers Squibb, ARIAD, Pfizer Neil Shah: Expert Testimony: Bristol-Myers Squibb, ARIAD, Pfizer.

V84 - CA180-034 final study results: Seven-year (yr) follow-up of chronic-phase chronic myeloid leukemia (CML-CP) patients (pts) with imatinib resistance or -intolerance receiving dasatinib, a 2nd-generation BCR-ABL tyrosine kinase inhibitor (TKI)

Müller M.C.1, Shah N.P.2, Rousselot P.3, Schiffer C.4, Rea D.5, Cortes J.6, Mohamed H.7, Healey D.7, Kantarjian H.6, Hochhaus A.8, Saglio G.9

1Medizinische Fakultät Mannheim der Universität Heidelberg, Mannheim, Germany, 2UCSF School of Medicine, San Francisco, United States, 3Hôpital Mignot, University Versailles Saint-Quentin-en-Yvelines, Le Chesnay, France, 4Karmanos Cancer Institute, Wayne State University School of Medicine, Detroit, United States, 5Hôpital Saint-Louis, Paris, France, 6The University of Texas MD Anderson Cancer Center, Houston, United States, 7Bristol-Myers Squibb, Princeton, United States, 8Universitätsklinikum, Jena, Germany, 9University of Torino, Ospedale San Luigi Gonzaga, Orbassano-Torino, Italy

Introduction: CA180-034 (NCT00123474), a prospective, randomized phase 3 study, compared the dose and schedule of dasatinib, a BCR-ABL TKI, for optimal benefit/risk ratio among imatinib-resistant or -intolerant CML-CP pts. Previous study results demonstrated significant efficacy of dasatinib, leading to its current approval at 100 mg once daily (QD) in this population. We present here the final 7-yr analysis for efficacy and safety in CA180-034, which is the longest follow-up of any 2nd-generation BCR-ABL TKI to date.

Methods: Pts (n = 670) were randomized to dasatinib: 100 mg QD (n = 167); 50 mg twice daily (BID; n = 168); 140 mg QD (n = 167); or 70 mg BID (n = 168). To manage inadequate response or adverse events (AEs), dose modifications (total daily dose [TDD] of 20-180 mg) were allowed. After 2 yrs, the protocol was amended to allow switching to a QD regimen with the same TDD.

Results: Approximately 55% (50 mg BID) and 51% (70 mg BID) of pts treated after the protocol amendment switched to QD dosing by the last recorded dose. Median duration of therapy was longer in the 100 mg QD arm (37.4 months [mos]) vs other dose groups. Rates for major molecular response (MMR), progression-free survival (PFS), and overall survival (OS) were similar across doses (Table). In an exploratory landmark analysis, pts in the 100 mg QD arm with BCR-ABL ≤10% (on the International Scale) at 3 mos had improved 7-yr PFS and OS rates relative to pts with BCR-ABL >10% (PFS [95% CI]: 56% [43-67] vs 21% [10-34]; OS [95% CI]: 72% [60-81] vs 56% [42-68], respectively). For 100 mg QD, most non-hematologic and hematologic AEs (all grades) first occurred within 24 mos. Fewer grade 3-4 AEs occurred at 100 mg QD. With a minimum of 7 yrs of follow-up, pleural effusion rate was lower in the 100 mg QD arm vs other dose groups (any grade [severe]: 28% [5%] vs 35% [8%], respectively). Overall, 128 pts (19%) received ≥7 yrs of dasatinib therapy.

Conclusions: Long-term follow-up with dasatinib demonstrates durable efficacy and benefit in imatinib-resistant or -intolerant pts with CML-CP, particularly if achieving BCR-ABL ≤10% at 3 mos. Dasatinib is well-tolerated, with most AEs occurring early in therapy; however, pleural effusion occurred throughout 7 yrs of treatment. No new safety signals were detected.


Disclosure: Martin Müller: Expert Testimony: Bristol-Myers Squibb, Novartis, ARIAD; Other Financial Relationships: Personal fees & non-financial support - Bristol-Myers Squibb, Novartis, ARIAD Giuseppe Saglio: Financing of Scientific Research: Novartis, Bristol-Myers Squibb, ARIAD, Pfizer.

V85 - Long-term 5-year results of the phase 3 DASISION trial (dasatinib versus imatinib in newly diagnosed chronic myeloid leukemia in chronic phase [CML-CP])

Stegelmann F.1, Cortes J.2, Saglio G.3, Baccarani M.4, Kantarjian H.2, Mayer J.5, Boqué C.6, Shah N.P.7, Chuah C.8, Casanova L.9, Narayanan G.10, Bradley-Garelik B.11, Manos G.11, Hochhaus A.12

1Universitätsklinikum, Ulm, Germany, 2The University of Texas MD Anderson Cancer Center, Houston, United States, 3University of Torino, Ospedale San Luigi Gonzaga, Orbassano-Torino, Italy, 4S. Orsola-Malpighi Hospital, University of Bologna, Bologna, Italy, 5University Hospital, Brno, Czech Republic, 6Institut Català d'Oncologia, Hospital Duran i Reynals, L'Hospitalet, Barcelona, Spain, 7UCSF School of Medicine, San Francisco, United States, 8Singapore General Hospital and Duke-National University of Singapore Graduate Medical School, Singapore, Singapore, 9Instituto Nacional de Enfermedades Neoplásicas, Lima, Peru, 10Regional Cancer Centre, Medical College, Thiruvananthapuram, Kerala, India, 11Bristol-Myers Squibb, Princeton, United States, 12Universitätsklinikum, Jena, Germany

Introduction: The randomized, phase 3 DASISION trial has demonstrated improved efficacy with dasatinib (DAS) versus imatinib (IM) in newly diagnosed CML-CP patients (pts). We report here the results of the final 5-year (yr) analysis of long-term efficacy and safety outcomes.

Methods: Treatment-naïve CML-CP pts were randomized to receive DAS 100 mg once daily (QD; n = 259) or IM 400 mg QD (n = 260) and were followed for the predefined minimum of 5 yrs.

Results: At the time of study closure, 61% and 63% of DAS- and IM-treated pts, respectively, were still on initial therapy. Confirmed complete cytogenetic response (cCCyR), major molecular response (MMR), and MR4.5 rates by 5 yrs continued to be higher for DAS versus IM (cCCyR: 83% vs 78%; MMR: 76% vs 64%; MR4.5: 42% vs 33%). DAS had fewer cases of transformation to accelerated/blast phase (AP/BP) on study or after discontinuation (4.6%) compared with IM (7.3%). Overall 5-yr progression-free survival (PFS) and overall survival (OS) rates were high and similar for DAS (PFS: 85%; OS: 91%) and IM (PFS: 86%; OS: 90%). A higher proportion of pts on DAS achieved BCR-ABL ≤10% at 3 months (84%) compared with IM (64%). The improved PFS, OS, and lower rates of transformation to AP/BP previously reported for pts who achieved BCRABL ≤10% versus >10% at 3 months were maintained at 5 yrs for both treatment arms. Fifteen pts on DAS and 19 pts on IM had BCR-ABL mutations. No unexpected safety events were identified in either treatment arm at 5 yrs; however, additional DAS-treated pts experienced a first occurrence of pleural effusion (PE). Overall, 74 DAS-treated pts (29%) experienced PE (grade1/2=67; no grade 5). Fifteen pts discontinued DAS due to PE. Arterial ischemic events overall were not common (DAS: n = 12; IM: n = 6). Six of 14 DAS-treated pts with pulmonary hypertension discontinued therapy. No DAS-treated pts had World Health Organization Group 1 pulmonary arterial hypertension (confirmed by right heart catheterization) or peripheral arterial occlusive events.

Conclusion: At 5 yrs, DAS 100 mg QD demonstrated superior efficacy compared with IM 400 mg QD as first-line therapy for CML. More pts on DAS achieved BCR-ABL ≤10% at 3 months, and there were higher cumulative rates of response and a lower rate of transformation on DAS. PFS and OS were similar for both treatment arms. No new safety signals were reported. These consistent results confirm that DAS offers meaningful advantages over IM for pts with newly diagnosed CML-CP.

Disclosure: Frank Stegelmann: Other Financial Relationships: Bristol-Myers Squibb, Novartis, Pfizer, ARIAD Andreas Hochhaus: Expert Testimony: ARIAD, Bristol-Myers Squibb, Merck Sharpe & Dohme, Novartis, Pfizer.

Wissenschaftliches Symposium

Long Term Survivorship

V86 - Social and financial consequences of cancer

Seifart U.1

1Klinik Sonnenblick, Marburg, Germany

The number of the long time “Cancer-Survivor” does increase, because of rising incidence and improved prognosis of patients (pts) [1]. In germany nearly 50% of the 1. 4 million cancer pts are younger than 65y. Only about 50% of them will return to work completely [2]. This means that every year approximately 100,000 tumour pts will face the question whether and how return to work.

The unfitness for work results in an increased risk of long-term unemployment or retirement. This may become for the affected persons a considerable financial distress (FD) which is linked with depressed quality of life and social isolation [3]. FD can show a significantly bigger load than possible physical or psychic side effects in particular for young adults, independent of the income.

The diminished social standing may also correlates with the prognosis our pts. For germany an evaluation of ten cancer registers showed that tumour pts from income-strong regions showed a better 5 year overall survival rates than cancer pts from socially weaker areas [4].

The risk a longer-term acquisition inability correlates, on one hand with the tumour entity, but also with the intensity of the therapy.

A risk decrease can be reached by professional help. This is available in the oncological rehabilitation [5], but also to the cancer advice centres.


1 Robert Koch-Institut (Hrsg.). Beiträge zur Gesundheitsberichterstattung des Bundes. Krebs in Deutschland 2009/2010. 9. Ausgabe, 2013.

2 Mehnert A Employment and work-related issues in cancer survivors. Crit Rev Oncol Hematol 2011;77(2):109-130.

3 Shankaran V et al. Risk factors for financial hardship in patients receiving adjuvant chemotherapy for colon cancer: a population-based exploratory analysis. J Clin Oncol 2012;30(14):1608-1614.

4 Jansen, L., et al for the GEKID Cancer Survival Working Group. Socioeconomic deprivation and cancer survival in Germany: An ecological analysis in 200 districts in Germany. Int J Cancer 2014;134: 2951-2960.

5 Mehnert A, et al . Predictors of employment among cancer survivors after medical rehabilitation - a prospective study. Scand J Work Environ Health 2013;39(1):76-87.

Disclosure: No conflict of interest disclosed.

V87 - Cognitive dysfunction with regard to return to work

Rick O.1

1Klinik Reinhardshöhe, Onkologie, Bad Wildungen, Germany

Cognitive dyfunction (CD) is a short-term, long-term or permanent functional disorder in attention, concentration, thought processes, memory (especially short-term memory), learning ability and the ability to perform complex tasks.

Depending on the investigated tumor disease, the prevalence of CD varies significantly between 12% and 68%, it can increase to 80% 6 months after the end of chemotherapy and is described up to 20 years after the cancer treatment.

CD is most likely multifactorial. Besides the actual tumor, genetic conditions, age and the anti-tumor therapy (chemotherapy, surgery, radiation therapy) play the key role. The interplay of these components ultimately determines the severity of the cognitive disorder.

The correlation between CD and participation in the labor market is only insufficiently investigated and returns partly contradictory outcomes. In patients with Hodgkin lymphoma, CD has a negative impact on the return to working life. Another study has shown that cancer patients with CD resume professional activity less frequently than patients without CD. In a study of 45 patients with breast cancer, the negative influence of CD on return to work could not be determined.

Despite the inhomogeneous and little evidence-based data, it can be assumed that especially patients with special professional requirements for cognitive function can be prevented from resuming professional activity by this kind of disorder. Moderate instances of memory impairment can make the performing of simple professional activities impossible. In particular disorders of the ultra-short-term memory (up to about sixty minutes back) normally impact performance to such a significant extent that the pursuit of a professional activity is no longer possible. It is essential to clarify, on a case-by-case basis, which individual professional activities are reduced or prevented by CD and how. This is best done by means of a differentiated vocational true-to-life stress test in the context of oncological rehabilitation.

Disclosure: No conflict of interest disclosed.

V88 - Sport and therapeutic exercise in the long-term treatment of young cancer patients

Koenig V.1

1Klinik Bad Oexen, Bad Oeynhausen, Germany

Functional impairments and therapy-related disorders may develop after cancer therapy, if intensifed therapeutic strategies are used, as is often the case in adolescents and young adults (AYA). Multimodal therapy (combination of surgery, radiotherapy and cytotoxic chemotherapy) might lead to general fatigue. To recover from cancer therapy, medical doctors often recommend physical inactivity. As a consequence, secondary problems may develop. However, for a sustanied physical improvement after cancer treatment, an adequate level of physical training is necessary, thus supporting the adjustment of body structures (i.e. strengthening of muscle fibers, increase in cardiac and pulmonary efficiency etc.). It has been observed that regular exercising after cancer therapy improves overall survival. In AYA physical fitness is markedly reduced after antineoplastic treatment compared to older adults, as AYA receive more aggressive therapeutic regimens. AYA should therefore undergo an intensified physical training program compared to older patients. This can be achieved within an age-specific group rehabilitation in a specialist clinic (in-patient program). A high-level exercise schedule within a peer group is effective by the competition initiated among the group members. Typically, such a program takes 4 weeks and is only a first step on the way back to life. For a complete recovery up to the old performance levels, the physical training program has to be continued for at least several months. The patients should be informed and motivated accordingly. Similar programs can also be applied in a single exercise environment in an out-patient setting, with reduced efficiency (more motivation needed to be successful).

Disclosure: No conflict of interest disclosed.

V89 - Impact and effects of sleep disturbances in younger cancer patients

Strik H.1

1Universität Marburg, Neurologische Klinik, Marburg, Germany

Sleep disturbances, daytime sleepiness and psychiatric symptoms are fequent, but only rarely asked for in daily oncological routine. Most probably, patients and their treating physicians consider sleep disturbances to be inevitable in cancer disease. Usually, oncologists ask their patients for tumor specific symptoms like pain, dyspnoea, loss of appetite or nausea. The quality of sleep is often disregarded.

Disturbances of sleep can have various reasons. Undesired effects of tumor therapy can inhibit the sleep as well as supportive treatment. Most frequently, sleep disturbances are caused by psychosocial distress which can be depression as well as financial worries. Moreover, indirect effects of the tumor disease or its treatment on the hormonal regulation regulation of sleep are possible. The extensive treatment of tumor diseases should therefore include a systematic assessment of sleep disturbances and treatment of their causes.

Disclosure: No conflict of interest disclosed.

Freier Vortrag

Kopf-Hals-Tumore I

V90 - Prospective multicenter phase II study of the anti-EGFR (epidermal growth factor receptor) antibody panitumumab (P) in patients with platinum pre-treated, advanced head and neck squamous cell cancer (HNSCC)

Siano M.1,2, Molinari F.3, Martin V.3, Mach N.4, Früh M.1, Crippa S.3, Ghielmini M.5, Frattini M.3, Espeli V.5

1Cantonal Hospital St. Gallen, Clinic of Oncology / Hematology, St. Gallen, Switzerland, 2Fondazione IRCCS Istituto Nazionale dei Tumori, Head and Neck Medical Oncology Unit, Milan, Italy, 3Institute of Pathology, Laboratory of Molecular Diagnostics, Locarno, Switzerland, 4Geneva University Hospital, Clinical Research Unit of the Dr. Henri Dubois-Ferrière Dinu Lipatti Foundation Oncology Center, Geneva, Switzerland, 5Oncology Institute of Southern Switzerland, Medical Oncology, Bellinzona, Switzerland

Introduction: We performed a multicenter phase II study with the fully human anti-EGFR-antibody P, administered as a single agent in platinum-pretreated HNSCC patients (pts). We present the first safety and efficacy data in this pt population.

Material and methods: Recurrent or metastatic HNSCC pts previously treated with platinum containing chemotherapy in four tertiary Swiss cancer centers were included. Previous anti-EGFR-antibody treatment (EAB) was allowed if pts had no progression during or within six months after therapy. P was administered intravenously every two weeks at a dose of 6mg/kg. Primary endpoint was response rate (RR), secondary endpoints were duration of response (DR), progression-free survival (PFS), and safety. Adverse events (AEs) were graded following the NCI Common Terminology Criteria (CTCAE V3.0). Tumor response was assessed every 8 weeks according to RECIST V1.1. Tumor tissue samples were collected and centrally analyzed for potential, predictive biomarkers.

Results: 33 pts received in total 151 and a median of 4 (range1-16) cycles of P. Ten pts (30.3%) were EAB pre-treated and 26 pts (78.8%) were platinum resistant or refractory. RR was 6.1% with two partial responses (PR) and 39.4% (13 pts) stable diseases (SD), resulting in a clinical benefit rate of 45.5%. Both PRs occurred in platinum-resistant pts.

Median PFS was 3.0 months (95% CI: 1.8, 5.5) and median duration of clinical benefit (PR+SD) 8.4 months (95% CI: 3.6, 17.7). Pts with PR were free of progression for 8 and 14 months.

54.5% of pts had only grade 1-2 toxicity, while 27.3% experienced grade 3-4 P-related adverse events. Cutaneous rash was the most frequent (27/33 pts in total [81.8%], of Grade 3 in 2 [6.1%], requiring treatment withdrawal in 1pt), followed by hypomagnesaemia (22/33 pts [66.7%], of Grade 3-4 in 5 [15.2%]). One pt died after the first P administration, presumably due to drug-related severe alveolitis since no other cause was identified, and one pt experienced serious hypokalaemia grade 4. P was in general well tolerated and no infusion-related reactions occurred.

Conclusion: We present the first efficacy and safety results of P monotherapy in platinum pre-treated HNSCC pts. P was well tolerated showing antitumor activity and remarkable duration of clinical benefit. Toxicity profile and convenience support consideration of P in this setting.

Disclosure: No conflict of interest disclosed.

V91 - Subgroup analysis according to differentiated thyroid cancer histology in phase 3 (SELECT) trial of lenvatinib

Elisei R.1, Schlumberger M.2, Tahara M.3, Robinson B.4, Brose M.5, Dutcus C.6, Zhu J.6, Newbold K.7, Kiyota N.8, Kim S.-B.9, Sherman S.10, Wirth L.11

1Endocrine Unit, Department of Clinical and Experimental Medicine, University of Pisa, Pisa, Italy, 2Department of Nuclear Medicine and Endocrine Oncology, Gustave Roussy and University Paris-Sud, Villejuif, France, 3Department of Head and Neck Medical Oncology, National Cancer Center Hospital East, Kashiwa, Japan, 4Kolling Institute of Medical Research, University of Sydney, New South Wales, Australia, 5Department of Otorhinolaryngology: Head and Neck Surgery, Abramson Cancer Center of the University of Pennsylvania, Philadelphia, United States, 6Eisai Inc, Woodcliff Lake, United States, 7The Royal Marsden National Health Service Trust, London, United Kingdom, 8Department of Medical Oncology and Hematology. Kobe University Hospital, Kobe, Japan, 9Department of Oncology, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Korea, Republic of, 10Department of Endocrine Neoplasia and Hormonal Disorders, Division of Internal Medicine, The University of Texas M. D. Anderson Cancer Center, Houston, United States, 11Department of Medicine, Massachusetts General Hospital, Boston, United States

Background: The previously reported results of the SELECT trial demonstrated that lenvatinib significantly prolonged progression-free survival (PFS) in patients with 131I-refractory differentiated thyroid cancer (RR-DTC) compared with placebo. Median PFS in lenvatinib and placebo were 18.3 and 3.6 months, respectively (hazard ratio [HR]: 0.21, 99% confidence interval [CI]: 0.14-0.31; P < 0.0001). This subgroup analysis of SELECT trial examined clinical outcomes of each histological characteristics.

Methods: In the SELECT trial, 392 patients with RR-DTC were enrolled and randomized 2:1 to lenvatinib (24 mg/day, 28-day cycle) or placebo. The primary endpoint was PFS and the secondary endpoints were including objective response rate (ORR), overall survival (OS) and safety. This analysis examines the clinical outcomes of prespecified subgroups divided based on histology: papillary thyroid cancer (PTC, n = 259) and follicular thyroid cancer (FTC, n = 133).

Results: PTC patients treated with lenvatinib and placebo had median PFS of 16.4 months and 3.5 months, respectively (HR: 0.27; 95% CI: 0.19-0.38). FTC patients treated with lenvatinib had a median PFS that had not yet been reached vs 3.7 months for placebo (HR: 0.10; 95% CI: 0.05-0.19). ORR results of the patients treated with lenvatinib were 63.9% (PTC) and 66.3% (FTC). Although the OS advantage with lenvatinib vs placebo had detected with no significant difference (HR: 0.73; 95% CI: 0.50-1.07) in the overall population, possibly due to the cross-over design, the clinically meaningful OS advantage was observed in lenvatinib-treated FTC group (HR: 0.41; CI: 0.18-0.97). A treatment-related TEAE (TR-TEAE) and grade ≥3 TR-TEAEs occurred in 96.4% and 76.3% in PTC patients and 98.9% and 75.0% in FTC patients, respectively.

Conclusions: Clinical benefits of lenvatinib treatment were observed in both examined PTC and FTC subgroups. These results such as the promising response in the FTC patients warrant further investigation to enrich the evidence.

Disclosure: Rosella Elisei: Advisory Role: Bayer, AstraZeneca, Genzyme; Financing of Scientific Research: Bayer, AstraZeneca, Genzyme; Other Financial Relationships: Fees from: Genzyme, Bayer, Sobi/Exelixis, AstraZeneca Lori Wirth: Other Financial Relationships: Fees from: Eisai, Novartis.

V92 - Phase II explorative trial to prospectively investigate predictive molecular biomarkers for efficacy of panitumumab (P) in platinum-pretreated head and neck squamous cell cancer (HNSCC)

Siano M.1,2, Molinari F.3, Martin V.3, Mach N.4, Früh M.1, Crippa S.3, Ghielmini M.5, Espeli V.5, Frattini M.3

1Cantonal Hospital St. Gallen, Clinic of Oncology / Hematology, St. Gallen, Switzerland, 2Fondazione IRCCS Istituto Nazionale dei Tumori, Head and Neck Medical Oncology Unit, Milan, Italy, 3Institute of Pathology, Laboratory of Molecular Diagnostics, Locarno, Switzerland, 4Geneva University Hospital, Clinical Research Unit of the Dr. Henri Dubois-Ferrière Dinu Lipatti Foundation Oncology Center, Geneva, Switzerland, 5Oncology Institute of Southern Switzerland, Medical Oncology, Bellinzona, Switzerland

Introduction: Monoclonal antibodies directed against EGFR (E-Mab) were extensively investigated in HNSCC. Even if clinical efficacy in terms of survival was shown to be marginal, there seems to be a subgroup of pts with clear benefit and durable responses. So far, no single biomarkers or biomarker combination pattern was found to identify this subgroup. The aim of this study was to analyze pre-specified biomarkers in a prospective phase II trial with P in HNSCC pts.

Methods: Pts with platinum-pretreated HNSCC included in a prospective phase II multicenter trial exploring P as 2nd line treatment consented for molecular biomarker analysis on available tumor tissue. As P is not active through indirect mechanisms like ADCC (antibody dependent cellular cytotoxicity), it seems to be an ideal candidate to examine the pure effect on EGFR pathway, held responsible for durable responses. To find a predictive marker pattern for response, a central laboratory investigated the following markers: KRAS, NRAS, HRAS, PI3KCA, BRAF gene mutations by Sanger sequencing; EGFR gene status by FISH; HPV genotyping.

Results: 25 pts consented for the biomarker sub-study. Tumor tissue was available in all pts. One case was excluded for bad quality of DNA. Two uncommon KRAS mutations (G48E, T50I, 8%) and 3 (12.5%) PIK3CA mutations (all E545K) were detected; all the other genes were wild type. HPV high-risk 16 was found in 9 (37.5%) and EGFR copy number gain (CNG) in 12 pts (50%). No correlation between response and molecular alterations was observed, with the exception of EGFR: all 3 PR patients showed EGFR CNG, a feature not identified in SD or PD patients.

Conclusion: EGFR CNG by FISH may identify HNSCC patients who will benefit from P administration and may be a useful marker to predict efficacy of P. This preliminary observation needs to be confirmed in a larger series. The cascade of MAP kinases (RAS, BRAF) was not predictive. The role of PIK3CA mutations remains to be elucidated.

Disclosure: Marco Siano: Other Financial Relationships: Amgen stellte das Medikament zur Verfügung für die Studie Milo Frattini: Other Financial Relationships: Amgen stellte das Medikament zur Verfügung für die Studie.

V93 - Prognostic and predictive factors correlated with treatment outcomes for radioactive Iodine-refractory differentiated thyroid cancer (RAI-rDTC) patients receiving sorafenib or placebo on the phase III DECISION trial

Paschke R.1, Schlumberger M.2, Elisei R.3, Pacini F.4, Jarzab B.5, Giannetta L.6, Bastholt L.7, de la Fouchardiere C.8, Worden F.P.9, Shong Y.K.10, Smit J.W.11, Kappeler C.12, Molnar I.13, Brose M.S.14

1Leipzig University, Department for Endocrinology and Nephrology, Leipzig, Germany, 2Gustave Roussy, Villejuif, France, 3University of Pisa, Department of Clinical and Experimental Medicine, Pisa, Italy, 4University of Siena, Unit of Endocrinology, Siena, Italy, 5Maria Sklodowska-Curie Memorial Cancer Centre and Institute of Oncology, Gliwice, Poland, 6Ospedale Niguarda Ca' Granda, Milan, Italy, 7Odense University Hospital, Odense, Denmark, 8Hospices Civils-Centre Anticancéreux, Consortium Cancer Thyroïdien, Lyon, France, 9University of Michigan Health System, Ann Arbor, United States, 10Asan Medicine Center, Seoul, Korea, Republic of, 11Radboud University Nijmegen Medical Center, Department of Internal Medicine, Nijmegen, Netherlands, 12Bayer Pharma AG, Berlin, Germany, 13Bayer HealthCare Pharmaceuticals, Whippany, United States, 14Abramson Cancer Center of the University of Pennsylvania, Department of Otorhinolaryngology, Head and Neck Surgery, Philadelphia, United States

Background: The DECISION trial established that sorafenib prolonged progression-free survival (PFS) compared to placebo in patients with progressive RAI-rDTC (Lancet 2014). Here we sought to identify prognostic and predictive factors correlated with treatment outcomes.

Methods: Multivariate Cox proportional hazards models adjusted for treatment effect and subgroup analyses defined by maximum target lesion size and existence of disease-related symptoms were used to explore the relationship between clinical baseline variables and PFS. Patients were deemed symptomatic at entry if they had symptoms/findings consistent with thyroid cancer reported in the medical history or pre-treatment adverse event dataset.

Results: A total of 417 patients were randomized to receive placebo (n = 210) or sorafenib (n = 207). Multivariate Cox model analyses indicated that lower maximum individual target lesion size, lower number of lesions, thyroglobulin levels at baseline less than the median (486 ng/ml) and region Asia versus Europe and North America were prognostic for longer PFS in placebo patients and in all patients when adjusted for treatment. Subgroup analyses indicated that patients whose maximum individual target tumor size were <1.5 cm had longer PFS and appeared to have less benefit from sorafenib treatment than patients with lesions ≥1.5 cm. Lesions ≥1.5 cm as well as lung metastases were predictive for better treatment effect with sorafenib. Both symptomatic and asymptomatic patients at entry had improved PFS following treatment with sorafenib.

Conclusions: Maximum tumor size, number of lesions, thyroglobulin levels at baseline and geographic regions were prognostic for longer PFS in RAI-rDTC patients. Individual tumor size ≥1.5 cm and lung-only metastases were predictive for better treatment effect with sorafenib. Patients appeared to benefit from sorafenib treatment irrespective of disease-related symptoms at baseline. Thus, based on these post hoc exploratory analyses, patients with progressive RAI-rDTC and maximum tumor size <1.5 cm appear to have a good prognosis and may be candidates for “watch and wait” before initiating sorafenib.

Disclosure: Ralf Paschke: Advisory Role: Bayer Pharma AG, Eisai; Financing of Scientific Research: Bayer Pharma AG, Eisai Marcia Brose: Advisory Role: Bayer HealthCare Pharmaceuticals, Exelixis, Onyx Pharmaceuticals; Financing of Scientific Research: Bayer HealthCare Pharmaceuticals; Expert Testimony: Bayer HealthCare Pharmaceuticals, Exelixis, Eisai, Novartis, Roche/Genentech.

V94 - Characterization of tumor-associated B cell subsets in head and neck squamous cell carcinoma

Lechner A.1,2, Rothschild S.I.2,3, Schlößer H.A.2,4, Thelen M.2, Shimabukuro-Vornhagen A.2, Beutner D.1, von Bergwelt-Baildon M.S.2

1Uniklinik Köln, Klinik und Poliklinik für Hals-, Nasen- und Ohrenheilkunde, Köln, Germany, 2Uniklinik Köln, Klinik I für Innere Medizin, Cologne Interventional Immunology, Köln, Germany, 3Universitätsspital Basel, Medizinische Onkologie, Basel, Switzerland, 4Uniklinik Köln, Klinik und Poliklinik für Allgemein-, Viszeral- und Tumorchirurgie, Köln, Germany

Background: Recent evidence suggests that B cells can both promote and inhibit the development and progression of tumors. Tumor-infiltrating B cells have been described in a large variety of solid tumors including head and neck squamous cell carcinoma (HNSCC). However, there has not been a detailed phenotypic characterization of the different B cell subsets in HNSCC. The aim of this study was to characterize the composition of the B cell infiltrates in HNSCC in order to gain further insight into their role.

Methods: In this study we characterized B cell subsets in primary tumors (n = 30), blood (n = 30) and non-cancerous mucosa (n = 10) of 30 previously untreated patients diagnosed with HNSCC. Furthermore, peripheral blood from samples of age-matched healthy donors (n = 10) was included as control. B cell subsets were analyzed by 10-color flow cytometry.

Results: Tumor-infiltrating B cells (CD19+) can be found in the majority of HNSCC samples. HNSCC tissue contained a significantly higher percentage of activated B cells (CD19+/CD20+/CD86+) compared to healthy controls, non-cancerous mucosa of HNSCC patients and peripheral blood of HNSCC patients. The percentage of memory B cells (CD19+/CD20+/IgD-/CD27+) was significantly higher in tumor tissue than in the peripheral blood of cancer patients. Moreover, we found a higher rate of regulatory B cells (CD24high/CD38high or TIM-1+) in tumor tissue compared to peripheral blood of healthy controls and tumor patients as well as non-cancerous mucosa.

Conclusion: B cells constitute a significant proportion of the immune infiltrate in HNSCC. The B cell infiltrate of HNSCC is characterized by an accumulation of CD86+ activated B cells and memory B cells which is suggestive of an antigen recognizing phenotype and a specific effector B-cell response. However, tumors are also infiltrated by a considerable number of regulatory B cells.

Disclosure: No conflict of interest disclosed.

Wissenschaftliches Symposium

Hoch-Risiko Smouldering Myelom (SMM): Pro und Kontra unverzügliche Behandlung

V96 - Treatment should immediately be started in every patient with high risk smouldering myeloma (SMM)

Engelhardt M.1, Wäsch R.1

1University of Freiburg Medical Center, Department of Hematology, Oncology & Stem Cell Transplantation, Freiburg, Germany

SMM is an asymptomatic clonal plasma cell (PC) disorder and bridges MGUS and MM, based on higher levels of circulating monoclonal immunoglobulin (IG) and bone marrow (BM) plasmocytosis (PCosis) without end-organ damage. Until the PETHEMA-GEM group reported fewer MM-related events and better overall survival (OS) among patients (pts) with high-risk (HR)-SMM treated with lenalidomide-dexamethasone, prior studies failed to show improved OS, assessing bisphosphonates, melphalan, prednisone or thalidomide as compared to observation. Risk factors (RF) of HR-SMM have now been defined (Table 1), and a subset of ultra-HR pts are reclassified by the International Myeloma Working Group as MM, based on biomarkers that identify pts with a >80% risk of progression.

Commonly defined RF for SMM progression are the number of bone lesions, e.g. abnormal whole-body (wb) magnetic resonance imaging (MRI [or other sensitive imaging]), size of the M-spike, IgA isotype, abnormal serum free light chain (sFLC) ratio, degree of PCosis, circulation and aberrancy of BMPCs and immunoparesis. Ultra-HR-SMM is now redefined as MM with BM PCosis >60%, sFLC ratio >100 and >1 focal lesion on wb-MRI. The number of these redefined pts is small (10-15%), but important to unravel, because their risk of progression is substantial (≥80% within 2 years). Pts with HR FISH cytogenetics (CG) (del17p, t(4;14), gain 1q21) are not yet considered for early treatment, because groups are heterogeneous and other RF than CG deemed to weight higher.

Albeit pts with ultra HR-SMM are now considered as MM and treated as such (Table 1), concerns do exist that earlier therapy may increase the risk of selecting resistant clones, induce side effects and costs. Therefore, the even more accurate identification of pts, who most likely benefit from interventions, needs to be performed, and clinical judgement and careful discussion of pros and cons of treatment initiation undertaken. For the greater majority of SMM pts, the standard of care remains observation until development of symptomatic MM occurs. The latter group remains to be observed every 3-6 months and is encouraged to participate in clinical trials.


Disclosure: No conflict of interest disclosed.

V97 - Treatment should immediately be started in every patients with high risk smoldering myeloma (SMM): No - defending the NO position

Ludwig H.1

1Wilhelminenspital, Wilhelminen Cancer Research Institute, Vienna, Austria

Introduction: Based on improved PFS and OS obtained in one small study (Metaeos MV et al., NEJM 2013) with immediate start of Lenalidomide/Dexamethasone therapy as compared to starting treatment only at time of clinical evident progression, the IMWG recommended immediate therapy in all patients with newly defined high risk SMM ( ≥ 1 focal lesions in MRI, or FLC ratio >100 and involved FL >100mg/dl).

Methods: Review of available evidence (Medline, ASH, EHA, IMWG, ASCO meetings) and risk calculation

Findings: Hillengass L. et al. showed that ≥1 focal lesion of >5 mm size detected in MRI associate with a 70% rate of progression, a finding which was confirmed by another small study (Kastridis E. et al.) showing similar results (69% rate of progression). The FLC ratio risk model (FLC ratio >100 and involved FLC concentration of >100 mg/dl) was associated with an 82% risk of progression after 2 years (Larsen JT et al., 2013), but when the risk for progression was assessed in another patient cohort using a FLC ratio >100 alone, only 53% of patients with SMM had progressed after 2 years (Waxmann AJ et al., 2015).

Risk modelling: These data show a 2 year rate of progression between 70 and 82% using either one of the risk models. Assuming a linear rate of transformation into active myeloma, the progression rate will be 20%, 40%, 60, and 80% after 6, 12, 18, and 24 months, and the rate of unnecessary treatment will amount to 80%, 60%, 40%, and 20% at the given time points, indicating significant overtreatment. Calculating the area under the curve patients require treatment and above the curve when patients do not require therapy shows a ratio of 40% to 60%, meaning that 60% of the time therapy was administered without medical need.

Conclusion: The recommendation to start therapy in every patient with high risk SMM, given the toxicity and burden of therapy, does not accord with one of the most important principles in medicine, namely ‘primum non nocere'. Modelling shows that roughly 60% of the treatment will be administered to patients without clinical signs of active myeloma if the recommendations are followed. This reality must be presented to and discussed with the individual patient to enable him to make an informed decision. In case the patient is unable or unwilling to decide treatment should be withheld, but the patient needs to adhere to careful regular control examinations in short intervals by an experienced myeloma specialist.

Disclosure: No conflict of interest disclosed.


Sarkom - Vergleich der Behandlungskonzepte

V104 - Osteosarcoma: A step backwards for poor responders? What´s next?

Bielack S.1, Cooperative Osteosarkom-Studiengruppe COSS

1Klinikum Stuttgart - Olgahospital, Päd. Onkologie, Hämatologie, Immunologie, Stuttgart, Germany

For the past 35 years, the treatment of osteosarcoma has been based upon surgery of the primary tumor and, if present, primary metastases plus pre- and postoperative chemotherapy. Many international groups would consider a combination of high-dose methotrexate, adriamycin (doxorubicin), and cisplatin (MAP) as standard. It has long been proven that the extent of histologic response of the primary tumor to upfront preoperative chemotherapy is correlated with survival expectancies, and patients whose tumors still contain more than 10% viable cells after standard induction carry an unfavorable prognosis. Until recently, no prospective randomized trial evaluating whether postoperative treatment modifications might improve the prognosis of these poor responders had been performed. The European and American Osteosarcoma Study EURAMOS1 (NCT00134030) filled this void. Poor responders to 10 weeks of MAP induction were randomized to either continue with MAP postoperatively until protocol week 29 or to receive postoperative MAP augmented by high-dose ifosfamide and etoposide until protocol week 40 (MAPIE). A total of 2.260 patients from 17 countries were registered into EURAMOS1, 1.059 of these were considered poor responders, of whom 618 participated in the poor response randomization (Marina et al., CTOS 2014, paper 032). At the predefined analysis of the primary endpoint, event-free survival (EFS), higher acute toxicities were observed with the augmented, experimental MAIE chemotherapy, as was an increased incidence of secondary acute leukemias. There was no evidence that the more toxic MAPIE was more efficacious than MAP, and EFS rates did not differ between arms. The four multinational osteosarcoma groups collaborating in EURAMOS1 therefore recommend to not use MAPIE in case of poor response to MAP, but to continue with MAP regardless of histologic response - the standard treatment which will still result in cure in well over 40% even of poor responders. Meanwhile, the search for novel treatment options continues. As of yet, no substance which might be available for large-scale, multi-institutional, multi-national phase III osteosarcoma trials has been identified. Current international efforts aim to describe genomic and immunologic targets for novel treatment approaches and to translate these findings from bench to bedside. The EURAMOS consortium intends to initiate their next large joint front-line trial once an agent suitable for such an approach is identified

Disclosure: Stefan Bielack: Advisory Role: Bayer, Celgene, IDM/Takeda Millennium, Lilly, Merck & Co., Roche.

V105 - Position papers and treatment recommendations for chordomas and desmoid tumours

Kasper B.1

1Mannheim University Medical Center, Interdisciplinary Tumor Center, Sarcoma Unit, Mannheim, Germany

Desmoid-type fibromatosis (DF) is a rare disease characterized by a monoclonal, fibroblastic proliferation and a variable and often unpredictable clinical course. The incidence is less than 3% of all soft tissue sarcomas. Due to the rarity and the long natural history of the disease, the level of evidence as it is available for more common tumour types is currently beyond reach for DF. Especially in the context of systemic treatment options, only few phase II trials are available, and most published data arises from retrospective case reports or small series. There is no published phase III randomized clinical study on DF and no established or evidence-based treatment approach is available as of today. With this background information in mind, a European joint effort was initiated bringing together sarcoma experts from the European Organisation for Research and Treatment of Cancer (EORTC) / Soft Tissue and Bone Sarcoma Group (STBSG) with patients and patient advocates from Sarcoma Patients EuroNet (SPAEN), and has led to the development of consensus recommendations based on patients' and professionals' expertise - for the first time ever in this disease. As a surrogate, a position paper has been published (Kasper B et al.: Eur J Cancer 2015;51:127-136).

Chordomas are very rare malignant bone tumours that have had a shortage of effective treatments for a long time. Although new treatment options are available for both local and metastatic disease, the degree of uncertainty in selecting the most appropriate treatment remains high and their adoption in daily life remains inconsistent. As a blueprint of the above described procedure, a consensus meeting on chordoma management was hosted by the European Society for Medical Oncology (ESMO) within their regular meeting updating the clinical practice guidelines on sarcomas. This meeting included more than 40 chordoma experts from several disciplines with the contribution and sponsorship of the Chordoma Foundation, a global patient advocacy group. The consensus reached at that meeting has recently been published in a position paper (Stacchiotti S et al.: Lancet Oncol 2015;16:e71-83).

The bullet points of the position papers and this innovative process of developing treatment recommendations and a therapeutic algorithm in rare diseases will be presented.

Disclosure: No conflict of interest disclosed.


Hodentumore - bestes Management

V106 - Management seminoma Stage IIB: Radiation therapy

Papachristofilou A.1

1Universitätsspital Basel, Radioonkologie, Basel, Switzerland

Optimal management of seminoma stage IIB remains a controversial issue. Standard radiation therapy of the paraaortal and ipsilateral pelvic lymphatic pathways leads to excellent local control, at the cost of distant recurrence and potential long term sequlae of large volume irradiation. In the past years innovative techniques have lowered the risk of excessive radiation dose to the kidneys and bowel, yet the problem of disease recurrence beyond the irradiated volume remains a challenge.

Innovative combined treatment concepts encompassing reduced volume as well as possibly reduced dose radiation therapy and less intensive systemic treatment may provide the optimal future treatment option.

Disclosure: No conflict of interest disclosed.

V107 - Management of seminoma stage IIB: Chemotherapy is the treatment of choice

Honecker F.1

1Tumor- und Brustzentrum ZeTuP, St. Gallen, Switzerland

Seminoma stage II B is defined by a N2 status diagnosed by abdominal CT scan. Definition of N2 is either a retroperitoneal lymph node mass of more than 2 but less than 5 cm, or multiple lymph nodes, any one mass extending between 2 and 5 cm in greatest dimension. The optimal management of stage IIB is controversial, and data from randomized trials is lacking. Treatment recommendations are as follows: the European Association of Urology (EAU) currently considers chemotherapy (CT) with 3 × PEB or 4 × PE as an alternative to radiotherapy (RT) with 36 Gy (in 2 Gy fractions) to paraaortic and ipsilateral iliac lymph nodes [1], whereas the European Society of Medical Oncology (ESMO) clinical practice guidelines from 2013 consider CT as the optimal choice [2]. Notably, when CT is considered, it must be performed according to the current standard, as use of 4 cycles of single agent carboplatin was insufficient and resulted in higher relapse rates [3]. Retroperitoneal lymph node dissection is highly experimental, and should not be carried out outside clinical trials. A systematic review and meta-analysis evaluated data from studies using either RT or CT in stage IIA/B [4]. 4 prospective and 9 retrospective studies, none of them randomized, and with a total of 890 patients (607 RT, 283 CT) were identified. Pooled relapse rate (RR) was 0.11 for RT (CI 0.08-0.14) and 0.08 for CT (CI 0.01-0.15). For assessment of overall mortality, 8 studies were available, and outcome was excellent with pooled mortality rates of 0.02 for RT (CI < 0.01-0.04) and 0.01 for CT (CI < 0.01-0.02). Despite statistical equivalence between RT and CT, a trend towards lower relapse rates and lower incidences of late toxicities, especially less GI toxicity and secondary cancers was observed. This is “suggesting an erosion of equipoise in favor of CT for stage IIB seminoma” [4]. An ongoing trial is assessing the efficacy and safety of combination therapy with single-agent carboplatin plus involved field RT in this situation. The trial is open in multiple centers in Germany and Switzerland, and referral of all suitable patients with stage II seminoma is strongly recommended.


1 Albers P et al.: Uroweb 2015, http://uroweb.org. 2 Oldenburg J et al.: Ann Oncol 2013;24(Suppl 6):vi125-vi132. 3 Krege S et al.: Ann Oncol 2006;17:276-280. 4 Giannatempo P et al.: Ann Oncol 2015;26:657-668.

Disclosure: No conflict of interest disclosed.

V108 - Management Seminoma IIB - pro retroperitoneal surgery

Albers P.1

1Universitätsklinikum Düsseldorf, Klinik für Urologie, Düsseldorf, Germany

Es gibt zur retroperitonealen Resektion der Metastasen im klinischen Stadium II der Seminome als singuläre Therapieoption kaum zitierbare Daten. In einer Serie aus Magdeburg, in der über 63 Patienten mit retroperitonealer Lymphadenektomie (RLA) bei Seminomen berichtet wird, fällt eine “in-field“ Rezidivrate von 9.5% (6/63) und eine Gesamtrezidivrate für Seminome ab Stadium IIC von 58% (7/12) auf. Dabei wurden 23 der 63 Patienten nach der RLA adjuvant therapiert. Die Operation wurde daher ab 1985 nur noch zurückhaltend durchgeführt. In der aktuellen Literatur findet man entsprechend nur noch Fallberichte (n = 4, Hu, 2015; n = 4, Mezvrishvili, 2006).

Pathophysiologisch gibt es keine Gründe, Seminome bezüglich operativer Strategien vor Chemotherapie grundsätzlich anders zu behandeln als Nicht-Seminome. Auch Seminome folgen in ihrer Metastasierung sehr lange den Lymphabflußwegen und metastasieren erst spät hämatogen. Aus klinischer Sicht gibt es aber folgende Gründe, metastasierte Seminome im Stadium IIA und IIB nach Ablatio testis primär retroperitoneal zu operieren:

marker-negative Tumoren mit unklarer Histologie, die einer Punktion nicht zugänglich sind

imperative Indikationen bzw. Kontraindikationen zur Chemotherapie wie z. B. Niereninsuffizienz (autosomal dominante Zystennieren)

Spät-Toxizität der Chemotherapie

Spät-Toxizität der Radiotherapie

Es müssen grundsätzlich drei Szenarien unterschieden werden: a) primär diagnostizierte Tumoren im Stadium II, b) Rezidiv unter Überwachung c) Rezidiv nach 1 × Carboplatin adjuvant. Die Standardtherapie dieser Stadien wäre 3 × BEP Chemotherapie oder Radiatio mit 36 Gy. Beide Therapieoptionen haben eine messbare Langzeittoxizität und werden undifferenziert und unabhängig von Größe, Lokalisation sowie Markerhöhe eingesetzt. Besonders Rezidive unter Überwachung und nach Carboplatin werden aber üblicherweise sehr früh in der Nachsorge erkannt (Tumordurchmesser oft <1.5 cm), so dass (insbesondere bei marker-negativen Rezidiven) im Rahmen prospektiver Studien eine primäre Resektion dieser kleinvolumigen Metastasen als singuläre Therapieoption zur Vermeidung von Spät-Toxizitäten geprüft werden sollte. Im Rezidivfall bleiben die bisherigen Primärtherapien als Rezidivtherapien erhalten.

Disclosure: No conflict of interest disclosed.

V110 - Palliative treatment of testicular germ cell cancer

Oechsle K.1

1Universitätsklinikum Hamburg-Eppendorf, Onkologisches Zentrum, II. Medizinische Klinik und Poliklinik, Hamburg, Germany

Introduction: Although 70-80% of germ cell cancer patients can be cured even in metastatic disease, prognosis remains poor in patients with multiple relapses during or after cisplatin-based chemotherapy or relapse after high-dose chemotherapy.

Methods: A systematic overview on current literature on treatment of patients with refractory or multiply relapsed germ cell cancer including recommendations for daily clinical practice will be presented.

Results: Single agent activity with response rates of about 20% could be observed for orally applied etopside, paclitaxel, gemcitabine and, oxaliplatin (Einhorn 1991, Motzer 1994, Bokemeyer 1999, Kollmannsberger 2002). Until today, the evaluation of targeted therapy only revealed some marginal activity for sunitinib with remission rates of 10% (Feldman 2010, Oechsle/Honecker 2011) and thalidomide with marker, but nor morphologic remissions in about 30% (Rick 2006). None of these agents showed significant impact on overall survival when applied as monotherapy.

Various studies on combination chemotherapy regimen combining two of the three agents, paclitaxel, gemcitabine, and oxaliplatin, achieved response rates of about 20-40% with overall survival times of 6-8 months (e.g. Hinton 2001, De Giorgi 2006, Einhorn 2007, Kollmannsberger 2004). In the following, a further German study evaluated the triple combination chemotherapy with gemcitabine, oxaliplatin plus paclitaxel (GOP-regimen) and demonstrated the highest response rate for the present of 51% in these heavily pretreated patients: 78% after high-dose chemotherapy (Bokemeyer 2008). After additional secondary surgery, complete remission was achieved in 20% of patients. Further follow-up confirmed long term survival >2 years in 17% of these patients (Oechsle 2011). Similar effects were present in a retrospective study on the triple combination chemotherapy of cisplatin, gemcitabine, and paclitaxel resulting in a 2 year overall survival rate of 30% after complete secondary surgery. (Necchi 2014)

Conclusions: Results of single agent treatments are still disappointing and only of palliative nature in patients with multiply relapsed or refractory germ cell cancer. The evaluation of new substances remains if enormous relevance for these patients. Triple combination with the GOP regimen could result in long term survival in about 15-20% of these patients if complete secondary surgery of all residual lesion could be achieved.

Disclosure: No conflict of interest disclosed.

Freier Vortrag

MDS klinisch I

V114 - Prognostic significance of rare single abnormalities in Myelodysplastic Syndromes

Schanz J.1, Tüchler H.2, Solé F.3, Sanz G.4, Garcia-Manero G.5, LeBeau M.6, Bennett J.M.7, Slovak M.L.8, Fenaux P.9, Malcovati L.10, Cazzola M.10, Pfeilstöcker M.11, Valent P.12, Ohyashiki K.13, Levis A.14, Sekeres M.15, Tauro S.16, Magalhaes S.17, Van de Loosdrecht A.18, Cermak J.19, Lübbert M.20, Stauder R.21, Germing U.22, Greenberg P.23, Haase D.24, On behalf of the International Work Group for Prognosis in MDS (IWG-PM)

1Universitätsmedizin Göttingen, Klinik für Hämatologie und Medizinische Onkologie, Göttingen, Germany, 2Hanusch Hospital, Boltzmann Institute for Leukemia Research, Vienna, Austria, 3Institut de Recerca contra la Leucèmia Josep Carreras, Barcelona, Spain, 4Hospital Universitario La Fe, Valencia, Spain, 5The University of Texas, MD Anderson Cancer Center, Houston, United States, 6University of Chicago Comprehensive Cancer Research Center, Chicago, United States, 7James P. Wilmot Cancer Institute, University of Rochester Medical Center, Rochester, United States, 8Quest Diagnostics Nichols Institute, Chantilly, United States, 9Hôpital Avicenne, Assistance Publique-Hôpitaux de Paris (AP-HP)/University Paris XIII, Bobigny, France, 10Department of Hematology Oncology, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy, 11Hanusch Hospital and L. Boltzmann Cluster Oncology, Vienna, Austria, 12Medical University of Vienna, Vienna, Austria, 13First Department of Internal Medicine, Tokyo Medical University, Tokyo, Japan, 14Fondazione Italiana Sindromi Mielodisplastiche c/o SS Antonio e Biagio Hospital, Allessandria, Italy, 15Cleveland Clinic Taussig Cancer Institute, Cleveland, United States, 16University of Dundee, Dundee, United Kingdom, 17Federal University of Ceara, Fortaleza, Brazil, 18Department of Hematology, VU University Medical Center, Amsterdam, Netherlands, 19Institute of Hematology and Blood Transfusion, Prague, Czech Republic, 20University of Freiburg Medical Center, Freiburg, Germany, 21University of Innsbruck, Innsbruck, Austria, 22Heinrich-Heine University Hospital, Düsseldorf, Germany, 23Division of Hematology, Stanford University Cancer Center, Stanford, United States, 24Universitätsmedizin, Göttingen, Germany

Introduction: The IPSS (Greenberg et al., 1997) and its recently published revision, the IPSS-R (Greenberg et al., 2012), defines the standard in risk stratification of patients with Myelodysplastic Syndromes (MDS). However, due to its low frequency, the prognostic impact of rare abnormalities remains unclear as yet. These abnormalities are coalesced in one group in the IPSS and IPSS-R. The main goal of the study presented here was to analyse the type, frequency and prognosis of rare single abnormalities in a large cohort of patients with primary, untreated MDS.

Methods: To date, we identified 382 pts. with rare single abnormalities derived from the international database of the IWG-PM, containing more than 7000 patients. A specific group was defined as having at least n = 5 cases with the same abnormality. The participating centers (in numerically order) were: Spain (n = 109; 28.5%), MD Anderson Cancer Center (73; 19.1%), IMRAW (43; 11.3%), France (34; 8.9%), Pavia (25; 6.5%), Vienna (21; 5.5%), Duesseldorf (16; 4.2%), Japan (14; 3.7%), Italy (11; 2.9%), Cleveland (10; 2.6%), Dundee (9; 2.4%), Brazil (8; 2.1%), Netherlands (5; 1.3%), Czech (2, 0.5%), Freiburg (1; 0.3%) and Innsbruck (1; 0.3%). For all abnormalities detected, the median overall- (OS) and AML-free survival (AFS) was calculated.

Results: Rare single abnormalities detected were: +1/+1q/dup1q (n = 9; 2.4%), t(1;var)(12; 3.1%), del(1q)(5; 1.3%), der(1;7)(19; 5.0%), del(3p)(7; 1.8%), t(3;var)(5; 1.3%), t(5q)(8; 2.1%), del(6q)(7; 1.8%); -9/del(9q)(15; 3.9%), t(11q23)(6; 1.6%), +13 (6; 1.6%), -13/del(13q)(20; 5.2%), +14(5; 1.3%), del(16q)(7; 1.8%), del(17p)(7; 1.8%), -19 (5; 1.3%), +21 (16; 4.2), -21/del(21q)(6; 1.6%), -X (7; 1.8%), and +mar (12; 3.1%). The remaining 198 patients (51.8%) showed rare abnormalities occurring in less than 5 patients. The median overall survival as well as the AML-free survival in each category will be presented in detail. Furthermore, a proposal to integrate these abnormalities in the cytogenetic module of the IPSS-R will be presented.

Conclusions: Due to their low frequency, rare abnormalities in MDS can only detected based on large, international cooperation projects. In the following study, the identification and prognostic impact of rare abnormalities, uninfluenced by therapy or additional abnormalities was comprehensively analyzed. The results will lead to a further refinement of the cytogenetic prognostic classification in patients with MDS.

Disclosure: No conflict of interest disclosed.

V115 - CMML and treatment with azacitidine in a routine care setting in Germany: Data from the second planned interim analysis of the non-interventional study “observational study on treatment patterns with VIDAZA®

Illmer T.1, Prange-Krex G.2, Lück A.3, Lollert A.4, Schwarzer A.5, Bruch H.-R.6, Bachinger A.7, Weiligmann C.8, Steudel C.8, Platzbecker U.9

1Onkologische Gemeinschaftspraxis, Dresden, Germany, 2Gemeinschaftspraxis Mohm und Prange-Krex, Dresden, Germany, 3Zentrum für Onkologie und Urologie in Rostock, Rostock, Germany, 4Gemeinschaftspraxis Dr. Neise & Dr. Lollert, Krefeld, Germany, 5Gemeinschaftspraxis Hämatologie/Onkologie, Leipzig, Germany, 6Praxis Bonn - Schwerpunktpraxis, Bonn, Germany, 7ClinAssess GmbH, Leverkusen, Germany, 8Celgene GmbH, München, Germany, 9Universitätsklinikum Carl Gustav Carus, Medizinische Klinik und Poliklinik I, Dresden, Germany

Introduction: Approval of azacitidine (AZA) in chronic myelomonocytic leukemia (CMML) is based on the pivotal AZA001 trial. This prospectively, randomized trial, which has shown prolonged overall survival (OS) in MDS patients (pts), included few CMML pts (n = 11). Additionally, the approval in CMML is restricted to the non-myeloproliferative disease. Here we present data from CMML pts out of the second interim analysis of a non-interventional multicentre trial in Germany. The aim of the study was to prospectively collect data on the clinical usage of AZA in the real world setting.

Methods: At the time of data cutoff (04/16/2015) 440 pts of 660 planned pts were enrolled in 66 German sites and observed for a maximum of 12 months with an optional one year of follow-up. A comprehensive set of data on demography, disease characteristics, AZA treatment regimen including dosage and duration, reasons for discontinuation and other factors were collected and analyzed. Survival data were estimated according to Kaplan Meier methodology.

Results: At the time of this analysis, 44 pts diagnosed with CMML had been recruited and were evaluable. The majority were elderly pts (median age of 75.5 years [range 57-89]) and treated with the approved schedule for 7 days (75.0%). Five pts (11.4%) received AZA in second or further line of treatment. Overall response rate (CR, CR-BM, PR or HI) was 36.4% (n = 16) and an additional 25.0% (n = 11) achieved SD while in 38.6% (n = 17) no response assessment data were available at the time of data cutoff. Six pts (13.6%) achieved a complete or partial cytogenetic response. One year survival rate was 59.1% (95% CI; 43.2-73.7). Treatment beyond one year was documented in one quarter of pts (n = 11). Main reasons for treatment discontinuation were death (27.3%, n = 9), pt request (15.2%, n = 5) and multiple reasons (24.2%, n = 8). Half of all pts experienced serious adverse events (54.5%, n = 24). The most common serious adverse events during treatment were general disorders and administration site conditions (18.2%, n = 8).

Conclusion: This report about 44 CMML pts treated with AZA in a routine care setting in Germany supports evidence that therapy with AZA is effective and safe in CMML.

Disclosure: Thomas Illmer: No conflict of interest disclosed. Uwe Platzbecker: Financing of Scientific Research: Celgene GmbH.

V116 - Progression of chromosomal aberrations correlates with an inferior prognosis in MDS patients with regard to survival and AML progression rates

Neukirchen J.1, Nolting A.-C.1, Hildebrandt B.2, Kobbe G.1, Haas R.1, Germing U.1

1Uniklinik Düsseldorf, Department of Hematology, Oncology and Clinical Immunology, Heinrich-Heine-University, Düsseldorf, Germany, 2Uniklinik Düsseldorf, Department of Human Genetics, Heinrich-Heine-University, Düsseldorf, Germany

Introduction: For MDS patients, several chromosomal abnormalities have been shown to have a significant influence of prognosis. The IPSS as well as in the recently introduced IPSS-R, uses the karyotypes as important factors for evaluation of prognosis. We analyzed the outcome of MDS undergoing subsequent karyotyping during the course of the disease and correlated the results with survival and AML development.

Methods: We screened the Duesseldorf MDS registry for those patients that revealed at least two chromosomal analyses during the course of the disease. We then categorized three groups: a) no change of karyotype b) worsening of karyotype, and c) normalization of karyotype and correlated the results with regard to overall survival as well of AML development.

Results: We identified in total 595 patients with at least two chromosomal analyses during the course of the disease. The median age was 63 years (17-86 years). 6.2% were categorized as IPSS-R very low, 28.8% as low, 25.5% as intermediate, 19.3% as high and 20.2% as very high. The median number of available karyotypes for each patient was 3 (range 2-25). Looking at all patients with at least two karyotype analyses available, 102 (17.1%) of the patients showed no change of the karyotypes, 292 (49.1%) progressed to a prognostic inferior karyotype and 201 (33.8%) showed a more favorable karyotype during the course of the disease as a result of treatment response. Looking at the AML evolution, time to AML progression was significantly shorter in patients that showed a progression of the karyotype (40 months vs. not reached, p < 0.0001). Accordingly, the survival was with 33 vs. 46 months shorter in the group of patients with karyotype progression (p = 0.03). After exclusion patients that received allogeneic stem cell transplant, the results were comparable.

Conclusion: Based on a large database of patients who were karyotyped as least two times, we could demonstrate that in MDS patients clonal evolution is associated with a worse prognosis with regard to survival and AML progression and that the normalization of cytogenetic alterations as a sign of treatment response correlates with a longer survival. These results underline the importance of karyotype analysis for a better prognostication of MDS patients, especially with regard to treatment decision.

Disclosure: Judith Neukirchen: No conflict of interest disclosed. Ulrich Germing: Expert Testimony: Celgene, Norvartis, Amgen, Jansen Cilag.

V117 - Evolution of chromosome 7 material loss in myeloid malignancies

Shirneshan K.1, Braulke F.1, Schanz J.1, Haase D.1

1Universitätsmedizin Göttingen, Klinik für Hämatologie und Medizinische Onkologie, Göttingen, Germany

Introduction: Loss of chromosome 7 (chr. 7) material (as mon(7) or del(7q)) is one of the most frequent chromosomal abnormalities in patients with acute myelogenous leukemia (AML) and myelodysplastic syndrome (MDS).It has already been demonstrated that the prognostic impact of isolated 7q deletion has to be regarded as prognostically more favorable compared to isolated mon(7) (Corduba et al., 2012, Schanz et al., 2012) and not as unfavorable as described in the IPSS.

Method: In 2% of MDS patients we observed a coexistence of mon(7) and del(7q)by FISH analysis of CD34+ peripheral blood cells. Two patients showed an increase of mon (7) clone size parallel to a decline in 7q- clone size during MDS progression.

Results: Here we report three additional MDS cases showing several clones with different chr.7 aberrations. The first case (A.M.): A 72-year old man with suspected MDS. Analysis of bone marrow (BM) revealed MDS RA. Banding analysis of BM cells showed three clones with different chr. 7 abnormalities: del(7q) in 22%, r(7) in 22% and mon(7) in 11% of the metaphases. Second case (A.L.): A 62 year old woman with suspected hematological malignancy. Karyotyping of BM cells revealed: A del(7q) in 12% and r(7) in 15% of metaphases. Third Case (K.B.): A 64 years old MDS Patient with the following karyotype abnormalities: del(7q) in 3%, mon(7) plus only a “naked” centromere of chr. 7 in 47% and complete mon(7) in 20% of BM cells.

Conclusion: These results could be a hint that chr. 7 can be prone to karyotype evolution during MDS progression. Due to the existence of the third clone with r(7) and a clone with remaining centromere 7 we assume that r(7) is a transitional stage of karyotype evolution between deletion of 7q and complete monosomy 7 which might result from loss of telomeres in 7q- cells promoting loss of further chromosomal material ending up in a r(7). It is known that ring chromosomes themselves are prone to be lost during cell division. And so finally the entire process ends with complete mon(7). By specific telomere FISH analysis we could confirm the deletion of telomeres in r(7). Our hypothesis could also explain why del(7q) has a better prognosis than mon(7): Del(7q) is occurring earlier during the course of the disease since it is the first step in mon(7)-evolution.

To prove this hypothesis, more data from patients with karyotype evolution involving chr. 7 will be accumulated and further specific examinations will be performed.

Disclosure: No conflict of interest disclosed.

V118 - Decitabine given at the 3-day schedule in older patients with refractory anemia with excess blasts in transformation (RAEBt), or low blast count acute myeloid leukemia according to WHO criteria

Becker H.1, Suciu S.2, Rüter B.H.1, Platzbecker U.3, Giagounidis A.4, Selleslag D.5, Labar B.6, Germing U.7, Salih H.R.8, Muus P.9, Pflüger K.-H.10, Hagemeijer A.11, Schaefer H.-E.1, Baron F.12, Ganser A.13, Aul C.4, de Witte T.9, Wijermans P.14, Lübbert M.1, EORTC Leukemia Cooperative Group, German MDS Study Group

1Medical Center - University of Freiburg, Freiburg, Germany, 2EORTC Headquarters, Brussels, Belgium, 3University of Dresden Carl Gustav Carus, Dresden, Germany, 4Marienhospital Düsseldorf, Düsseldorf, Germany, 5AZ St-Jan, Brugge, Belgium, 6University Hospital Center Rebro, Zagreb, Croatia, 7University Hospital, Düsseldorf, Germany, 8Eberhard Karls University, Tübingen, Germany, 9Radboud University Medical Center, Nijmegen, Netherlands, 10DIAKO, Bremen, Germany, 11University of Leuven, Leuven, Belgium, 12C.H.U. Sart-Tilman, Liège, Belgium, 13Hannover Medical School, Hannover, Germany, 14Haga Ziekenhuis, The Hague, Netherlands

Treatment of older patients with high-risk myelodysplastic syndromes (MDS) or acute myeloid leukemia (AML) is challenging. In the randomized phase III study 06011 of the EORTC Leukemia Group and German MDS Study Group, we compared decitabine (15 mg/m2 every 8 hours for 3 days, every 6 weeks) with best supportive care (BSC) in patients ≥60 years with MDS by FAB criteria and not eligible for intensive therapy (Lübbert et al.: J Clin Oncol 2011;29:1987-96). In the present analysis, we investigate trial 06011 for the efficacy of decitabine in refractory anemia with excess blasts in transformation (RAEBt), and, after regrouping the study cohort according to WHO criteria, in AML (WHO) with low blast counts.

In RAEBt, rates of best response in the decitabine arm (n = 40) were as follows: complete (CR) or partial remission (PR), 15%; hematologic improvement (HI), 15%; progressive disease (PD), 30%. In the BSC arm (n = 35), no patient achieved CR, PR or HI; 80% had PD. The favorable responses in the decitabine arm translated into a significantly longer progression-free survival (PFS; HR 0.30, 95%-CI 0.18-0.51; P < 0.001) and, by trend, longer overall survival (OS; HR 0.68, 95%-CI 0.42-1.11, P = 0.12) than in the BSC arm. Post-progression treatment, particularly allogeneic hematopoietic stem cell transplantation (alloHSCT), may influence survival, and the number of patients receiving alloHSCT was imbalanced between the treatment groups. Thus, we censored OS at the time of alloHSCT and observed that the difference between the treatment groups increased (HR 0.63, 95%-CI 0.38-1.04; P = 0.067). This was particularly true for patients aged 60-74 years (HR 0.48, 95%-CI 0.26-0.89; P = 0.018), i.e. potential candidates for alloHSCT after reduced-intensity conditioning. The impact of decitabine on the outcome in RAEBt patients differed from that in patients with refractory anemia with excess blasts (RAEB). After regrouping the study cohort according to the nowadays more widely used WHO classification, AML (WHO) patients in the decitabine arm (n = 27) also had a longer PFS (P = 0.008), and their median OS was 9.8 months compared to 5.9 months in the BSC arm (n = 23).

In conclusion, 3-day decitabine is active in MDS and/or AML with 5-30% blood or 20-30% marrow blasts and may be used as bridge to alloHSCT. The 3-day schedule is not feasible in the outpatient setting, but our data point to it as a valid treatment option for hospitalized patients with above features.

Disclosure: No conflict of interest disclosed.

V119 - Underestimation of 12p-deletion in myelodysplastic syndromes? Results from a German diagnostic study in comparison with an international control group

Braulke F.1, Müller-Thomas C.2, Götze K.2, Platzbecker U.3, Germing U.4, Hofmann W.-K.5, Giagounides A.A.N.6, Lübbert M.7, Greenberg P.L.8, Bennett J.M.9, Sole F.10, Slovak M.L.11, Ohyashiki K.12, Le Beau M.M.13, Tüchler H.14, Pfeilstöcker M.15, Hildebrandt B.16, Aul C.17, Stauder R.18, Valent P.19, Fonatsch C.20, Bacher U.1, Trümper L.1, Haase D.1, Schanz J.1

1Universitätsmedizin Göttingen, Hämatologie und medizinische Onkologie, Göttingen, Germany, 2Technische Universität München, Hämatologie u. Onkologie, München, Germany, 3Uniklinik Dresden, Hämatologie u. Onkologie, Dresden, Germany, 4Universität, Düsseldorf, Germany, 5Universitätskrankenhaus, Mannheim, Germany, 6Marienhospital, Düsseldorf, Germany, 7Universität Freiburg - Klinik für Innere Medizin I, Freiburg, Germany, 8Stanford University Cancer Center, Stanford, United States, 9University of Rochester Medical Center, Rochester, United States, 10Institut de Recerca Contra la Leukemia Josep Carreras, Badalona, Spain, 11Sonora Quest Laboratories, Phoenix, United States, 12Tokyo Medical University, Tokyo, Japan, 13University of Chicago, Chicago, United States, 14Hanusch Hospital, Boltzmann Institute for Leukemia Research, Vienna, Austria, 15Hanusch Hospital and L. Boltzmann Cluster Oncology, Vienna, Austria, 16Universität Düsseldorf, Humangenetik, Düsseldorf, Germany, 17St. Johannes Hospital, Duisburg, Germany, 18Innsbruck Medical University, Innsbruck, Austria, 19Division of Hematology and Hemostaseology and Ludwig Boltzmann Cluster Oncology, Medical University of Vienna, Vienna, Austria, 20Medical University of Vienna, Vienna, Austria

Introduction: In myelodysplastic syndromes (MDS), 12p-deletion (del(12p)) as a sole anomaly is a characteristic, but rare abnormality (0.6-5% at initial diagnosis). It is described to occur as an additional aberration in up to 4% in the context of clonal evolution during the course of the disease. According to current scoring systems, del(12p) is associated with a good to intermediate prognosis as a single aberration. Del(12p) is usually a small interstitial deletion on the short arm of chromosome 12 with a common deleted region between 12p12.2-12p13.1 and ETV6/TEL as the affected gene.

Methods: We present the results of a systematic del(12p) testing in our German prospective diagnostic study (clinicaltrials.gov: NCT01355913) on 351 MDS patients in whom CD34+ peripheral blood (PB) cells were analysed by sequential Fluorescence-in-situ-Hybridization (FISH) analyses using a large probe panel including TEL/AML1 (Abbott®, Germany). We compared the CD34+PB-FISH results with the results of chromosome banding analyses (CBA) of bone marrow (BM) metaphases of 2902 previously published MDS patients.

Results: Out of 351 MDS patients analysed by CD34+PB-FISH, 28 (7.6%) showed a del(12p) by CD34+PB FISH: 23 at initial screening at the time of study entry, 5 additional patients showed del(12p) later during the course of the study in the context of a karyotype evolution. Most of the del(12p) patients had primary MDS (85.7%), 4 patients (14.3%) were diagnosed as t-MDS. The clone sizes measured by CD34+PB-FISH and CBA differed significantly for both, non-complex and complex karyotypes involving del(12p), the median clone size of a del(12p) was 65% by CD34+PB-FISH and 100% by CBA of BM metaphases. In contrast, only 45 MDS patients (1.6%) of the CBA control group showed a 12p-deletion. Del(12p) is often associated with other aberrations (93% by CD34+ PB-FISH vs 60% by CBA).

Conclusions: The probability to detect del(12p) was significantly higher by FISH as compared to CBA (7.6% by CD34+ PB-FISH vs 1.6% by CBA, p < 0.001). The detection rate of del(12p) could be increased by repeated FISH analyses in a patient over time. It is important for MDS patients to distinguish between a normal karyotype, a single anomaly and double or complex aberrations for the prediction of prognosis according to international prognostic scoring systems. Therefore, we recommend including a probe for del(12p) to common FISH probe panels in MDS.

Disclosure: Friederike Braulke: Expert Testimony: Die CD34+FISH-Studie wurde von Celgene und Novartis unterstützt.; Immaterial Conflict of Interests: nein. Julie Schanz: Employment or Leadership Position: nein.; Advisory Role: nein.; Stock Ownership: nein.; Honoraria: nein.; Financing of Scientific Research: nein.; Expert Testimony: Die CD34+FISH-Studie wurde von Celgene und Novartis unterstützt.; Other Financial Relationships: nein.; Immaterial Conflict of Interests: nein.


Palliativmedizin Entscheidungsprozesse und -kriterien

V121 - What can the nurse contribute to the decision making process?

Monteverde S.1,2

1Berner Fachhochschule, Fachbereich Gesundheit, Bern, Switzerland, 2Universität Zürich, Institut für Biomedizinische Ethik und Medizingeschichte, Zürich, Switzerland

As in other domains of medical and nursing practice, also in oncology the goal of collaborative decision making is the patient's health and wellbeing. This is achieved by means of the integration of patient preferences, available scientific evidence and a sensitivity for the relationship between burden and benefit. Patients are encouraged and requested to exercise their right to self-determination and give their consent to the oncological treatment that best fits their values as well as the medical condition. Nevertheless, evidence shows that the threat of the disease and the uncertainties and risks inherent to the medical treatment render the patient and the family particularly vulnerable. Patients and families have multiple needs of information, counseling, assistance and encouragement in making meaningful treatment decisions. These needs are best met when each of the professions involved in the delivery of oncological care plays an active role within the process of shared decision making.

Therefore, interprofessional collaboration between doctors and nurses not only covers medical treatment, monitoring and reporting, but also decision making. Due to their continuous presence and proximity to the patients and families, nurses play a crucial role in helping patients to make meaningful decisions. This task is germane to the sphere of co-responsibility shared by doctors and of nurses (see for example Swiss Health Insurance Law (KVG, KLV Art. 7).

However, communication research within oncology suggests that the perception of patients, doctors and nurses with regard to the quality of communicative interaction, the understanding of the information delivered, the empathic response and the weighting of risks and benefits of a proposed treatment differs significantly. Similarly, doctors and nurses have different perceptions of the degree to which the other profession is involved in decision making processes. Of particular importance are those decisions that involve the redirection of care towards palliation and quality of life when suffering increases and no benefit is seen for the patient in continuing a curative approach. In redirecting care towards palliation and advance care planning, careful communication, clarification of patient values and readiness for interprofessional negotiation are indispensable. Here, clinical ethics tools like case deliberations, ethics consultations or family conferences are important means to identify meaningful goals of care.

Disclosure: No conflict of interest disclosed.

Freier Vortrag

Allogene Transplantation klinisch III

V124 - The combined impact of amino acid polymorphism at HLA-DPB1 on T cell alloreactivity defines a functional distance between patient and donor independently predictive of survival after 10/10 matched unrelated HSCT for AML, ALL and MDS

Crivello P.1, Beelen D.W.2, Heinold A.3, Heinemann F.M.3, Rebmann V.3, Lindemann M.3, Ottinger H.2, Horn P.A.3, Fleischhauer K.1

1Institut für Zelltherapeutische Forschung, Universitätsklinikum, Essen, Germany, 2Klinik für Knochenmarktransplantation, Universitätsklinikum, Essen, Germany, 3Institut für Transfusionsmedizin, Universitätsklinikum, Essen, Germany

Introduction: We have previously experimentally determined the differential impact of individual amino acid polymorphisms on the alloreactive T cell response to HLA-DPB1*09:01, translated these findings into numeric “functional distance” (FD) scores for all HLA-DPB1 alleles, and showed that these FD scores correlate with T cell epitope groups determining non-permissive mismatches in unrelated HSCT (Fleischhauer et al.: Lancet Oncol 2012; Crivello et al.: Biol Blood Marrow Transplant 2014).

Methods: A cohort of 379 consecutive patients (pts) (age median 56 yrs [18-73 yrs]), who underwent high-resolution HLA-A,B,C,DRB1,DQB1 matched, but DPB1 mismatched unrelated HSCT for AML (n = 272 [72%]), ALL (n = 58 [15%]), or MDS (n = 49 [13%]) at the University Hospital of Essen were included in the analysis. FD scores of DPB1 alleles were calculated as previously described, and the ΔFD scores for each transplant were calculated as the absolute number of [FDpatient-FDdonor]. Receiver Operator Curves (ROC) were constructed to calculate the best cut-off values for the endpoint of overall survival (OS).

Results: With a median follow-up of 4 yrs for surviving pts, the 5-yrs OS for pts in early disease stages (n = 158 [42%]) was 56% (95%-confidence interval [CI] 48-64%). For pts in advanced disease stages (n = 221 [58%]) this estimate was 38% (95%-CI 31-45%) (p < 0.0005). ΔFD score distribution ranged from 0.01 to 7.46 with a median of 2.12. ROC analysis indicated stratification into 2 subgroups with ΔFD scores < = 2.665 (n = 252 [66%]) and >2.665 (n = 127 [44%]) as the best predictor. In these subgroups, the 5-yrs OS were 51% (95%-CI 44-57%) and 37% (95%-CI 28-46%), respectively (p < 0.008). In multivariate analysis, independent predictors of OS were time-dependent aGvHD (p < 0.0002) and cGvHD (p < 0.0001), the use of anti-thymocyte globulin (p < 0.0001), patient age (p < 0.004), and the stratified ΔFD score (p < 0.03).

Discussion: In this monocentric cohort, the stratified DPB1 ΔFD score was a significant independent risk factor for OS after 10/10 matched unrelated HSCT, possibly owing to the strength of the alloimmune response to mismatched HLA-DPB1 in this setting, which was also reflected by an independently increased non-relapse mortality in pts with ΔFD scores >2.665. These findings could pave the way for the identification of non-permissive DPB1 mismatches in unrelated HSCT based on an experimentally defined numerical weighting system for amino acid sequence polymorphism.

Disclosure: No conflict of interest disclosed.

V125 - Clinical phase I peptide vaccination trial with a CMVpp65-derived peptide can prevent and clear CMVpp65 antigenemia after allogeneic hematopoietic stem cell transplantation

Greiner J.1,2, Schmitt A.3, Hofmann S.1, Götz M.1, Michels B.3, Hückelhoven A.3, Maccari B.4, Lindner D.3, Wang L.3, Wuchter P.3, Mertens T.1, Döhner H.1, Ho A.3, Bunjes D.1, Dreger P.3, Kuball J.5, Schrezenmeier H.4, Schauwecker P.4, Wiesneth M.4, Schmitt M.3

1University of Ulm, Department of Internal Medicine III, Ulm, Germany, 2Diakonie Hospital Stuttgart, Department of Internal Medicine, Stuttgart, Germany, 3University of Heidelberg, Department of Internal Medicine V, Heidelberg, Germany, 4German Red Cross Blood Transfusion Service Baden-Württemberg-Hessen, Institute of Clinical Transfusion Medicine and Immunogenetics, Ulm, Germany, 5University Medical Center Utrecht, Department of Hematology, Utrecht, Netherlands

We performed a phase I clinical peptide vaccination trial using an immunodominant viral epitope peptide derived from the phosphoprotein 65 of cytomegalovirus (CMVpp65) for patients after allogeneic stem cell transplantation with a high risk of CMV reactivation, disease and mortality.

The CMV phosphoprotein 65 derived nonamer peptide NLVATVPMV has been well characterized as immunogenic. Therefore, we designed a vaccine with 300 microgram of the peptide in an oil-in-water emulsion (Montanide™).

Ten CMV-seropositive patients after allogeneic stem cell transplantation received 4 vaccines s.c. at a biweekly interval. Patients were monitored for clinical course as well as CMVpp65 antigenemia. CD8+, Tregs and gamma-delta T cells were analyzed by multi-color flow cytometry. ELISpot assays for Interferon and granzyme B and tetramer assays were performed and correlated to clinical parameters.

Peptide vaccination was well tolerated and no drug-related serious adverse events were detected.

Eight of nine patients with CMVpp65 antigenemia cleared the CMV after four vaccinations and are still free from viremia until now. One patient received prophylactic vaccination and did not develop viremia. One patient with CMV reactivation showed no clinical response and resented persisting CMV antigenemia.

An increase in frequency of CMV specific T-cells was detected in eight patients and frequency of regulatory T-cells was found in a part of the patients. Moreover, the frequency of gamma-delta T cells increased in several but not in all patients with immune and clinical response after vaccination.

Taken together, administration of CMVpp65 peptide vaccination for patients after allogeneic stem cell transplantation at high risk for CMV reactivation was save, well tolerated and clinically encouraging.

Disclosure: No conflict of interest disclosed.

V126 - Prognostic impact of aberrant MN1 expression in patients with Acute Myeloid Leukemia (AML) undergoing allogeneic stem cell transplantation in complete remission after Non-Myeloablative Conditioning (NMA-SCT)

Jentzsch M.1, Bill M.1, Leiblein S.1, Schubert K.1, Wildenberger K.1, Bergmann U.1, Pleß M.1, Weidner H.1, Knyrim M.1, Grimm J.1, Lange T.1,2, Cross M.1, Franke G.-N.1, Pönisch W.1, Vucinic V.1, Behre G.1, Niederwieser D.1, Schwind S.1

1Universitätsklinik Leipzig, Hämatologie und internistische Onkologie, Leipzig, Germany, 2Asklepios Klinik Weißenfels, Hämatologie und intern. Onkologie, Weißenfels, Germany

Introduction: Meningeoma-1 (MN1) is highly expressed in primitive hematopoietic cells & has dismal prognostic impact in AML patients (pts) treated with chemotherapy. The prognostic significance in AML pts receiving NMA-SCT with a therapeutic effect relying on immunologic graft-versus-leukemia (GvL) effects remains unknown.

Patients and methods: We analysed 98 pts (median age 64.1 years [y], range 38.2-75.3y) who received NMA-SCT (90 mg/m2 Fludarabine & 2Gy total body irradiation) in complete remission with pre-treatment bone marrow (BM) material available. Donors were human leukocyte (HLA) matched related (18%) or HLA-matched (57%) or mismatched (24%) unrelated. Medical Research Council (MRC) classification was: 4% favourable, 71% intermediate & 24% adverse. Common surface marker expression was assessed by flow cytometry. Mutation status of NPM1, CEBPA & FLT3-ITD & BAALC & ERG expression were evaluated. MN1 expression was normalized to ABL & the 25th percentile of the normalized gene expression was used to define high & low expressers. Median follow up was 4.3y.

Results: High pre-treatment MN1 expression associated with higher CD34+ (P < 0.001), higher CD34+/CD38- (P < 0.001) & by trend higher CD117+ (P = 0.08) BM cell burden, lower white blood count (P = 0.01), lower platelets (P = 0.02), abnormal karyotype (P = 0.003), NPM1 wild type (P < 0.001), high BAALC (P < 0.001) & high ERG expression (P < 0.001).

High MN1 expressers had higher cumulative incidence of relapse (CIR) by trend in the entire group (P = 0.07, Figure 1A) & a significantly higher CIR in the MRC intermediate risk subgroup (P = 0.04, Figure 1C). However, this did not translate into shorter overall survival (OS) for the whole or the MRC intermediate risk cohort (P = 0.29 & P = 0.45 respectively, Fig. 1. B&D).


Conclusion: AML pts with high MN1 expression had a higher burden of immature surface marker i.e. CD34 & CD117 & more CD34+/CD38- cells, which harbour the leukemic stem cell population. High MN1 expressers had higher CIR after NMA-SCT, but in contrast to pts after chemotherapy no shorter OS, suggesting that the dismal impact of high MN1 may in part be diminished by the GvL effect after NMA-SCT.

Disclosure: No conflict of interest disclosed.

V127 - T-cell-replete HLA-haploidentical transplantation using post-transplantation high-dose cyclophosphamide post-transplantation in high-risk and advanced ALL: Feasibilty and early outcome

Zoellner A.-K.1, Fritsch S.1, Prevalsek D.1, Köhnke T.1, Hubmann M.1, Lippl S.1, Kruger S.1, Hellmuth J.1, Mumm F.1, Barlow S.1, Ledderose G.1, Subklewe M.1, Spiekermann K.1, Hiddemann W.1, Albert M.2, Schmid C.3, Hausmann A.1,4, Tischer J.1

1Ludwig-Maximilians-University Hospital of Munich-Grosshadern, Department III of Internal Medicine, Hematopoietic Stem Cell Transplantation, Munich, Germany, 2Ludwig-Maximilians-University of Munich, Dr. von Haunersches Kinderspital, Munich, Germany, 3Department of Internal Medicine II, Klinikum Augsburg, Augsburg, Germany, 4Department I of Internal Medicine, Klinikum München-Schwabing, Munich, Germany

Haematopoietic stem cell transplantation (allo-HSCT) is a potential curative treatment option for patients suffering from high-risk ALL, but less true for patients with advanced disease. However, not in all of our patients a suitable HLA-matched donor could be identified in time. To evaluate the outcome of T-cell-replete (TCR) HLA-haploidentical haematopoietic stem cell transplantation (haplo-HSCT) utilizing high-dose cyclophosphamide post-transplantation in the context of intensification of conditioning in patients with high-risk, relapsed and refractory ALL, we retrospectively analysed the course of 25 patients (B-ALL n = 23, T-ALL n = 2) transplanted between 2010 and 2015 in four German transplant centres. Disease was advanced in 19 patients, including 11 patients with relapse after a first allo-HSCT. Conditioning was TBI-based in 14 patients and consisted of fludarabine and cyclophosphamide (CY) plus either 12 Gy TBI in all remission patients or 8 Gy TBI in all patients being older than 55 years; all infants (n = 3) received 12 Gy TBI plus etoposide. In adults with relapse after a first allogeneic transplantation conditioning was drug-based: fludarabine, CY plus treosulfan (3 x 10-12g/m2) and etoposide. Post-grafting immunosuppression was high-dose CY, tacrolimus and MMF in all patients. 23/25 patients engrafted, 2 patients died early in aplasia. No primary graft rejection was observed. Acute GvHD grade II-IV occurred in 5 patients (20%), while 6 patients (24%) suffered from mostly mild to moderate chronic GvHD. Severe toxicity (grade III-IV) was observed in 11 patients (44%); most commonly mucositis (36%), transient elevation of transaminases (32%) and diarrhoea (32%). Kidney failure requiring haemodialysis occurred in 3 patients. CMV reactivated in 8 patients and EBV in 3 patients while no patient developed CMV disease or PTLD. Proven invasive aspergillosis was diagnosed in 2 patients. One-year non-relapse mortality was 12%. After a median follow up of 16.6 months, estimated one-year overall survival and relapse-free survival was 74% and 48%, respectively. In summary, intensification of conditioning in the setting of TCR haplo-HSCT using PTCY is well tolerated with low NRM in patients with high-risk and relapsed ALL, while providing an effective anti-leukemic activity in advanced disease. Thus, we suggest that donor availability can be expanded in patients with high-risk and advanced ALL who lack a conventional donor or suffer from aggressive disease.

Disclosure: No conflict of interest disclosed.

V128 - Predictive markers for CD34+ mobilization in healthy peripheral blood stem cell donors

Körper S.1, Hauber D.1, Fürst D.1, Reinhardt P.1, Schauweker P.1, Mytilineos J.1, Schwarz K.1, Bunjes D.2, Wiesneth M.1, Schrezenmeier H.1

1Universitätsklinik Ulm/DRK Butspendedienst Baden-Württemberg-Hessen, IKT-Ulm, Ulm, Germany, 2Universitätsklinik Ulm, Medizinische Klinik III, Ulm, Germany

A few prognostic factors for stem cell mobilization have been established. Body weight (BW) and male sex (MS) are accepted predictors for good mobilization. In search for other prognostic factors we performed a retrospective analysis of the records of 316 healthy stem cell donors.

A single platform assay was used to determine the CD34+ cell count/µl in peripheral blood after G-CSF mobilization (aim: 40 µg G-CSF per kg BW, divided in 8 dosages). The concentration of CD34+ cells was correlated with different parameters in all 316 donors. Spearmans Test was used to determine significance (P ≤ 0.01).

The study population included 104 women and 212 men. The median age was 38.5 years. The median CD34+ cell concentration after G-CSF was 63/µl (7 to 280/µl; non-Gaussian distribution). For further analysis groups of poor (PM) and good mobilizers (GM) were defined by the 10. (28/µl) and 90. (132/µl) percentiles.

Colony assays differed between PM and GM with BFUE of 49 (18-228) and CFU-GEMM of 10 (4-36) in PM and BFUE of 226 (83-3048) and CFU-GEMM of 47 (16-487) in GM (unit: /105 viable cells).

BMI, BW and ALT, and cholinesterase (obtained before G-CSF administration) showed a weak but significant correlation with CD34+ mobilization in men and women. To compare the ultrasound examinations of the liver the presence of steatosis hepatis was examined in the PM and GM. Steatosis of grade 1 or 2 was present in 6% of PM and 41% of GM, a grade 2 steatosis was not seen in PM but in 9% of GM.

Further factors were shown in the table.


Our data confirmed MS and BW as positive predictors for CD34+ mobilization. As expected colony growth and stem cell yield was tightly connected to mobilization. Interestingly higher counts of red blood cells and eosinophils (prior to G-CSF) were correlated with better mobilization in an univariate analysis. Indicators of obesity like elevated liver enzymes and steatosis hepatis were also linked to better CD34+ mobilization.

Disclosure: Sixten Körper: Employment or Leadership Position: Abteilungsleiter Im IKT Hubert Schrezenmeier: Employment or Leadership Position: Ärztlicher Direktor am IKT; Expert Testimony: Beobachtunrgsstudie zur Anwednung von Filgrastim Hexal.

V129 - Donor lymphocyte infusion (DLI) in patients with hematological malignancies and the detection of cytotoxic T-cell (CTL) responses against several leukemia-associated-antigens (LAA)

Hofmann S.1, Götz M.1, Herbst C.1, Schneider V.1, Wiesneth M.2, Döhner H.1, Bunjes D.1, Greiner J.1

1University of Ulm, Department of Internal Medicine III, Ulm, Germany, 2University of Ulm, Institute of Clinical Transfusion Medicine, Ulm, Germany

T-cell responses play a central role in maintaining remission and prolonging overall-survival in patients with hematologic malignancies after allo-HSCT and delayed DLI. The role of LAA and mHag has to be elucidated although there is evidence that the graft-versus-leukemia-effect observed after DLI is based on CTL-mediated immunity which is reactive against mHag and LAA.

In this study, we analysed peripheral blood and serum samples of 10 patients (pts) with AML (3 pts), CML (2 pts), Multiple Myeloma (3 pts) and CLL (2 pts) in the course of allo-HSCT and before and after DLI for specific CTL responses against several LAA and established a cytokine profile. 3 pts received preemptive DLI and 8 pts received therapeutic DLI. 5 pts underwent a single DLI, 4 pts received a second and 1 pt a third DLI. Epitopes derived from PRAME, NPM1mut, RHAMM, WT-1 and other LAA were tested. Immune reactions of CTL were measured in ELISpot assays (INF-gamma and granzyme B). The corresponding cytokine profile of several cytokines was analysed for further interpretation of the clinical data.

In general, 4 of the 10 pts who underwent DLI developed GvHD with 3 pts receiving ongoing immunosuppression. In all patients we could detect a CTL response against at least one of the tested LAA in the course of DLI. The two pts with AML with NPM1-mutation (NPM1mut) who received preemptive DLI in molecular relapse showed a polyspecific CTL-response against several epitopes including those derived from the mutated region of NPM1, Survivin and WT1. CTL responses against RHAMM were detected in AML, Multiple Myeloma, CML and CLL. All tested patients with Multiple Myeloma and CLL showed CTL responses against NYESO1. WT1 was detected in AML and Multiple Myeloma in a high frequency. The pattern of CTL-responses changed during the course, interestingly, patients with GvHD and immunosuppression also showed CTL positivity against several LAA.

Immunoinhibitory as well as immunostimulating cytokines were analysed. Cytokine profiles differed in accordance to the clinical situation including immunosuppression, GvHD, time point after DLI application and disease status.

Taken together, CTL responses against several LAA were detected in the course of DLI, this might support the hypothesis that these LAAs are a therapeutic target in remission induction in patients with allogeneic transplantation. Patients who developed GvHD after DLI showed a combination of LAA positivity.

Disclosure: No conflict of interest disclosed.

Freier Vortrag

Immuntherapie I

V130 - Targeting HLA.A2-restricted MDM2 (81-88) epitope as a new antigen for TCR gene therapy of multiple myeloma

Amann E.1, Antunes E.1, Jacobi B.1, Theobald M.1,2,3, Echchannaoui H.1,3

1University Medical Center of the Johannes Gutenberg University Mainz, Third Medical Department (Hematology, Oncology and Pneumology), Mainz, Germany, 2Johannes Gutenberg University Mainz, Research Center Immunology (FZI), Mainz, Germany, 3German Cancer Research Center (DKFZ), German Cancer Consortium (DKTK), Frankfurt/Mainz, Germany

Background: Adoptive transfer of T cells retrovirally equipped with tumor-associated antigen (TAA)-specific T cell receptor (TCR) has shown efficacy in cancer patients. The human homologous of the murine double-minute 2 protein (MDM2) TAA is overexpressed in a variety of human tumors, including soft tissue sarcoma, multiple myeloma (MM), glioblastoma and melanoma. MDM2 protein overexpression is particularly observed in invasive and metastatic melanoma and is associated with enhanced survival of MM cells. We have generated and optimized an HLA.A2.1-restricted CD8-dependent MDM2 (81-88)-specific TCR isolated from a high-avidity murine CTL clone derived from CD8xA2Kb transgenic mice. Our aim is to target MM in adoptive immunotherapy using human T cells genetically equipped with MDM2-specific TCR.

Methods: The nucleotide sequence of the MDM2-TCR-construct was codon-optimized (opt.) and TCR chains cloned in a bicistronic retroviral vector containing the self-cleaving 2A virus-derived peptide. We also introduced an additional inter-chain disulfide bond (cys.) between TCR α and β constant domains to prevent mixed heterodimers formation. Human T cells were retrovirally transduced with TAA-specific TCR and expression levels were analyzed by flow cytometry. HLA.A2.1 MM cell lines were screened for MDM2 expression by Western Blot. In a xenograft mouse model MM cell line was injected subcutaneously in the flank of NOD-scid IL2R gamma chainnull (NSG) mice and human T cells transduced with cys.opt. TCR or mock vector were injected intravenously when the tumors were palpable.

Results: We observed a strong correlation between MDM2 expression level and MDM2 TCR-mediated lysis in vitro. In a xenograft mouse model we could observe a prolonged overall survival and tumor-infiltrating T cells in mice which received MDM2-specific TCR transduced T cells compared to Mock-treated group. Interestingly, ex vivo tumor cells exhibit a down-regulation of MDM2 expression and concomitantly an up-regulation of p53 expression which results in a lower recognition of these tumor cells by the MDM2-specific TCR. Preliminary data showed a potent lysis of ex vivo tumor cells by MDM2/p53 dual TCR-modified T cells as compared to single TCR-equipped T cells.

Conclusion: Our data show that MDM2-specifc TCR can target MM in vitro and in vivo. Using MDM2- and p53-specific dual TCR transduced T cells may represent a novel approach to circumvent tumor escape mechanisms like antigen down-regulation.

Disclosure: No conflict of interest disclosed.

V131 - Next-generation dendritic cell vaccination in postremission therapy of AML: results of a clinical phase I trial

Lichtenegger F.S.1,2, Schnorfeil F.M.1,2, Brüggemann M.3, Moosmann A.4, Köhnke T.1, Bücklein V.1, Altmann T.1, Wagner B.5, Hiddemann W.1, Bigalke I.6, Kvalheim G.6, Subklewe M.1,2

1Klinikum der Universität München, Department of Internal Medicine III, Munich, Germany, 2Helmholtz-Zentrum München, Clinical Cooperation Group Immunotherapy, Munich, Germany, 3University Hospital Schleswig-Holstein, Department of Hematology, Kiel, Germany, 4Helmholtz-Zentrum München, Clinical Cooperation Group Immunooncology, Munich, Germany, 5Klinikum der Universität München, Department of Transfusion Medicine, Cellular Therapeutics and Hemostaseology, Munich, Germany, 6The Norwegian Radium Hospital, Oslo University Hospital, Department of Cellular Therapy, Oslo, Norway

Introduction: Postremission therapy for acute myeloid leukemia (AML) is critical for elimination of minimal residual disease (MRD). In patients not eligible for allogeneic stem cell transplantation, therapeutic vaccination with autologous, tumor antigen-loaded dendritic cells (DCs) is a promising treatment strategy to induce anti-leukemic immune responses and to eradicate chemorefractory cells. We have developed a GMP-compliant 3-day protocol including a TLR7/8 agonist to differentiate monocytes of intensively pretreated AML patients into next-generation DCs.

Methods: We are conducting a proof-of-concept phase I/II (6+14 patients) clinical trial using next-generation DCs as postremission therapy for AML of non-favorable genetic risk in CR after intensive induction therapy (NCT01734304). DCs are loaded with RNA encoding the leukemia-associated antigens WT1 and PRAME as well as CMVpp65 as adjuvant and surrogate antigen. Patients are vaccinated intradermally with 5×106 DCs of each antigen species up to 10 times within 26 weeks. The primary endpoint of the trial is feasibility and safety of the vaccination. Secondary endpoints are immunological responses and disease control.

Results: 9 patients have been enrolled into the trial. With one dropout, DCs of sufficient number and quality were generated from leukapheresis in 7/8 cases. DC analysis showed a positive costimulatory profile, secretion of IL-12p70, migration towards a chemokine gradient, antigen expression and specific T cell activation in vitro. 4 patients have completed the vaccination schedule; the 5th patient has received 7/10 vaccinations. We observed delayed-type hypersensitivity (DTH) responses at the vaccination site in 5/5 patients, accompanied by slight erythema and indurations at the injection site, but no grade III/IV toxicities. Multimer analysis revealed the induction of antigen-specific T cell responses in 3/3 patients tested. We detected an increase of WT1-specific T cells in one patient and strong inductions of CMVpp65-specific T cells in two CMV-seronegative patients. TCR repertoire analysis by next-generation sequencing revealed an enrichment of particular clonotypes at DTH sites. In an individual treatment attempt, we treated one patient with impending relapse with a combination of DC vaccination and 5-azacytidine, resulting in MRD conversion.

Conclusion: Vaccination with next-generation DCs in AML is feasible and safe, and induces anti-leukemia-specific immune responses in vivo.

Disclosure: No conflict of interest disclosed.

V132 - Selective engagement of the RIG-I pathway synergizes with checkpoint blockade in cancer immunotherapy

Heidegger S.1, Kreppel D.1, Bscheider M.1,2, Wintges A.1, Bek S.1, Schmickl M.1, Fischer J.C.1, Lin C.-C.1, Peschel C.1, Haas T.1, Poeck H.1,3

1Klinikum rechts der Isar der Technischen Universität München, III. Medizinische Klinik, München, Germany, 2Stanford University School of Medicine, Department of Pathology, Stanford, United States, 3Memorial Sloan-Kettering Cancer Center, Departments of Immunology and Medicine, and Cell Biology, New York, United States

Introduction: Targeting of regulatory T cell receptors such as CTLA-4 has been shown to enhance anti-tumor immune responses. Given that checkpoint inhibitors can augment T-cell reactivity against tumor neoantigens, we aimed at improving this response with a vaccination protocol that combines selective engagement of the nucleic acid-sensing pattern recognition receptor RIG-I, antigen and CTLA-4 blockade.

Methods: We used wild-type and genetically-altered mice as well as an established combination of antibiotics to characterize the molecular pathways and external influences affecting our combinatorial approach on a whole organism level. Specifically, mice were vaccinated with ovalbumin (OVA) and the specific RIG-I ligand 5'-triphosphat RNA (3pRNA) together with anti-CTLA-4 antibody. The frequency of OVA-specific T cells and specific lysis of target cells was determined. For tumor challenge, identically treated mice were injected iv with B16.OVA melanoma cells and the number of lung metastases was analyzed. By using 3pRNA/OVA-stimulated dendritic cells (DCs) isolated from gene-deficient mice, we assessed the relevance of these respective pathways for cross-presentation and subsequent cross-priming of co-cultured cytotoxic T cells.

Results: We found that vaccination and RIG-I ligation strongly synergized with CTLA-4 blockade to induce expansion and activation of antigen-specific CD8+ T cells and potent anti-tumor immunity (Figure 1). Cross-priming of cytotoxic T cells as well as anti-tumor immunity required the adapter protein MAVS and type I interferon (IFN) signaling and were mediated by DCs. In addition, the benefit of the combined immunization with anti-CTLA-4 was reduced by systemic antibiotics pointing to the prerequisite of an intact commensal microbiota in this context.

Conclusion: We established a novel combinatorial strategy to generate peptide-specific CD8+ T cells and anti-tumor immunity and characterized the respective signaling pathways, cell types and environmental influences responsible for this beneficial response. The combination of RIG-I-induced type I IFN and checkpoint blockade has the potential to enhance personalized anti-cancer vaccines.


Disclosure: No conflict of interest disclosed.

V133 - Triplebody 33-3-19 eliminates biphenotypic (CD19 plus CD33) leukemia cells selectively

Roskopf C.C.1, Braciak T.A.1, Fenn N.2, Kobold S.3, Jacob U.4, Fey G.H.4, Hopfner K.-P.2, Oduncu F.S.1

1Klinikum der Universität München, Medizinische Klinik und Poliklinik IV, Hämatologie/Onkologie, München, Germany, 2Genzentrum, München, Germany, 3Klinikum der Universität München, Medizinische Klinik und Poliklinik IV, Klinische Pharmakologie, München, Germany, 4SpectraMab GmbH, München, Germany

Introduction: Targeted immune-interventions with antibodies and antibody derivatives have shown improved tumor-cell specificity for cancer therapy. However, as target antigens are tumor-associated (TAA) but not tumor-specific, the elimination of healthy non-cancerous cells may occur. To improve tumor cell-selectivity we envisioned a “dual-targeting” approach: Antibody derivatives in the triplebody format simultaneously target two different TAAs on the surface of the same tumor cell and engage immune effector cells such as NK or T cells via a trigger antigen.

Methods: Triplebody 33-3-19 was constructed from single chain variable fragment (scFv) building blocks, expressed recombinantly in suspension-adapted FreeStyle™ 293-F cells and purified. Protein properties and cytotoxic activity were analyzed using molecular biology and flow cytometric techniques as well as redirected lysis assays.

Results: 33-3-19 binds the B lymphoid marker CD19 and the myeloid marker CD33 specifically; an antigen combination that is exclusively present on biphenotypic leukemia cells. Furthermore, it recruits T cells via the CD3 epsilon chain and activates these effectors as efficiently as the recently described mono-targeting triplebody 19-3-19 (Roskopf et al. 2014). The triplebody induced specific lysis of established myeloid (MOLM13, THP-1) and B-lymphoid cell lines (SEM, Raji) and of primary patient cells at low picomolar concentrations. EC50 values ranged from 0.1 to 28 pM. At 1 nM concentration 33-3-19 induced preferential lysis of double- over single-positive leukemia cells at a ratio of 2-to-1 in a target cell mixture: (CD19 plus CD33) double-positive BV173 target cells were eliminated more efficiently in the presence of 33-3-19 plus T cells than the CD19 single-positive SEM target cells, in spite of comparable overall target antigen densities (50,000-60,000/cell). 19-3-19 or a bispecific T cell engager mixture of 19-3 and 33-3, however, led to similar elimination of both cell lines at 1nM.

Conclusions: Dual-targeting agents such as triplebodies may be used for an efficient and more selective immune-intervention in cancer and may induce the elimination of leukemia-initiating cells.

Disclosure: No conflict of interest disclosed.

V134 - Combining immune checkpoint inhibition with the CD33 BiTE® antibody construct AMG 330 to enhance T cell mediated lysis of primary AML cells

Krupka C.1,2, Kufer P.3, Kischel R.3, Zugmaier G.3, Köhnke T.1,2, Lichtenegger F.S.1,2, Altmann T.1,2, Spiekermann K.1,4, Vick B.4,5, Jeremias I.4,5, Hiddemann W.1,4, Subklewe M.1,2,4

1Klinikum der Universität München, Department of Internal Medicine III, München, Germany, 2Helmholtz Institute Munich, Clinical Co-operation Group Immunotherapy, München, Germany, 3AMGEN Research (Munich) GmbH, München, Germany, 4German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ), Heidelberg, Germany, 5Helmholtz Zentrum München, Department of Gene Vectors, München, Germany

Introduction: BiTE® antibody constructs are very efficient in inducing T-cell activation and secretion of proinflammatory cytokines (Krupka, Blood 2014).

The proinflammatory cytokine milieu favors the upregulation of immune checkpoint molecules on target cells. In this study the relevance of immune checkpoint molecules in CD33/CD3 BiTE® antibody construct (AMG 330) mediated lysis of primary AML cells was evaluated.

Methods: AMG 330 mediated lysis of primary AML cells was analyzed in a long-term culture system. The assay system was validated for proliferation of AML cells, antigen expression pattern, clonal evolution and persistence of leukemia initiating cells (LIC). The expression level of checkpoint molecules was assessed by flow cytometry (MFI ratio).

Results: Ex vivo analysis of primary AML cells in long-term cultures showed consistent AML cell proliferation or persistence (n = 38), stable antigen expression pattern (n = 19) and clonal stability (n = 2) throughout at least 28 days of culture.

In vivo xenotransplantation experiments confirmed the propagation of LICs in the system (n = 4).

Although not constitutively expressed on primary AML cells (n = 123), PD-L1 was strongly upregulated upon the addition of AMG 330 to the ex vivo cytotoxicity experiments (p < 0.0001; n = 27). This phenomenon was cytokine-mediated as the sole addition of IFN-γ and TNF-α also induced expression (n = 6).

Furthermore, we observed a significant upregualtion of PD-1 on activated T cells (MFI ratio: control vs AMG 330 3.1 : 12.9, p = 0.0002, n = 18).

In ex vivo cytotoxicity experiments blocking of the PD-1/PD-L1 interaction significantly enhanced AMG 330 mediated lysis efficacy (median lysis: AMG 330 58% vs + α-PD-1/PD-L1 75%, n = 9, p = 0.03) which was accompanied by a significant increase in T cell proliferation and a markedly increase in IFN-γ secretion. The effect was most prominent in cultures with low E:T ratios (PD-1: lysis E:T 1:1: AMG 330 100% vs + α-PD-1 100%; lysis E:T 1:9: AMG 330 90% vs + α-PD-1 98%; PD-L1: lysis E:T 1:1: AMG 330 95% vs + α-PD-L1 100;% lysis E:T 1:5: AMG 330 83% vs + α-PD-L1 97%). Our work thus provides evidence that PD-L1 upregulation is a relevant mechanism of primary AML cells to escape cytokine-mediated immune responses. We demonstrate that AMG 330 mediated cytotoxicity is enhanced by blockade of inhibitory receptors on AML cells.

Conclusion: Our results support the use of combinatorial approaches of BiTE® antibodies with immune checkpoint blockade.

Disclosure: Christina Krupka: Expert Testimony: Die Studie wurde teilweise durch AMGEN Research (Munich) finanziert Marion Subklewe: Expert Testimony: Die Studie wurde teilweise durch AMGEN Research (Munich) finanziert.

V135 - Modulation of glycolytic metabolism promotes in vitro generation of EBV-reactive and redirected virus-specific human CD8+ cytotoxic T lymphocytes with stem-cell-memory and central-memory properties

Weber I.1, Bhatti A.1, Krebs L.1, Theobald M.1, Hartwig U.F.1

1III. Medizinische Klinik, Universitätsmedizin der Johannes Gutenberg-Universität Mainz, Hämatologie, Internistische Onkologie & Pneumologie, Mainz, Germany

Introduction: Adoptive T cell therapy has advanced as a powerful tool to treat opportunistic viral infections and leukemia relapse. However, terminally differentiated, high avidity effector T cells (TEFF) are often limited to establish durable responses whereas less differentiated stem-cell-memory (TSCM) and central-memory (TCM) T cells can elicit potent antitumor immunity, prolonged survival and memory. Moreover, TSCM and TCM depend less on glucose consumption to drive oxidative phosphorylation (OXPHOS) as their primary source of ATP whereas TEFF utilize aerobic glycolysis to generate additional ATP. We therefore studied means of modulating T cell metabolism to generate EBV-specific TSCM and TCM for adoptive transfer and, additionally, for redirecting robust viral specificities to leukemia by e.g. retroviral T cell receptor (TCR)-transfer.

Methods: Naive HLA-A2+CD8+CD45RA+ T cells and total CD8+ T cells were stimulated by autologous DCs and subsequently PBMCs loaded with EBV-peptides in the presence of low glucose (1 mM), glutamine, an optimized cytokine cocktail and 1 mM of either glucose analog 2-deoxy-glucose (2-DG), galactose or 9-oleic acid. In addition to phenotypic and functional analyses glucose uptake and lactate production was determined. OXPHOS and aerobic glycolysis was assessed by measuring oxygen-consumption rate (OCR) and extracellular acidification rate (ECAR) on a Seahorse Analyzer.

Results: Repetitive restimulation of naive T cells revealed EBV-reactive CTL expressing a CD8+CD45RA+CD45RO- TSCM and CD8+CD45RA-CD45RO+ TCM phenotype in the presence of low glucose, glutamine and 2-DG when compared to untreated CTL. This effect was also seen in total CD8+ T cells although less pronounced. In contrast, supply of galactose or oleic-acid to glucose free (but glutamine containing) medium did not result in reduced TEFF differentiation, suggesting OXPHOS by galactose metabolism and fatty acid oxidation. Moreover, 2-DG treated TSCM and TCM showed less lactate production and ECAR as compared to untreated controls, suggesting that differentiation to TEFF requires aerobic glycolytic ATP production. 2-DG treated TSCM and TCM revealed EBV-reactivity comparable to controls, elicited superior migration properties in vitro and could be sucessfully reprogrammed with an AML-reactive TCR.

Conclusions: These studies demonstrate that modulating T cell metabolism may be a promising approach to generate TSCM/CM in vitro for improved adoptive cellular therapy.

Disclosure: No conflict of interest disclosed.


Genetische Beratung

V137 - Genetic counselling

Müller H.1

1Universitätsspital Basel, Medizinische Genetik, Basel, Switzerland

Genetic counselling is a communication process between patients or persons at risk of a disorder that may be hereditary and health care professionals. It seeks to assist the counselees to understand medical and genetic facts of the disorders. However it does not stop with informing about those facts, but must include a variety of other actions if it is to be effective. Genetic counselling gains practical importance in haematology and oncology due to the progress of molecular genetic testing.

The following topics will be discussed:

- The general concept of genetic counselling (diagnostic aspects, estimation of risks, supportive role) and its realization in daily practice,

- Genetic counselling in Mendelian and in common disorders,

- Counselling of already affected individuals,

- Genetic risk estimation for healthy family members,

- Ways of communication and empathy for counselees,

- Genetic testing and counselling of children and adolescent,

- Inclusion of healthy family members (related and unrelated),

- Autonomy of the counselee, shared decision making, non-directiveness, informed consent,

- Gene- versus gene panel-sequencing,

- Handling of an uncertain or even wrong diagnosis,

- Interdisciplinary collaboration between the medical specialists involved/the role of case managers,

- Perception of consanguinity and ethnicity,

- Sources for current scientific information concerning genetic disorders,

- Legal aspects concerning genetic testing and counselling,

- Teratogenic and genetic consequences of chemotherapy and radiation.

Disclosure: No conflict of interest disclosed.

Wissenschaftliches Symposium

Kolon- und Rektumkarzinom Optimierungen der Strategien

V143 - Are less intensive/sequential therapies in elderly and fragile patients with mCRC are justified?

Eisterer W.1

1Medizinische Universität Innsbruck, Innere Medizin V, Innsbruck, Austria

Colorectal cancer has a high prevalence in the aging population, with a median age of 69 years at diagnosis. The vast majority of patients with colon cancer will require chemotherapy treatments during their disease course, challenging oncologists with the task of tailoring therapy for older patients with CRC in the face of limited evidence-based data to guide them. Older patients, particularly those with a higher numberof comorbidities, are less likely to be referred to a medical oncologist after diagnosis and once referred, they have a lower likelihoodof receiving chemotherapy. Recent Phase-III studies (1-3) showed no inferior overall survival (OS) if 5-fluorouracil (5FU) or capecitabine was chosen as first-line therapy versus combination chemotherapy.

The MRC FOCUS2 trial (3) was conducted specifically among older and frail patients with untreated mCRC. Patients were randomly assigned to receive capecitabine or 5FU, with or without oxaliplatin, at a 20% dose reduction. Capecitabine and 5FU showed similar clinical benefit in progressions-free survival (PFS) and OS. The addition of Oxaliplatin with a 20% dose reduction resulted in improved Response rate and a lack of improvement in OS.

The randomized phase III AVEX Trial (4) compared capecitabine alone or in combination with bevacizumab and enrolled untreated patients 70 years or older who were not candidates for oxaliplatin or irinotecan. AVEX demonstrated prolonged PFS, better response and a trend toward improved OS.

Conclusion: Less intensive/sequential therapies are justified in elderly and/or fragile patients with mCRC.


1 Seymour MT et al. Chemotherapy options in elderly and frail patients with metastatic colorectal cancer (MRC FOCUS2): an open-label, randomised factorial trial. Lancet 377:1749-1759,2011.

2 Koopman M et al. Sequential versus combination chemotherapy with capecitabine, irinotecan, and oxaliplatin in advanced colorectal cancer (CAIRO): A phase III randomised controlled trial. Lancet 370:135-142,2007.

3 Ducreux M et al.: Sequential versus combination chemotherapy for the treatment of advanced colorectal cancer (FFCD 2000-05): An open-label, randomised, phase 3 trial. Lancet Oncol 12:1032-1044,2011.

4 Cunningham D, et al.: Bevacizumab plus capecitabine versus capecitabine alone in elderly patients with previously untreated metastatic colorectal cancer (AVEX): an open-label, randomized phase 3 trial. Lancet Oncol 14:1077-85, 2013.

Disclosure: No conflict of interest disclosed.


HIV-assoziierte Tumoren

V147 - Aggressive HIV-associated B-cell lymphoma: options and limits of therapeutic approaches

Hübel K.1

1Universitätsklinik Köln, Klinik I für Innere Medizin, Köln, Germany

Patients infected with HIV have an increased risk of developing aggressive B-cell non-Hodgkin's lymphoma (NHL). Recent data indicate that among AIDS-related death NHL is the most frequent event. The majority of patients have poor prognostic features such as advanced-stage disease or high International Prognostic Index (IPI). Since the introduction of combination antiretroviral therapy (cART) therapeutic options and prognosis have considerable improved. However, the overall outcome remains worse compared to immunocompetent patients.

Work-up and staging of patients should be performed in accordance with guidelines established for HIV-negative NHL. In diffuse large B-cell lymphoma (DLBCL), most centers use 6 cycles of R-CHOP 21 as first-line treatment achieving a complete remission in about 60% of patients. Date clearly show that the addition of rituximab improves outcome compared to CHOP alone. However, if CD4 cell counts is below 50/µl, rituximab is not recommended due to increased risk of infection. The use of R-EPOCH may further improve prognosis compared to R-CHOP. In Germany, a prospective trial evaluating the R-CHOEP regimen as first-line treatment in DLBCL will be initiated soon. In the relapse situation, patients may be candidates for high-dose therapy followed by autologous stem cell transplantation as in the HIV-negative setting. In Burkitt lymphoma, does-intensive regimens such as the B-ALL/NHL2002 protocol of the GMALL or the Hyper-CVAD/HD-MTX protocol are successfully applied to patients who are in an acceptable condition. Patients with rare, aggressive diseases as plasmablastic lymphoma or primary effusion lymphoma represent a therapeutic challenge for clinicians since no therapeutic approach has been clearly recommended yet.

There is no doubt the cART applied concurrently to chemotherapy is an essential module to a successful therapeutic outcome provided that pharmacokinetik interactions are being considered.

In summary, significant advances have been made in the understanding and treatment of HIV-associated B-cell lymphomas in recent years. Prospective trials are needed to develop treatment algorithms and to investigate novel therapeutic approaches for patients with HIV-lymphoma.

Disclosure: No conflict of interest disclosed.

V148 - Non-AIDS-defining malignancies: current therapeutic concepts

Hentrich M.1

1Rotkreuzklinikum, München, Germany

Combined antiretroviral therapy (cART) has dramatically improved the life expectancy of patients living with HIV. However, the risk of cancer is substantially increased in HIV-infected patients. In western countries the number of cases of non-AIDS-defining malignancies (NADM) now equals or exceeds the number of cases of AIDS-defining cancers, and NADM are a leading cause of mortality for HIV-infected subjects. Important risk factors for NADM are advanced age, the length of time one has been infected with HIV, exposure to cigarette smoking, and oncogenic viruses such as Epstein-Barr virus, Hepatitis B and C virus, and human papillomavirus. The most frequent NADM are lung cancer (LC), Hodgkin lymphoma (HL), anal cancer (AC), liver cancer (HCC) and head & neck cancer. Basically, patients with NADM should be treated in an identical manner to patients without HIV infection. Most cases of LC occur in the setting of limited immune deficiency and a long-lasting viral suppression. However, as in the HIV-negative setting long-term overall survival can only be achieved at limited LC stages. Improved survival rates were reported for patients undergoing stage-adapted treatment of HIV-HL. Two cycles of ABVD followed by involved-field (IF) radiation therapy (RT) can be regarded as standard treatment for early favorable HL. Four cycles of ABVD followed by IF-RT are standard of care for early stage unfavourable HL, and patients with advanced stage HL should receive 6 cycles of BEACOPP baseline or 6-8 cycles of ABVD. The management of AC with chemoradiotherapy (CRT) achieves similar outcomes as the general population. However, CRT is associated with significant prolonged CD4 suppression. The outcome of HCC in the setting of HIV remains poor. During radio- or chemotherapy patients should receive concurrent cART. However, interactions between cytotoxics and antiretrovirals must be considered, as chemotherapy-related toxicity may be markedly increased by concomitant use of antiretrovirals. Antimicrobial prophylaxis should be considered in accordance with current guidelines. In particular, prophylaxis against pneumocystis jiroveci pneumonia is indicated if the CD4 cells are <200/µl.

Disclosure: No conflict of interest disclosed.

Wissenschaftliches Symposium

Maligne Gliome

V152 - Recent diagnostics, therapy standards and perspectives in patients with anaplastic gliomas and glioblastomas (WHO grade III/IV)

Hau P.1

1Universität Regensburg, Neurologie, Regensburg, Germany

Anaplastic gliomas and glioblastomas cannot be cured at this time. However, genetic markers increasingly enable prognostication and allow stratification into specific treatment groups, even if the standard histology is not distinctive.

The most widely used drug in the treatment of gliomas is the alkylating agent Temozolomide. In addition, other alkylating agents and the antiangiogenic substance Bevacizumab are used. Bevacizumab generates impressive efficacy in a subgroup of patients with gliomas. However, after two negative Phase III-trials, the substance is not approved in all countries. Immunotherapeutic strategies become increasingly interesting. Relevant, beyond others, are the induction of humoral and celluar immune responses against EGFRvIII and the mutated form of IDH1 (IDH1R132H), autologous dendritic cell vaccinations with e.g. peptide cocktails (ICT-107), and use of immune checkpoint blocking agents against PD-1/PD-L1 and CTLA-4. A novel concept uses tumor treating fields (TTF, Optune) that induce apoptosis. A final decision on this treatment can be made after full publication of a completed Phase III-trial that is expected this summer.

Therapeutic decisions increasingly include genetic markers. The methylation of the promotor of O6-Methyl-Guanin-Methyl-Transferase (MGMT) and the codeletion of chromosome arms 1p and 19q (LOH1p1/19q) have been evaluated as predictive markers in a number of trials. They are routinely used in at least two situations for treatment stratification, namely in elderly patients with glioblastoma (where MGMT is used to stratify to radiotherapy or chemotherapy) and in patients with anaplastic glioma with 1p19q codeletion. In these patients, a combination of radiotherapy and chemotherapy with Procarbacin, CCNU and Vincristin doubles overall survival in comparison to radiotherapy as monotherapy.

The development in the use of genetic markers for prognostication and therapy stratification is dynamic. Considering this, a new classification of gliomas will soon include genetic markers and will therefore more thoroughly inform the treating physician about the best treatment of choice.

Disclosure: No conflict of interest disclosed.

V153 - Recommendations for the treatment of recurrent malignant glioma

Weller M.1

1Universitätsspital Zürich, Klinik für Neurologie, Zürich, Switzerland

Newly diagnosed glioblastoma is now commonly treated with surgery as feasible or biopsy, followed by radiotherapy plus concomitant and adjuvant temozolomide. The treatment of recurrent glioblastoma is less well standardized and depends on multiple factors, including efficacy and tolerability of prior treatment, time from diagosis, topographic pattern of relapse, and MGMT promoter methylation status.

A minority of patients are candidates for second surgery or reirradiation, based on tumor size and location. Progression-free survival rates at 6 months of 20%-30% have been reported with nitrosoureas, temozolomide rechallenge in various dosing regimens, or bevacizumab. Temozolomide rechallenge is not active in patients with tumors lacking MGMT promoter methylation (DIRECTOR) whereas bevacizumab is not approved in the EU.

Data from a phase III trial (EORTC 26101) comparing the combination of lomustin and bevacizumab versus lomustin alone are expected near the end of the year. Emerging therapies may include vaccination against mutant epidermal growth factor receptor variant III (EGFRvIII) (ReACT) and the PD-1 antibody nivolumab.

More research is needed to better define patient profiles that predict benefit from the limited therapeutic options available after the current standard of care has failed.

Disclosure: Michael Weller: Financing of Scientific Research: Celldex, Immunocellular, Isarna, Magforce, MSD, Merck & Co, Northwest Biotherapeutics, Novocure, Pfizer, Roche, Teva; Expert Testimony: Acceleron, Actelion, Alpinia Institute, Bayer, Isarna, MSD, Merck & Co, Novocure, PIQUR, Roche.


Kopf-, Hals-, Schild- und Speicheldrüsentumore - Update 2015

V154 - Thyroid cancers: which molecular marker are relevant?

Siano M.1,2

1Cantonal Hospital St. Gallen, Clinic of Oncology / Hematology, St. Gallen, Switzerland, 2Fondazione IRCCS Istituto Nazionale dei Tumori, Head and Neck Medical Oncology Unit, Milan, Italy

Even if rarely encountered by the medical oncologist, his knowledge is required for iodine refractory differentiated thyroid cancer (DTC), poorly differentiated and anaplastic TC. In de-differentiated carcinomas as in iodine refractory, molecular alterations play an important role. There is a ‚career of dedifferentiation' according to mutational status (Xing 2013). Moreover, mutations have direct impact on iodine uptake and so on effectiveness of radioiodine therapy. With targeted therapies e.g. it is possible to convert iodine refractoriness in follicular TC (Hu 2013, Rothenberg 2015). This shows that even if molecular markers are not yet established in decision-making, they will gain importance as prognostic and predictive markers.

Mutational landscape was soundly analyzed particularly for papillary TC (Agrawal 2014). Different gene signatures were identified (BRAF-like und RAS-like). Affinity towards specific targets, tumor pathways and gene expression signatures lead to possible assumptions about best treatment or best use of novel tyrosine kinase inhibitors (TKI). Even if tempting, this approach is for the moment, not supported by data. Beside mutational status, angiogenesis seems to be of major importance. TKI with anti-angiogenetic properties, show to be very active (Lenvatinib, Axitinib). This confirms that some genetic alterations are not drivers and that TC serves more mechanisms.

Prognostic value of BRAF mutation, even if controversially discussed at primary diagnosis, is associated with higher recurrence rates (Xing 2015) and seems prognostic in advanced disease (in iodine refractoriness). TERT mutation indicates poor prognosis, notably in papillary TC and seems to be associated with distant metastasis and increased disease specific mortality (Melo 2014, Gandolfi 2015).

For DTC Sorafenib and Lenvatinib and for medullary TC Vandetanib and Cabozantinib are or will soon be registered. Still these agents are not applied in a selected patient population. BRAF mutational status is not predictive for better outcome with Sorafenib treatment (Brose 2014). Vemurafenib und Dabrafenib are investigated in selected patients harboring a BRAF mutation showing satisfactory response rates.

No predictive molecular marker has been identified or was soundly validated so far. Only for Cabozantinib treatment in medullary TC, RET M918T mutation was able to show in a retrospective analysis, predictive potential for overall survival (Elisei 2013, Schlumberger 2015).

Disclosure: Marco Siano: Advisory Role: Beratertätigkeit: Bayer, Eisai.


Ethik Advance Care Planning

V159 - “Existential communication” at the end of life - conceptual and practical aspects

Gieseler F.1, Schäfer V.1, Theobald W.2

1UKSH Campus Lübeck, Med I, Palliativmedizin und Ethik in der Onkologie, Lübeck, Germany, 2Agrar- und Ernährungswissenschaftliche Fakultät, Ethik in den Lebenswissenschaften, Kiel, Germany

Empathic patient-centered communication is an integral part of the therapeutic care in palliative oncology. The integration in the German National Cancer Plan and the Patients´ Rights Act underlines this consent. Thus, the doctor is obliged to explain in an understandable way, all significant information for the diagnosis, all treatment options, and probable side effects (SGB V § 630 c BGB). The requirement is that all service providers have the necessary communication skills (NKP Ziel 12a).

These objectives and demands do not consider that terms and conditions in palliative oncology have changed considerably including external environment (space, time frame, etc.), the background of the patient (family, friends, counselors) and the patients themselves (age structure, level and sources of information). We believe that a communication style should be developed, taking account of these changing conditions and thus leading to more patient satisfaction and less “burn-out symptoms” among physicians.

Despite the developments of modern oncology, the existential threat by the cancer diagnosis is still vivid. Also doctors and nurses have to cope with finiteness and limitations, e.g. on the time available, treatment options and financial resources. We work on the subject with in the “Projektkolleg Erfahrung und Umgang mit Endlichkeit” of the Collegium Philosophicum at the CAU Kiel. The term of our subproject “Existenzielle Kommunikation” we have borrowed from the philosophy of Carl Jaspers, but augmenting it with the aspect of communication in a situation of existential threat.

We found that ethical aspects and communication style represent stressors for patients and physicians alike. One hypothesis is that the communication style is a part of the treatment concept and should be adjusted to the specific situation. Styles include the patriarchal approach, shared decision making and patient centered communication in the sense of service oriented approach. Advance care planning needs to be integrated into the concept. We present here among others a didactic method in teaching communication (“double reflection”).


Birninger, Gieseler. “Communication between nurses and patients in oncology” Oncol Res Treat 37,2014;S1:110.

Mannhardt, Ogbonnaya, Gieseler. “Double Reflection” Oncologist 18 (2013):1058.

Gieseler, Schäfer, Theobald “Entscheidungen im Schatten der Endlichkeit “. In “Endlichkeit“, Bihrer (Hrg.), transcript-Verl, accepted.

Disclosure: No conflict of interest disclosed.

Freier Vortrag

Allogene Transplantation klinisch I

V161 - Ruxolitinib for corticosteroid-refractory graft-versus-host disease: analysis of 95 patients treated at multiple medical centers

Zeiser R.1, Burchert A.2, Lengerke C.3, Verbeek M.4, Maas-Bauer K.1, Metzelder S.2, Spoerl S.4, Ditschkowski M.5, Ecsedi M.3, Sockel K.6, Ayuk F.7, Ajib S.8, Sicre de Fontbrune F.9, Na I.-K.10, Penter L.10, Holtick U.11, Wolf D.12, Schuler E.13, Meyer E.14, Apostolova P.1, Bertz H.1, Marks R.1, Lübbert M.1, Wäsch R.1, Scheid C.11, Ordemann R.6, Bug G.8, Kobbe G.13, Negrin R.14, Brune M.15, Spyridonidis A.16, Schmitt-Gräff A.17, van der Velden W.18, Huls G.18, Grigoleit G.U.19, Kuball J.20, Blazar B.R.21, Arnold R.10, Kröger N.7, Passweg J.3, Halter J.3, Socié G.22, Beelen D.5, Peschel C.4, Neubauer A.2, Finke J.1, Duyster J.1, von Bubnoff N.1

1Freiburg University Medical Center, Department of Hematology, Oncology and Stem Cell Transplantation, Freiburg, Germany, 2Marburg University Medical Center, Department of Hematology, Oncology and Stem Cell Transplantation, Marburg, Germany, 3University Hospital Basel, Hematology, Basel, Switzerland, 4Technical University of Munich, III Department of Internal Medicine, Munich, Germany, 5West German Cancer Center, University Hospital Essen, Department of Bone Marrow Transplantation, Essen, Germany, 6Universitätsklinikum Carl Gustav Carus an der Technischen Universität Dresden, Department of Hematology, Oncology and Stem Cell Transplantation, Dresden, Germany, 7University Hospital Hamburg-Eppendorf, Department of Hematology, Oncology and Stem Cell Transplantation, Hamburg, Germany, 8University Hospital Frankfurt/Main, Frankfurt, Germany, 9Saint Louis Hospital, APHP, Hematology Stem cell transplant Unit, Paris, France, 10Charité University Medicine Berlin, Department of Stem Cell Transplantation, Berlin, Germany, 11University Hospital Cologne, Department of Internal Medicine I, Cologne, Germany, 12University Hospital Bonn (UKB), Medical Clinic III, Oncology, Hematology and Rheumatology, Bonn, Germany, 13Universitätsklinikum Düsseldorf, Department of Hematology, Oncology and Clinical Immunology, Düsseldorf, Germany, 14Stanford University Medical School, Department of Bone Marrow Transplantation, Stanford, United States, 15University of Gothenburg, Sahlgrenska University Hospital, Gothenburg, Sweden, 16Patras University Medical School, Department of Bone Marrow Transplantation, Patras, Greece, 17Freiburg University Medical Center, Department of Pathology, Freiburg, Germany, 18Radboud University Medical Centre, Department of Hematology, Oncology and Stem Cell Transplantation, Nijmegen, Netherlands, 19University Medical Centre Würzburg, Department of Hematology, Oncology and Stem Cell Transplantation, Würzburg, Germany, 20University Medical Center Utrecht, Department of Hematology, Utrecht, Netherlands, 21University of Minnesota, Minneapolis, Division of Blood and Marrow Transplantation, Minnesota, United States, 22Saint Louis Hospital, APHP, Paris, Hematology Stem cell transplant Unit, Paris, France

Introduction: Despite major improvements in allogeneic hematopoietic cell transplantation (allo-HCT) over the last decades, severe acute and chronic graft-versus-host disease (GvHD) still remains a life-threatening complication associated with high mortality. Pre-clinical evidence has indicated an anti-inflammatory effect of ruxolitinib, therefore we analyzed responses to ruxolitinib as salvage treatment in patients suffering from corticosteroid refractory GvHD.

Methods: A total of 19 Stem Cell Transplant Centers in Europe and United States reported outcome data from 95 patients who received ruxolitinib for corticosteroid-refractory GvHD (skin, mucosa, intestine, liver, lung, musculoskeletal) between 01/2012 and 04/2015. Patients were classified as having acute (n = 54, only grade 3 or 4) or chronic (n = 41, only moderate or severe) GvHD. The median number of previous GvHD-therapies was 3 for acute GvHD (range: 1-7) and 3 for chronic GvHD (range: 1-10).

Results: The overall response rate was 81.4% (44/54) in acute GvHD comprising 25 CRs (46%). In chronic GvHD the overall response rate was 85% (35/41). Clinical improvement was at a median time to response of 1.5 (1-10) weeks and 3 (1-25) weeks after initiation of ruxolitinib treatment in acute and chronic GvHD, respectively. GvHD relapsed in 6.8% (3/44) and 5.7% (2/35) of the ruxolitinib-responsive patients with acute or chronic GvHD, respectively. The median follow-up was 22 (3-98) and 25 (2-112) weeks for acute or chronic GvHD patients. Cytopenias (anemia, leukopenia or thrombocytopenia) and CMV reactivation were observed during the time of ruxolitinib treatment in both acute (30/54, 55.5% and 18/54, 33.3%) or chronic (7/41, 17% and 6/41, 14.6%) GvHD patients. Cytopenias had preceded ruxolitinib treatment in 51.8% (28/54) and 14.6% (6/41) of the patients with acute or chronic GvHD, respectively. Relapse of the underlying malignancy occurred in 9.2% (5/54) and 2.4% (1/41) of the patients with acute or chronic GvHD, respectively.

Conclusions: Overall, these data collected in multiple centers, using different strategies for GvHD prophylaxis and treatment, suggest that ruxolitinib is a very promising agent in the treatment of corticosteroid-refractory acute or chronic GvHD and may be successfully used to salvage a major subset of patients beyond 2nd line of GvHD treatment and needs to be validated in a prospective trial.

Disclosure: No conflict of interest disclosed.

V162 - Genetic polymorphism of cytochrome P450 1B1 (C432G) is associated with lower overall survival in male patients undergoing allogeneic haematopoietic stem cell transplantation

Stute N.1, Beelen D.1, Koldehoff M.1

1Clinic for Bone Marrow Transplants, University of Duisburg-Essen, Essen, Germany

Introduction: Human cytochrome P450 1B1 (CYP 1B1) is a key enzyme involved in estrogen/drug metabolism, steroid synthesis and pro-carcinogen activation. It is highly expressed in some human cancers and some myeloid cells but not in liver or kidney. The CYP 1B1 codon 432 polymorphism (one of four identified SNP) leads, among other things, to a four-fold higher km in the 4-hydroxylation of 17ß-estradiol.

Methods: In a single center retrospective study 382 patients (53% male) who underwent allogeneic HSCT (95% myeloablative) for various diseases (51% acute leukemia) and their respective donors were genotyped for CYP 1B1 (C432G) polymorphism by rt-PCR and the influence on outcome was analyzed. Median follow-up was 76 months.

Results: 169 patients (44%) were genotyped as homozygous wild-type (wt) gene C/C, 157 (41%) as heterozygous genotype C/G and 56 (15%) as homozygous gene mutated G/G.

Patients' demographic, disease and treatment characteristics were equally distributed and showed no difference between the three subgroups (C/C, C/G and G/G) except that CYP 1B1 C/C was more common in females (52%) than in males (38%), p=.02.

Five-year estimate for overall survival (OS) was 58% ±4 for the C/C group and 48% ±3 for the C/G- and G/G groups (p=.036). Surprisingly, this difference was primarily evident in males (p=.024), where the group with CYP 1B1 gene mutations did significantly worse: 58% ±6 vs. 42% ±4, whereas it was virtually absent in females.

Transplant-related mortality (TRM) in male patients was higher for the group with mutated (mut) genes compared to the group with wt gene: One-year estimate for TRM 15% ±3 C/C vs. 20% ±3 C/G vs. 38% ±4 G/G (p=.048).

Three-year estimate for relapse rate in male patients was 31% ±6 (wt) vs. 41% ±5 (mut), p=.097. Interestingly, male patients had more relapses than female patients if mut recipient SNP 41% ±5 vs 24% ±5 (p=.007) or mut donor SNP.

Multivariate analysis (Cox regression) for OS in male patients revealed only two prognostic factors: mut donor SNP relative risk (RR) 1.4 (CI: 1.1-1.9), p=.007 and advanced disease RR 2.8 (CI: 1.9-4.1), p=.000.

Conclusion: These results suggest that male patients with genetic polymorphism of CYP 1B1 have a lower OS due to a higher TRM and relapse rate after HSCT. Genotyping for CYP 1B1 (C432G) might help to identify patients with higher risk for allogeneic HSCT and may help explain gender differences in outcome. Prospective studies or confirmation in independent datasets are warranted.

Disclosure: No conflict of interest disclosed.

V163 - Hepatic iron overload predicts for poor long-term overall survival in AML and MDS patients undergoing allogeneic stem cell transplantation

Eckoldt J.1, von Bonin M.1, Sockel K.1, Middeke M.1, Stölzel F.1, Schetelig J.1, Ehninger G.1, Theurl I.2, Bornhäuser M.1, Platzbecker U.1, Wermke M.1

1Uniklinik Dresden, Medizinische Klinik I, Dresden, Germany, 2Uniklinik Innsbruck, Department für Innere Medizin VI, Innsbruck, Austria

Background: Iron overload (IO) is a frequently observed condition in AML and MDS patients undergoing allogeneic stem cell transplantation (allo-SCT). Its definition and prognostic implications are a matter of constant debate. We have shown recently, that IO characterized by a liver iron content (LIC) ≥125 µmol/g (7 mg/g) as assessed by magnetic resonance imaging (MRI) correlated with a significantly increased early non-relapse mortality. However, this study as well as all other LIC-based studies published thus far, were characterized by a short follow up leaving the impact of IO on long-term outcomes such as overall survival (OS) and chronic GvHD (cGvHD) elusive.

Patients and methods: Here we report the long-term outcome of our previously published cohort of 64 AML and 24 MDS patients undergoing allo-SCT. Pre-transplantation IO was measured by serum ferritin, transfusion burden or liver MRI.

Results: After a median follow-up of 41 (range 4-66) months, IO defined by a LIC of ≥125 µmol/g was associated with a significantly shortened overall survival (Figure 1, p = 0.013). Elevated LIC was identified as an independent adverse risk factor for OS in a multivariate Cox regression model (HR 2.15, 95% CI 1.16-3.98, p = 0.015), whereas an increased serum ferritin above 2500 ng/ml or a transfusion burden of ≥20 red blood cell concentrates (RBC) did not predict for adverse prognosis. Hepatic IO was not associated with an increased likelihood of relapse (3 year cumulative incidence: IO: 36.6% vs. non-IO: 41.9%, p = 0.966) or cGvHD (3 year cumulative incidence: IO: 47.5% vs. Non-IO: 66.3%, p = 0.138). Infections after allo-SCT occurred at a similar frequency in IO (65.9%) and non-IO (68.2%) patients. Infections and GvHD were the most common causes of mortality each accounting for a third of all deaths without obvious differences between patients with and without an elevated LIC.


Conclusion: The increased early NRM observed in iron-overloaded AML and MDS patients undergoing allo-SCT translated into a reduced long-term overall survival. The causative relationship between IO and mortality warrants further investigations within prospective clinical trials.

Disclosure: Julia Eckoldt: No conflict of interest disclosed. Martin Wermke: Expert Testimony: Novartis.

V164 - Post-transplant cyclophosphamide reduces haplo-identical transplant related mortality in non hodgkin lymphoma yielding similar results as with sibling donors

Dietrich S.1,2, Finel H.2, Martinez C.2, Tischer J.2, Blaise D.2, Chevallier P.2, Castagna L.2, Milpied N.2, Bacigalupo A.2, Corradini P.2, Mohty M.2, Sanz M.2, Velardi V.2, Hausmann A.2, Montoto S.2, Hermine O.2, Schmitz N.2, Schouten H.2, Anna S.2, Robinson S.2, Dreger P.2

1Medizinische Klinik V, Heidelberg, Germany, 2EBMT LWP, Paris, France

Background: Allogeneic stem cell transplantation (SCT) can provide long-term disease control for non hodgkin lymphoma (NHL), but its use depends on the availability of a matched donor. Haplo-identical relatives represent a reasonable alternative but data for haplo-transplants in NHL is sparse.

Methods: Information of patients with mantle cell lymphoma (MCL), DLBCL, T-cell lymphoma (TCL) and follicular lymphoma (FL) who received an SCT from a sibling donor (SIB), 10/10 matched unrelated donor (MUD), haplo-identical donor (HAPLO) or cord blood (CORD) between 2007 and 2012 was downloaded from the EBMT database.

Results: 2798 patients with NHL met the inclusion criteria. 2065 received a transplant from a SIB, 447 from a MUD, 167 from CORD (18 MCL, 36 DLBCL, 43 FL, 70 TCL) and 119 from a HAPLO donor (16 MCL, 30 DLBCL, 22 FL, 51 TCL). For 99 patients we received additional information on immunosuppression details after haplo-transplantation. 60 patients were treated with post-transplant cyclophosphamide (post-SCT cy) and 39 patients received other haplo-protocols. Patient characteristics were balanced except for higher median age in the post-SCT cy group. OS was significantly better for post-SCT cy treated patients than for alternative haplo-protocols (p = 0.0052, HR 2.3). More non relapse mortality (NRM) related deaths among alternative haplo-transplants accounted for the difference in OS (p = 0.032, HR 1.8), whereas relapse rates were similar. Observed trends for better OS of post-SCT cy treated patients held separately true for B- and T-cell lymphomas.

Comparison of post-SCT cy haplo-transplants with alternative donor types demonstrated similar OS of SIB-, MUD- and post-SCT cy haplo-transplants, although haplo-transplants were performed in more advanced disease stages, patients had poorer performance status and were older. OS of CORD and alternative haplo-transplants were significantly worse than for SIB, MUD and post-SCT cy haplo-transplants (p < 0.001), which was attributable to lower NRM incidences for SIB, MUD and post-SCT cy haplo-transplants (p < 0.001). Acute severe GvHD (grade 3-4) incidences were not different between donor groups, but post-SCT cy haplo-transplants had a trend towards lower chronic GvHD incidences.

Conclusions: Haplo-identical donors and immunosuppression with post-SCT cy is a valuable alternative to SIB or MUD transplants in NHL patients whereas CORD- and alternative haplo transplants are associated with increased mortality.

Disclosure: No conflict of interest disclosed.

V165 - CMV infection plays a crucial role in the immune reconstitution after allogeneic stem cell transplantation

Hartjen A.S.1, Krumbholz A.2, Thieme F.3, Bulduk M.1, Humpe A.1, Gramatzki M.1, Günther A.1

1CAU / UKSH Kiel, Sektion für Stammzell- und Immuntherapie, II. Med. Klinik, Kiel, Germany, 2CAU / UKSH Kiel, Institut für Infektionsmedizin, Kiel, Germany, 3UKSH, Institut für Klinische Chemie, Kiel, Germany

Introduction: The renewal of the immune system after allogeneic stem cell transplantation is a complex process not fully understood. Recently the use of reduced intensity conditioning regimens, new anti-viral drugs and intensified GvHD treatment changed the landscape of allogeneic transplantation. Here, we analyzed the influence of several parameters on the development of the cellular and humoral immunity of 136 patients consecutively transplanted in a single center.

Methods: All adult patients (median age 52 years) transplanted over three years starting in September 2010 were analyzed retrospectively in respect of routinely measured immunoglobulin levels (turbidimetric method) and lymphocyte subset counts (FACS staining). Only in 9% of patients full conditioning regimen was used; nearly all patients received either anti-thymoglobulin (ATG) or alemtuzumab (25/136 matched related donors) leading to a relatively low rate of chronic GvHD (40%). The immune parameters achieved one year after stem cell transplantation were correlated with the conditioning regimen, diagnosis, relapse, age, sex and history of CMV infection, acute and chronic GvHD using Pearson correlation, t-test and regression analysis (SPSS).

Results: A normal CD4/CD8 ratio at day +360 was only seen in 14% of available patients (3% elevated, 83% reduced) mostly caused by impaired CD4 cell count (90% < normal) while CD8 cell count was increased in 42%. The IgG level was normal in 51%, reduced in 31% and increased in 17% of tested patients. The CD8 cell count was significantly higher in patients after CMV infection (p < 0.001) and without alemtuzumab (p = 0.017) in the conditioning regimen; the IgG level correlated with history of CMV infection, CD4, NK (all p = 0.002) and CD8 (p = 0.025) cell count; age, sex, diagnosis, relapse, acute and chronic GvHD had no significant impact.

Conclusions: Despite progress in recent years nearly all patients show a compromised immune system one year after allogeneic stem cell transplantation mainly characterized by a reduced CD4 cell count. In contrast, the CD8 cell count is often elevated probably reflecting response to viral infections. Interestingly, the immunoglobulin levels are highly variable ranging from antibody deficiency to hyper-immunoglobulinemia positively correlating with previous CMV infection. Surprisingly, GvHD had only a minor impact on the immune reconstitution of the investigated collective.

Disclosure: No conflict of interest disclosed.

V166 - Regulatory B-cells can be identified early post-transplant and are raised in low grade acute Graft versus Host disease

Chakupurakal G.1, Garcia-Marquez M.1, Shimabukuro-Vornhagen A.1, Schloesser H.2, Theurich S.3, Scheid C.1, Hallek M.1, Holtick U.1, von Bergwelt-Baildon M.1

1Uniklinikum Koeln, Innere Medizin I, Koeln, Germany, 2Uniklinikum Koeln, Klinik für Allgemein-, Viszeral- und Tumorchirurgie, Koeln, Germany, 3Uniklinikum Koeln, Max-Planck-Institute for Metabolism Research, Koeln, Germany

Allogeneic stem cell transplantation (alloSCT) is the curative therapeutic option for a variety of haematalogical malignancies. Graft versus Host disease (GvHD), with an incidence of around 40-60%, remains a major post transplant complication. The role of B-cells and especially the Interleukin-10 (IL-10) producing regulatory B-cells (B10-cells) in the pathophysiology of GvHD is not clearly understood to date. We studied B cells in allogeneic transplant recipients in the early post transplant phase.

Samples, on 95 patients, transplanted at the University of Cologne and 10 healthy donors were collected prospectively after obtaining informed consent Patients were transplanted mostly for haematological malignancies (n = 94). Male: Female = 52:43. 86/95 90% received RIC regimens (90%). No patient received a B-cell depleting agent in the conditioning regimen or in the 6 weeks prior to transplant . 43 (46%) had no GvHD, and 10 (10%), 9 (9%),17 (18%)and 16 (17%) had grade 1-4 GvHD as per the modified Glucksberg criteria 6 respectively.

Day 30 post transplant the total B-cell percentages in transplant recipients were significantly reduced in comparison to the controls (p=< 0.0001). Transitional B-cells were significantly more in transplant recipients (p = 0.004). Omission of data on the MAC patients did not affect the results. B10-cells could be demonstrated in 22 transplant recipients studied (12 with acute GvHD). The percentages of B10-cells were significantly reduced in the transplant recipients on comparison with the control cohort (p < 0.0001). The mean percentage of B10-cells in patients with acute GvHD (1.94±1.2), though lower than control cohort 5.01±2.2, was significantly higher than those without GvHD 0.84±0.6 (p = 0.0003).

This is the first study of B-cells and B-cell subsets suggesting that B-cell reconstitution commences immediately following engraftment. This is the first report demonstrating B10-cells in stem cell transplant recipients in the early post transplant (30 days) period and that they are enriched within the transitional cell compartment. Contrary to expectations, the percentage of B10-cells in patients with GvHD was significantly higher than those without GvHD (Fig 2C). The frequency of B10 cells were significantly higher in patients with lower grade GvHD suggesting a parallel to the observation on T-regs- where higher the frequency of T-regs the lower the GvHD grade and lower the non-relapse mortality (NRM) associated with an alloSCT.

Disclosure: No conflict of interest disclosed.


Leichtketten-Amyloidose: Aktueller Stand der Diagnostik und Therapie

V169 - Light-chain amyloidosis: Current status of diagnostic and treatment

Hegenbart U.1, Schönland S.1

1Universitätsklinikum Heidelberg; V. Medizinische Klinik, Amyloidose-Zentrum, Heidelberg, Germany

Systemic amyloidoses are rare life-threatening disorders. Misfolded proteins which circulate in the blood are deposited in several organs (excluding the central nervous system) leading to organ dysfunction or failure. The prognosis is still poor: patients with advances cardiac involvement may die within 6-12 months of cardiac failure. There are various causes which must be differentiated. Amyloidoses are discriminated due to the causative pathologic protein (e.g. light chain, SAA or transthyretin) and in hereditary and non-hereditary forms. The amyloid light-chain (AL) amyloidosis is most common in Europe. In Germany about 800 patients will be diagnosed per year. A certain differentiation of the amyloid types is essential to avoid wrong treatment. First symptoms are non-specific; therefore the diagnosis is often (very much) delayed. Due to the availability of new diagnostic tools and treatments prognosis has improved; however the early diagnosis plays a crucial role to avoid organ failure or death of the patients. Goal of the chemotherapy is to destroy the clonal plasma cells. The missing replenishment of amyloidogenic light chains is able to stop the pathologic process of protein deposition which leads to improvement of organ function and quality of life in a large number of patients (within months to years). The multidisciplinary collaboration of several medical disciplines has an outstanding role to care for those severely ill patients.

Disclosure: Ute Hegenbart: Financing of Scientific Research: Janssen, Celgene, Binding Site. Stefan Schönland: Financing of Scientific Research: Janssen, Celgene, Binding Site; Expert Testimony: Janssen, Celgene.



P170 - Feasibility of semi-automated MRD analysis by flow cytometry in acute myeloid leukemia

Köhnke T.1,2, Rechkemmer S.1, Bücklein V.L.1,2, Hiddemann W.1, Subklewe M.1,2

1LMU München, Medizinische Klinik und Poliklinik III, München, Germany, 2Klinische Kooperationsgruppe Immuntherapie am Helmholtz Zentrum München, München, Germany

Introduction: The relevance of minimal residual disease (MRD) detection in AML has increased within the past decade and currently, clinical trials are evaluating the value of MRD-guided therapy. Analysis of this data is performed manually, by creating gates defining a leukemia-associated immunophenotype (LAIP) at primary diagnosis, which is then applied to follow-up samples. However, antigen expression has been shown to display some variability during therapy, which might reduce the sensitivity of this strategy.

Methods: To test the feasibility of the recently proposed viSNE (visualization of t-distributed stochastic neighbor embedding) algorithm to detect aberrant populations, we analysed patient samples acquired within 3 months prior to hematological relapse, which were MRD-negative by traditional LAIP-gating. 400.000 events were acquired using an 8 color, 10 parameter panel on a Navios® flow cytometer (Beckman Coulter, Brea, CA, USA). For the bioinformatic analysis, the patient's sample and a healthy donor sample measured on the same machine were combined digitally and viSNE clustering was applied to the resulting dataset. As previously suggested, MRD positivity was defined as the presence of a distinct cluster of >100 cells which consisted of >90% patient cells.

Results: From our database, we identified 12 patients with comprehensive MRD flow assessment available within 3 months prior to hematological relapse who were in CR by cytomorphology at the time of MRD assessment. 9 of these patients were MRD-positive by traditional LAIP-gating (>0.1%). This first analysis therefor focused on the remaining 3 patients with MRD-levels at 0.01%, 0.03% and 0.07% at 21, 42 and 37 days prior to hematological relapse. Using viSNE, distinct clusters of 5058, 225 and 1043 events consisting of 95.5%, 92.6% and 93.8% patient cells were identified, consistent with MRD positivity as defined above. In all three cases, the cells identified by the viSNE algorithm showed the immunophenotype seen at relapse, while there were immunophenotypical shifts compared to primary diagnosis, possibly explaining false-negative MRD-assessment by traditional gating.

Conclusion: viSNE clustering detected minor, aberrant populations in 3 patients with subsequent hematological relapse, which were not detected by traditional LAIP-gating strategy. viSNE might therefor aid in the analysis of high-dimensional flow cytometry data to detect small, aberrant populations. Further validation is ongoing.

Disclosure: No conflict of interest disclosed.

P171 - LSD1 inhibition induces differentiation in murine models of AML

Barth J.1, Scheder A.-M.2, Schulz-Fincke J.1,3, Schmitt M.3, Walter A.3, Lübbert M.4, Jung M.3, Serve H.2, Berg T.1,2

1German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ), Heidelberg, Germany, 2Goethe Universität Frankfurt, Medizinische Klinik II - Hämatologie/Onkologie, Frankfurt, Germany, 3University of Freiburg, Institute of Pharmaceutical Sciences, Freiburg, Germany, 4University of Freiburg, Department of Medicine I, Hematology, Oncology and Stem Cell Transplantation, Freiburg, Germany

The histone demethylating enzyme LSD1 represents a promising epigenetic target in the treatment of AML and inhibition of LSD1 has been shown to facilitate a response of AML cells to all-trans retinoic acid (ATRA). In this project, we have now modeled the effect of pharmacological inhibitors of LSD1 in different murine leukemia models.

Hematopoietic progenitor cells isolated from murine bone marrow were transformed by retroviral overexpression of a combination of Hoxa9 and Meis1 or MN1 turning them into cytokine-dependent progenitor cultures that elicit AML upon transplantation into mice. We have then used these cells to study the effect of different pharmacological inhibitors of LSD1. In detail, we treated the Hoxa9/Meis1- and MN1-transformed cells with three irreversible (tranylcypromine, AW69, AW84) and two reversible LSD1 inhibitors (MS120 (=Namoline), MS142) alone or in combination with ATRA.

AW69 and AW84 reduced the short-term proliferation of MN1 cells by a maximum of 50% at day 3 of treatment, while the proliferation of Hoxa9/Meis1 cells was not affected by any of the inhibitors. When analyzing the morphology of Hoxa9/Meis1 cells after treatment with AW69, AW84 or high-doses of tranylcypromine we observed signs of granulocytic differentiation. Flow cytometric analysis showed an increase of Mac1 and a reduction in CD117 (C-kit) after treatment with AW69 in Hoxa9/Meis1 and an increase in the expression of FcERI in MN1 cells. Colony formation of Hoxa9/Meis1 cells treated with AW69 for 96hrs was reduced by 70% compared to control cells. Treatment with AW69 and AW84 also enhanced the response of leukemia cells to ATRA by increasing the anti-proliferative effect and by reducing colony formation by 70% in MN1 and by 99% in Hoxa9/Meis1 cells in the combination. The two reversible LSD1 inhibitors MS120 and MS142 exhibited only minor effects on differentiation even when used at high concentrations.

We conclude that pharmacological inhibition of LSD1 by tranylcypromine and its derivatives induces differentiation in Hoxa9/Meis1 cells and in MN1 cells. It is now planned to assess the effect of LSD1 inhibitors on leukemic stem cells and in vivo. This project will allow to select new LSD1 inhibitors for treatment and facilitate molecular studies investigating the role of LSD1 in the development of AML.

Disclosure: No conflict of interest disclosed.

P172 - Dronabinol leads to disease control via release of differentiation blockage in an elderly patient with secondary acute leukemia

Kampa-Schittenhelm K.1, Kanz L.1, Schittenhelm M.M.1

1Universitätsklinikum, Tübingen, Germany

Dronabinol displays considerable growth-inhibiting antitumor efficacy in several tumor models and we have recently shown a direct proapoptotic effect in native leukemia patient blasts ex vivo. As we discuss separately, we have evidence that these effects are at least in part mediated via epigenetic hypomethylation (separate abstract provided).

Here we report on a multimorbid elderly patient with secondary acute myelomonocytic leukemia treated with dronabinol for best supportive care considerations due to tumor kachexia and emesis. After initial cytoreduction with hydroxyurea no further specific cytoreductive therapy was administered and the patient was referred to palliative home care. Tantalizingly, while still taking dronabinol, the patient recovered and presented two months later with normal blood counts and absence of leukemia blasts.

To confirm that clinically relevant doses are achievable in vivo, we set up a plasma inhibitory assay and cultured referenced Jurkat cells in serum of this patient. A strong proapoptotic effect of approx. 50% was noted after 72 hours of incubation.

Further follow up of this patient revealed a steadily increasing monocyte count, suggesting leukemia relapse. However, cytomorphology confirmed adequately differentiating granulo- and monocytes and immunophenotyping revealed absence of CD34+ blasts (at diagnosis entire population reacting), while now being highly positive for CD13, CD14 and CD11c - indicating maturing cells. Interestingly in a leukemia mutation screen, an ASXL1 mutation was detected in this population, strengthening the hypothesis that this maturing population stems from the former leukemia population. Notably, ASXL1 interacts with OGT, which is upregulated upon dronabinol (discussed in more detail in a separate protocol). In theory, upregulation of OGT may circumvent inactivating mutations of ASXL1 - and thus provide a stimulus for differentiation.

Conclusions: Clinically active antileukemic doses of dronabinol can be achieved in vivo. Our findings provide a strong rationale for further exploration of dronabinol as an agent with remarkable antileukemic efficacy. Whether specific cohorts may in particular benefit from dronabinol (such as mut-ASXL1) needs to be addressed in future studies.

Disclosure: No conflict of interest disclosed.

P173 - Response-adapted sequential azacitidine and induction chemotherapy in patients > 60 years old with newly diagnosed AML eligible for chemotherapy (RAS-AZIC): Results of the phase I of the DRKS00004519 study

Jäkel N.1, Hubert K.1, Krahl R.1, Cross M.1, Niederwieser D.1, Al-Ali H.K.1

1University Hospital of Leipzig, Department of Haematology/Oncology, Leipzig, Germany

Outcome in elderly patients (pts) with AML after intensive chemotherapy (IC) remains unsatisfactory. The hypomethylating agent azacitidine (AZA) prolongs OS in elderly pts even with >30% marrow blasts. But it is unlikely that a single agent could accomplish the goal of attaining rapid responses and translating them into long-term survival in all pts because of the heterogeneity of the disease. Thus, integrating epigenetic therapy and IC in well-designed trials might further optimize outcome in elderly patients. The feasibility of priming with AZA prior to IC was studied in the phase I of the DRKS00004519 (RAS-AZIC) study.

Patients and methods: RAS-AZIC is a prospective, multicentric, phase I/II trial evaluating the feasibility (phase I) and efficacy (phase II) of priming with AZA followed by a sequential, and response-adapted therapy with AZA or IC in eligible pts > 60 years with newly diagnosed AML. The safety and the dose of priming with AZA 75 mg/m2/day s.c. for 5 (level 1) or 7 days (level 2) followed by IC on day 17 (Mitoxantrone/cytarabine) was established through a 3+3 design. The level at which not more than one of 6 pts experienced a dose limiting toxicity (DLT) would be used in the phase II. DLT was defined as an event at least possibly related to trial therapy within the first 56 days as following: Grade ≥ 3 liver or renal toxicity, or delay in WBC regeneration (≥ 1×109/L) beyond day 45 after IC.

Results: No DLT was documented in both dose level 1 and 2. The phase I part was completed after enrolling 9 pts. Median age was 71 years. Secondary/therapy-related AML, and unfavourable cytogenetics were 56%, and 33% respectively. Baseline median marrow blasts were 44%. AZA priming was well-tolerated and given as out-patient therapy in 67%. In 8/9 pts IC could be started as per protocol. Median WBC regeneration > 1 x109/L following IC occurred after 24 days. No death till day 90 ensued. CR/CRi on day 56 was 78%. After a median follow-up of 13 months, median survival was not reached.

Conclusions: The results of priming with AZA for 7 days followed by IC on day 17 in elderly pts with newly diagnosed AML were encouraging in terms of safety and efficacy. The phase II part is now actively enrolling.

Disclosure: Nadja Jäkel: No conflict of interest disclosed. Haifa Kathrin Al-Ali: Financing of Scientific Research: honoraria and research funding.

P174 - Leukemic cells from patients with acute myeloid leukemia (AML) functionally inhibit healthy CD34+ hematopoietic stem and progenitor cells (HSPC)

Jäger P.1, Cadeddu R.-P.1, Zilkens C.2, Fenk R.1, Germing U.1, Kobbe G.1, Haas R.1, Schroeder T.1, Geyh S.1

1Klinik für Hämatologie, Onkologie und Klinische Immunologie, Universitätsklinikum, Düsseldorf, Germany, 2Orthopädische Klinik, Düsseldorf, Germany

Introduction: Hematopoietic insufficiency is the hallmark of AML with cytopenia-related complications representing major causes of death. Albeit recent experiment evidence pointed towards an important role of the bone marrow (BM) microenvironment, the underlying mechanisms mediating hematopoietic insufficiency are still insufficiently understood. Since the BM of patients with AML is dominated by leukemic blasts accompanied by a reduction of normal CD34+ HSPC, we reasoned that leukemic cells might directly suppress normal HSPC.

Methods: To experimentally address this hypothesis we modelled the situation of BM infiltration in vitro by exposing healthy BM-derived CD34+ HSPC to supernatants derived from leukemic cells. Conditioned media (CM) were harvested from 3 AML cell lines (THP-1, HL-60, MV4-11) as well as from a total of 23 newly-diagnosed AML patients covering all relevant WHO subtypes after 3 days of cultivation. Healthy CD34+ HSPC were incubated for 3 days in the presence of leukemic or control media. Subsequently, proliferation, cell cycle behaviour and differentiation of these CD34+ HSPC was investigated using cell counting, dye staining with Ki-67 and Hoechst 33342 as well as semisolid clonogenic assays.

Results: Exposure to conditioned media derived from AML cell lines and primary patients samples significantly inhibited proliferation as indicated by a profound reduction of viable healthy CD34+ HSPC. Complementary with this, we observed a clear shift of the cell cycle state of healthy CD34+ HSPC towards a resting phenotype when cultivated in AML-derived media with the majority of cells being in inactive G0 phase. Performing semisolid clonogenic assays demonstrated a strikingly lower colony-forming capacity of CD34+ HSPC following incubation with AML-derived supernatants.

These inhibitory effects on healthy hematopoiesis were markedly related to the CD34+ leukemic cell population, but not to the MNC fraction as indicated by a comparison of paired MNC and CD34+ immunomagnetically enriched AML samples. PCR-screening of well-known negative regulators of hematopoiesis revealed a significant overexpression of TGF beta suggesting a potential role of this candidate molecule for suppression of healthy hematopoiesis by leukemic cells.

Conclusion: Overall, these data indicate that leukemic cells mediate direct suppressive effects on important functions of healthy CD34+ HSPC thereby contributing to hematopoietic insufficiency in AML.

Disclosure: No conflict of interest disclosed.

P175 - CD33-BiTE® antibody construct mediated lysis of AML cells is influenced by the choice of the pretherapeutic cytoreductive agent in vitro

Brauneck F.1,2, Krupka C.1,2, Lichtenegger F.S.1,2, Kufer P.3, Kischel R.3, Zugmaier G.3, Köhnke T.1,2, Altmann T.1,2, Schneider S.1, Fiegl M.1, Spiekermann K.1,4, Hiddemann W.1,4, Subklewe M.1,2,4

1Department of Internal Medicine III, Klinikum der Universität München, Munich, Germany, 2Clinical Co-operation Group Immunotherapy at the Helmholtz Institute, Munich, Germany, 3AMGEN Research (Munich) GmbH, Munich, Germany, 4German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ), Heidelberg, Germany

Introduction: In our previous work, we demonstrated that the CD33-BiTE® antibody construct AMG 330 is able to induce activation and proliferation of autologous T cells and effectively mediates lysis of primary AML cells.

We hypothesized that in AML patients with high leukocyte counts a cytoreductive phase prior to AMG 330 therapy might be beneficial to increase the E:T ratio and to reduce cytokine mediated toxicity. Ideally, the cytoreductive drug does not impair T-cell function.

Methods: We evaluated the effect of cytarabine (20µM), azacitidine (1µM, 5µM) and hydroxyurea (10µM, 100µM, 1000µM) on T-cell proliferation and function in close analogy to potential treatment algorithms for AML. Healthy donor T cells were pre-incubated with the cytoreductive drugs for 72 hours and subsequently co-cultured with HL60 cells with or without AMG 330. After 3 days, AMG 330 mediated lysis of AML cells and T-cell proliferation were assessed by flow cytometry.

Results: Pretreatment of T cells with cytarabine completely abrogated T-cell function (lysis of HL60 cells: untreated (no tx): 98.2% vs 20µM: 7%) and significantly impaired T-cell proliferation (no tx: 57.4% vs 20µM: 4.3%, n = 3). These findings correlated to data using primary AML samples collected 3 and 6 days after discontinuation of cytarabine treatment in vivo. After a 3-day chemotherapy-free interval, we observed no relevant T-cell proliferation and lysis of AML cells upon addition of AMG 330 to the ex-vivo long-term culture system (lysis of AML cells on day 12: 30%; fold change T-cell expansion 0.9). After a 6-day treatment-free interval, high T-cell proliferation and cytotoxicity were observed (lysis of AML cells on day 12: 61%; fold change T-cell expansion: 3.1).

Azacitidine treatment only marginally impaired T-cell function (lysis of HL60 cells: no tx: 100% vs 1µM: 94.9% vs 5µM: 86.8%; proliferation: no tx: 90.9% vs 1µM: 80% vs 5µM: 66.8%). Pretreatment with hydroxyurea had the least impact on T-cell performance. It did not impair T-cell function (lysis of HL60 cells: no tx: 99.9% vs 10µM: 96.5% vs 100µM: 97.7% vs 1000µM: 99.9%) and proliferation compared to untreated controls (no tx: 79.5% vs 100µM 74.6% vs 10µM 75.2% vs 1000µM: 81.9%, n = 3).

Conclusion: As the BiTE® antibody construct technology essentially relies on T-cell function chemotherapeutic approaches need to be carefully evaluated. Our data support the use of hydroxyurea in AML patients that require cytoreduction prior to AMG 330 treatment.

Disclosure: Marion Subklewe: received research funding from AMGEN Research (Munich) GmbH

P176 - The gain-of-function KIT D816V mutation associated with systemic mastocytosis (SM) is sensitive towards the FLT3 inhibitor crenolanib (Cb) and sensitizes cells towards cladribine (2-CdA) - a rationale to combine 2-CdA with Cb

Kampa-Schittenhelm K.1, Frey J.1, Ramachandran A.2, Kanz L.1, Schittenhelm M.M.1

1Universitätsklinikum, Tübingen, Germany, 2AROG Pharmaceuticals Inc., Dallas, United States

Activating D816 mutations of the class III receptor tyrosine kinase KIT are associated with the majority of patients with SM, making mutant-KIT an attractive target using tyrosine kinase inhibitors (TKI) - however, so far clinical efficacy of TKI fell short of the expectations. Cb is a novel potent FLT3 inhibitor with decent co-activity against the imatinib-resistant KIT D816 isoforms (Kampa-Schittenhelm DGHO 2014), however combination regimens might increase clinical efficacy. We have evidence that gain of the D816V enzymatic pocket type mutation within the tyrosine kinase domain 2 (TK2) sensitizes mast cells to 2-CdA - making it an attractive compound to combine with Cb.

HMC1.1, HMC1.2 and the p815 mastocytosis cell lines were treated with Cb and/or 2-CdA. Cell cycle analyses and induction of apoptosis were performed flow cytometrically. Antiproliferative efficacy was measured using XTT-based assays. Tyrosinphosphorylation of KIT and downstream signaling was assessed by western immunoblots. For synergy analysis isobolograms were computed using Calcusyn software.

HMC1.1 cells, harboring a regulatory KIT V560G mutation, treated with 2-CdA in a dose-dependent manner demonstrated a potent antiproliferative effect in the hundred nanomolar ranges - surprisingly, no significant proapoptotic effect was observed. We therefore evaluated time- and dose-dependent cell cycle effects and observed a dose-independent permanent G2/M phase arrest. Consequently, combination of 2-CdA with Cb (which accumulates cells in G1/G0 as most class III TKI) did not enhance proapoptotic effects. Interestingly, the sister cell line HMC1.2, harboring an additional D816V mutation displayed cell cycle independent high antiproliferative as well as proapoptotic sensitivities towards 2-CdA. High proapoptotic efficacy for 2-CdA was confirmed in murine p815 cells (harboring a D814Y mutation, corresponding to the D816 codon in humans). Combination of 2-CdA with Cb resulted in an additive proapoptotic effect, compared to either substance alone. It has been shown, that knockdown of p21 leads to resistance towards 2-CdA in colorectal tumor models. Tantalizingly, p21 is regulated via the PI3K/AKT pathway, which is directly activated via the KIT TK2 domain, providing a mechanistic model for our observation - in detail currently evaluated by our group.

To summarize, we provide a rationale to combine 2-CdA and Cb in KIT D816 positive SM. Clinical evaluation is warranted.

Disclosure: No conflict of interest disclosed.

P177 - The Adult Comorbidity Evaluation-27 (ACE-27) score is associated with survival in patients with acute myeloid leukemia

Hitz F.1, Müller-Tidow C.1, Müller L.P.1, Wass M.1

1Universitätsklinik und Poliklinik für Innere Medizin IV, Universitätsklinikum Halle, Halle (Saale), Germany

Introduction: The role of comorbidities as an independent prognostic factor for patients with AML is ill defined. The ACE-27 comprises a tool to assess comorbidities.

The aim of this study was to determine the impact of pretreatment comorbidities categorized by ACE-27 on the survival of patients with AML.

Methods: In a single-center retrospective study the ACE-27 was obtained for 194 adult AML patients (median age 61y) treated with intensive chemotherapy between 1996 and 2012. Data on demographics, cytogenetics according to ELN and outcome were collected. Kaplan-Meier methods and Cox regression were used to assess survival.

Results: ACE-27 was 0 (none) in 20% of patients, 1 (mild) in 25%, 2 (moderate) in 38%, and 3 (severe) in 16%. The most frequent comorbidities were cardiovascular disease (60%), prior malignancy (39%) and diabetes mellitus (16%). The median overall survival was 17 months. However, it was associated with ACE-27 risk groups with 96, 18, 14 and 8 months for patients with none, mild, moderate and severe comorbidities respectively (p=.006). In univariate analysis, cardiovascular disease and renal impairment were associated with inferior survival (26 vs 12 months, p=.005; 17 vs 7 months, p=.016). Interestingly complete remission (CR) rates differed versus risk groups: 80%, 79%, 63% and 43% for patients with none, mild, moderate and severe comorbidities, respectively (p=.002). Also early death rate differed between patients with none or mild vs moderate or severe comorbidities (6% vs 14%). Multivariate analysis showed that higher ECOG score (HR 2.9, p=.003), poor cytogenetics (HR 3.8, p=.007) and ACE-27 ≥ 2 (p=.002) had a significant impact on overall survival. Multivariate adjusted HR were 2.1, 2.6 and 4.5 for mild, moderate and severe comorbidities, respectively, compared with no comorbidities. Interestingly, higher age (≥ 60 years) had no significant impact in multivariate analysis when comorbidities based on ACE-27 were included.

Conclusion: Comorbidities assessed by ACE-27 seem to have a significant impact on survival of patients with AML. Patients with severe comorbidity had a greater than 50% decrease in survival, independent of age, ECOG and cytogenetics. A pretreatment assessment of severity of comorbidities according to the ACE-27 may help to identify patients with poor outcome. It appears that age at least partially impacts survival as a marker for comorbidities.

Disclosure: No conflict of interest disclosed.

P178 - Resistance to sorafenib in acute myeloid leukemia with FLT3-ITD may be overcome by down-regulating the nuclear repressor Ski by inhibitors of histone deacytelases

Jehn L.B.1, Frech M.1, Metzelder S.K.1, Teichler S.1, Stabla K.1, Neubauer A.1

1Department of Hematology, Oncology and Immunology, Philipps University of Marburg and University Hospital of Giessen and Marburg, Marburg, Germany

Acute myeloid leukemia (AML) is genetically a heterogenous disease. Besides age, cytogenetic and molecular aberrations are the most important prognosticators. AML with monosomy 7 or deletion of 7q (-7/del7q) has a poor prognosis and is associated with an up-regulation of the nuclear oncogene SKI. AML with -7/del7q reveal an up-regulation of SKI expression as microRNA29a, encoded on 7q32, regulates SKI-expression (Teichler et al., Blood 2011). Ski protein represses all-trans retinoic acid (ATRA) signaling and myeloid differentiation in AML cells in vitro via interacting with N-CoR (nuclear receptor corepressor) and cooperating with histone deacetylases (HDAC). This inhibitory effect of wild-type Ski can be abrogated by the HDAC blocking agent valproic acid (VPA) (Ritter et al., Leukemia 2006). Activating internal tandem duplication (ITD) mutations in the fms-like tyrosine kinase 3 (FLT3) gene (FLT3-ITD) with normal cytogenetics are associated with poor outcome in AML (Thiede et al., Blood 2002). The therapeutic options for relapsed or refractory FLT3-ITD-positive AML are limited, particularly in case of prior allogenic stem cell transplantation (SCT) or poor performance status. Here, the use of the multi-targeted tyrosine kinase inhibitor sorafenib is an effective treatment option with high response rates (Metzelder et al., Blood 2009 and Leukemia 2012). However, the development of drug resistance to this targeted intervention remains an important clinical problem. Statistical analyses of patients (n = 14) with normal karyotype and positive for the FLT3-ITD mutation revealed a significantly shorter survival with a high Ski protein level compared with patients with a low Ski protein level. We further have investigated the influence of HDAC inhibitors (HDACi) on sorafenib-sensitive and -resistant FLT3-ITD-positive AML cells with differing levels of Ski expression in vitro. Cells were treated with increasing concentrations of different HDACi. Cell viability was measured using trypan blue staining and MTT assay. Western analysis was performed to investigate changes in Ski expression during HDACi treatment. We found that high expression of Ski in sorafenib-resistant FLT3-ITD-positive cells was predictive of better responses to treatment with HDACi in vitro. In conclusion, the combined use of HDACi and multi-targeted tyrosine kinase inhibitors like sorafenib in relapsed or refractory FLT3-ITD-positive AML with high Ski expression deserves further consideration.

Disclosure: No conflict of interest disclosed.

P179 - Prospective functional and patient-reported outcomes study of AML patients aged >60 years receiving induction chemotherapy (INCIDER trial): first feasibility analysis

Maurer H.1, Müller M.J.1, Ihorst G.2, Bogatyreva L.1, Lübbert M.1

1Universitätsklinikum Freiburg, Klinik für Innere Medizin I, Freiburg, Germany, 2Albert-Ludwig-Universität Freiburg, Department für Medizinische Biometrie und Medizinische Informatik, Freiburg, Germany

It is increasingly accepted that not only disease but also host-specific factors contribute to the treatment tolerance and outcome in elderly AML/MDS patients (pts). We could previously show that patient-reported outcome parameters can predict overall survival of AML/MDS patients >60 year unfit for standard chemotherapy and therefore receiving non-intensive treatment or sole best supportive care (Deschler et al. 2013). Only very few studies have successfully shown that assessments also in AML pts fit for induction chemotherapy can yield prognostic parameters, e.g. Klepin et al. (Blood 2013). The INCIDER prospective study aims at defining functional and patient-reported outcomes parameters that might predict overall survival in this patient group.

Methods: Since 5/2012, all AML pts >60 years receiving induction chemotherapy at our center were screened for a pre-treatment assessment and up to 3 subsequent assessments. The following physician-assessed instruments were applied: G8 (geriatric symptoms), ECOG performance status, Barthel-index (Activities of daily living), HCT-CI (comorbidity burden). Patient-reported outcomes instruments: EORTC-C30 (quality of life), HADS (anxiety, depression), RS-11 (psychological resilience). Tests were applied by a team of 2 physicians and a study nurse.

Results: 34 consecutive pts met the eligibility criteria for the study, of whom 25/34 (74%) were successfully assessed prior to induction (timepoint T0). Another 9/34 pts were not enrolled, reasons were as follows: 3 declined participation due to cognitive overload, 3 had rapidly progressive disease and evolving cognitive disability, 3 were not approached. Feasibility of subsequent assessments: at timepoint T1 (at 3 months) 15 of the 19 pts still alive were assessed; timepoint T2 (at 6 months): 12/14; timepoint T3 (at 12 months): 5/9. Thus the main drop-out reason was death.

Conclusions: This feasibility analysis demonstrates that the assessment tests chosen were well accepted by most of the pts, and were feasible despite treatment initiations also on weekends. Also pts with rapidly progressive disease and difficult clinical conditions could be included by the team. Thus in this single-center pilot trial, patient compliance and logistics allowed for successful assessment of the majority of eligible pts. The study will be continued with particular attention to further improvement of patient coverage and timeliness of pre-therapy assessments, and expansion to more centers.

Disclosure: No conflict of interest disclosed.

P180 - Successful treatment of acute promyelocytic leukemia in pregnancy with single agent ATRA

Nellessen C.1, Mayer K.M.1, Merz W.M.2, Flöck A.2, Brossart P.1, Janzen V.1

1Medizinische Klinik III, Hämatologie-Onkologie, Uniklinik, Bonn, Germany, 2Universitätsfrauenklinik, Abteilung für Geburtshilfe und Pränatalmedizin, Uniklinik, Bonn, Germany

Introduction: The diagnosis of acute promyelocytic leukemia (APL) in pregnancy is an uncommon, life-threatening emergency. Choice of treatment and management of complications is challenging, especially as published data are rare.

Case: A 41-year old woman was referred to our emergency department at 25 weeks of gestation with spontaneous hematoma and pancytopenia. Diagnosis of APL was made by bone marrow cytology and confirmed by PCR. A delivery at that time was judged to be of unacceptable risk to the mother due to the presence of severe, APL-associated coagulopathy. Therefore, a joint decision was made to immediately start induction therapy with all-trans retinoic acid (ATRA) supplemented by dexamethasone to prevent ATRA differentiation syndrome. Induction treatment was continued with ATRA alone without addition of anthracyclines with the intention to minimize the risk of toxicity to the fetus. During the entire induction cycle no signs of differentiation syndrome were observed. Transfusion with red blood cells and platelets as well as substitution with clotting factors within the first few days was necessary and sufficient to prevent major bleeding incidents. At day 32 after initiation of therapy the blood cell counts had normalized, and the patient was discharged from hospital with continuation of ATRA therapy.

Fetal wellbeing was assessed regularly by ultrasound and Doppler examination. Delivery was scheduled for 34 weeks of gestation and an apparently healthy female baby was delivered. Remission control of peripheral blood and bone marrow after delivery (day 72 of ATRA treatment) showed complete cytomorphologic and molecular remission. We immediately proceeded with the first consolidation course of ATRA and arsenic trioxide. Currently (May 2015), the patient is in her third consolidation course, remaining in complete remission.

Conclusion: This case illustrates that treatment with ATRA alone in the second half of pregnancy was safe for both mother and fetus, achieving complete remission of APL and delivery of a healthy newborn. Close surveillance of both, mother and fetus, and management of disease related complications, and interdisciplinary teamwork with obstetricians was mandatory for successful outcome.

Disclosure: No conflict of interest disclosed.

P181 - Metabolic reprogramming of acute myeloid leukemia blasts by bone marrow stromal cells

Braun M.1, Jitschin R.1, Mackensen A.1, Mougiakakos D.1

1Universitätsklinikum Erlangen; Hämatologie und Onkologie, Erlangen, Germany

Acute myeloid leukemia (AML) represents the most common form of acute leukemia in adults. Despite the enormous efforts during the last decades treatment resistance is still observed at a high rate. Previous studies have shown that bone marrow stroma promotes an increased resilience of AML blasts towards chemotherapeutics. Furthermore, current data suggest that alterations of the malignant cells' metabolism could represent a strong determinant for the disease's (including AML) course and/or treatment resistance. In fact, a deregulated metabolism could lead to a reduced sensitivity towards therapy and it remains to be elucidated whether this is a mechanism contributing to the blast-protective effects elicited by the bone marrow stroma. Here, we sought out to characterize the impact of stroma cells on the AML blasts' metabolism.

The human bone marrow stromal cell line HS-5 was utilized for establishing the in vitro niche model. We compared in our assays AML cell lines as well as primary blasts cultured on a HS-5 monolayer or alone. In line with previous observations we could detect an increased proportion of AML cells in the S-phase of the cell cycle upon co-culture with HS-5. When evaluating the cells' metabolism we observed a shift towards glycolysis despite presence of oxygen, i.e. aerobic glycolysis or the “Warburg”-effect. Basal glycolysis as well as maximal glycolytic capacity upon blocking ATP production in the mitochondrial respiratory chain was increased. Respiration was not significantly affected. However, mitochondrial biogenesis appeared reduced.

Increased glycolysis was accompanied by an increased uptake of fluorescently labeled glucose as well as an increased expression of glucose transporters. The expression of several glycolytic molecules found to be increased upon HS-5 co-culture. Noticeably, cell-to-cell contact was not a pre-requisite for the metabolic shift. Our data was further corroborated by direct observations from AML patients: AML blasts isolated from the bone marrow exhibited an upregulated glycolysis as compared to their counterparts from the periphery collected at the same time point. Taken together, our data indicates a stromal cell-mediated metabolic shift in AML blasts towards aerobic glycolysis. This metabolic phenotype is linked to an unfavorable prognosis and increased chemo resistance. The underlying molecular pathways remain to be elucidated and could represent a promising target for future interventions.

Disclosure: No conflict of interest disclosed.

P182 - Blastic plasmacytoid dendritic cell neoplasm: A very rare, unfamiliar and scary disease

Schmidt A.1, Neuberger C.2, Schmid M.1, Lienhard R.3

1Stadtspital Triemli, Internal Medicine, Zürich, Switzerland, 2Stadtspital Triemli, Pathology, Zürich, Switzerland, 3Spital Affoltern am Albis, Internal Medicine, Affoltern am Albis, Switzerland

Introduction: Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a very rare haematological disease currently categorized as an entity of “acute myeloid leukaemia and related precursor neoplasm” in the WHO Classification of 2008.This malignancy is characterized by CD4+/CD56+/CD123+ immunophenotype, aggressive course and poor prognosis. BPDCN often initially presents with solitary or multiple heterogenic skin lesions, accompanied by regional lymph node involvement. In advanced stages of disease, bone marrow infiltration and leukemic transformation occurs regularly.

BPDCN is usually diagnosed by immunohistological staining. Therapeutic approaches are heterogeneous and despite high response rate to first line therapy, early relapse with disease progression and dismal prognosis is the rule.

Methods: Based on an own case of an 71 year old man with at the beginning only cutaneous manifestation of BPDCN, we made an extensive literature research in PubMed for articles in several languages relating to small series and case reports of BPDCN. We reviewed 70 articles from 1997 to now considering initial clinical manifestation, chosen therapeutic regimen, evolution and survival. Special attention was laid to the time of starting therapy and use of novel substances like bortezomib.

Results: Reviewing available literature (containing about 400 patients) confirmed BPDCN as a very rare disease of any age with heterogeneous clinical and histopathological presentation, often leading to misdiagnosis. Currently the only curative approach, showing long-term remission in selected cases, is allogeneic stem cell transplantation. Most patients were treated with ALL- or AML-like chemotherapy-regimen, associated with high response but early relapse rate, with a median survival of only approximately 12 months. Overall, evidence is scarce and newer drugs barely used. In our case, delaying start of therapy until needed and addition of Bortezomib to a conventional R-CHOP-regimen didn't improve prognosis.

Conclusion: BPDCN is a very rare disease with commonly inhomogeneous cutaneous manifestation that typically occurs in elderly patients. Currently no standardised treatment exists and early relapse with fulminant deterioration affects majority of patients. Further investigations and registries are needed to better understand pathophysiological processes of BPDCN and develop diagnostic and therapeutic guidelines to reduce misdiagnosis and improve long-term remission rate.

Disclosure: No conflict of interest disclosed.

P183 - Acute promyelocytic leukemia resulting from a cryptic insertion responds well to ATRA plus ATO therapy: a case report

Hecht A.1, Nolte F.1, Fabarius A.1, Haferlach C.2, Hofmann W.-K.1, Lengfelder E.1

1Universitätsklinikum Mannheim, Hämatologie und Internistische Onkologie, Mannheim, Germany, 2MLL Münchner Leukämie Labor, München, Germany

Introduction: Normally, diagnosis of acute promyelocytic leukemia (APL) is based on the pathognomonic finding of the characteristic translocation t(15;17) resulting in the PML-RARA fusion gene. Submicroscopic cryptic insertions are rare abnormalities that can be found in approximately 3.6% of APL cases only (Grimwade et al., Blood 2000). Therefore, results on treatment response and outcome are solely based on case reports.

Here we present a case of APL due to a cryptic insertion with an unusually favorable initial presentation and good response to ATRA plus ATO.

Case report: In August 2014 an inquiry reached our international office concerning the treatment of a Ukrainian patient with APL. One week later a 27 year old woman presented with a history of prolonged fatigue. Diagnosis of APL had been based on a bone marrow (BM) smear one week before administration to our hospital. Though the patient had not undergone any kind of treatment, she did not present with any symptoms of bleeding or coagulation dysregulation. Laboratory results showed leukocyte count of 2,400/µl with no peripheral promyelocytes, hemoglobin level of 12.3 g/dl and platelet count of 206,000/µl. Mild hypofibrinogenemia and subtly elevated levels of d-dimers were the only hints at coagulopathy. The initial BM smear showed a total infiltration by promyelocytic blasts with multiple auer rods. On the day of administration we performed another BM biopsy to confirm the results. Cytomorphology showed a 50% proportion of abnormal promyelocytes. Molecular genetic analysis proved the presence of the PML-RARA fusion transcript (long isoform). However, cytogenetic analysis showed a normal female karyotype. At last, FISH analysis revealed a submicroscopic insertion of the RARA locus on chromosome 17 into the PML locus on chromosome 15 (ins(15;17)(q24;q21q21).

The patient started treatment with ATRA and ATO according to the APL0406 protocol and achieved complete remission within the first 28 days. There were no APL or treatment related complications and she is now undergoing her last cycle of consolidation therapy.

Conclusions: To our best knowledge this is the first report on response to the ATRA plus ATO combination therapy in an APL patient bearing a cryptic insertion. Interestingly, this patient also presented with an unusually benign course of APL. This adds knowledge to the few cases reported with this abnormality.

Disclosure: No conflict of interest disclosed.

P184 - Dronabinol reveals antileukemic efficacy via epigenetic upregulation of O-linked β-N-acetylglucosamine transferase (OGT) resulting in release of differentiation blockage and sensitizing cells towards induction of apoptosis

Kampa-Schittenhelm K.1, Salitzky O.1, Akmut F.1, Bonin M.2, Kanz L.1, Schittenhelm M.M.1

1Universitätsklinikum, Tübingen, Germany, 2Universität Tübingen, Medizinische Genetik, Microarray Facility, Tübingen, Germany

Dronabinol, the natural (−)-Δ9-Tetrahydrocannabinol, has growth-inhibiting antitumor efficacy - including acute leukemia. We now reveal a novel mechanism-of-action via epigenetic modulation of OGT, an enzyme linked to genes involved in leukemogenesis such as AKT, MLL5, TET2 or ASXL1, releasing leukemia blasts from differentiation blockage in vivo and sensitizing cells towards induction of apoptosis.

gDNA methylation gene arrays using Jurkat leukemia cells revealed global modulation of methylation patterns upon dronabinol treatment. OGT was identified as the highest altered gene (-42%, pval 3,68E-38) - correlating with an increase of OGT protein expression in Western immunoblots. Consistently, hypomethylation of the transcription start site of OGT and induction of OGT protein expression upon dronabinol were confirmed in an independent array using native patient samples. To study the underlying mechanisms, Jurkat cells were pretreated with CB1 (LY320135) and/or CB2 (JTE-907) antagonists, and exposed to dronabinol. Inhibition of either receptor reduced induction of apoptosis - and was most profound when inhibiting both receptors simultaneously. Importantly, upregulation of OGT protein expression upon dronabinol was suppressed by inhibition of either receptor, arguing for a receptor-mediated epigenetic effect of dronabinol. Retroviral knockdown of OGT in Jurkat and native leukemia blasts rendered cells less susceptible towards induction of apoptosis.

We had the chance to follow a patient with secondary acute myeloid leukemia who was supportively treated with dronabinol for tumor kachexia and emesis. Direct disease control linking to dronabinol was noted (case report provided as separate abstract). Tantalizingly, besides induction of apoptosis upon dronabinol, the leukemic clone was maturing - overriding the differentiation blockage. Mimicking this observation, we treated cells of this and other patients with dronabinol ex vivo and revealed loss of CD34 and upregulation of CD11c by flow cytometry - again a sign for maturation. Interestingly this differentiation was abrogated by lentiviral OGT-interference arguing for a role of OGT in overriding the differentiation blockage in acute leukemia.

Conclusions: Our findings provide a strong rationale for further exploring dronabinol as an agent with remarkable antileukemic efficacy achievable in vivo. In specific, overriding the differentiation blockage in leukemia cells may open up alternative therapeutic approaches.

Disclosure: No conflict of interest disclosed.

P185 - Expression of inhibitory (i)ASPP (Apoptosis Stimulating Protein of p53) in acute myeloid leukemia (AML)

Schittenhelm M.M.1, Bajrami Saipi M.1, Kanz L.1, Kampa-Schittenhelm K.1

1Universitätsklinikum, Tübingen, Germany

Inactivation of the p53 pathway is a frequent event in human cancers promoting tumorigenesis and resistance to chemotherapy. Inactivating p53 mutations are uncommon in non-complex karyotype leukemias suggesting that the p53-pathway must be inactivated by other mechanisms. ASPP proteins are a family of p53-binding proteins that consists of three members: the pro-apoptotic ASPP1 and ASPP2 proteins and the apoptosis-inhibiting iASPP. All play a crucial role in the regulation of p53-induced apoptosis.

We have shown that ASPP2, but not ASPP1, is an independent haploinsufficient tumor suppressor in vivo and is upregulated upon cellular stress. Further, ASPP2 is significantly attenuated in AML - and this uniquely links to the patient cohort failing induction chemotherapy. We now study changes of ASPP2 expression and its counterplayer iASPP during chemotherapy to evaluate whether ASPPs may provide a useful tool to predict and monitor therapy response.

ASPP2 as well as iASPP mRNA levels were determined using standard qRT-PCR. Protein expression was measured using flow cytometry and immunoblot assays. Basal levels in bone marrow samples as well as in peripherel blood samples of so far 25 leukemia patients were normalized to a control cohort comprising 16 healthy bone marrow donor samples. Blood samples were obtained before and during initial therapy at defined timepoints (12, 24, 48 and 72 hours) and expression levels of ASPP2 and iASPP were measured and correlated with blast count and initial therapy response.

We confirmed lower basal ASPP2 mRNA as well as protein levels in AML samples compared to the donor controls. During induction therapy, a proportion of patients revealed adequate upregulation of ASPP2 in vivo. Vice versa, we define a subgroup of patients that fail to upregulate ASPP2 during therapy. iASPP levels were generally higher in the patient compared to the donor cohort and tended to increase at the beginning of therapy. Interestingly, highest iASPP levels were found in patients expressing low ASPP2 levels - and these findings again correlated with poor clinical outcome.

We believe, that different members of the ASPP family play a distinct role in acute leukemia and that complex interactions between the different family members are in part responsible and predictive for therapy outcome.

Further studies are necessary to shed light into the role of iASPP in leukemia and to understand the complex interactions within the ASPP family.

Disclosure: No conflict of interest disclosed.

P186 - Accelerated senescence of MSC from patients with high risk ALDH-numerous AML

Horn P.1, Hoang V.T.1, Hoffmann I.1, Benes V.2, Wuchter P.1, Ho A.D.1

1Universitätsklinikum Heidelberg, Innere Medizin V, Heidelberg, Germany, 2European Molecular Biology Laboratory, Genomics Core Facility, Heidelberg, Germany

The microenvironment in the bone marrow (BM) has significant effects on maintenance, regulation, and safeguarding of hematopoietic stem cells (HSC), both in health and development of leukemia. We have compared mesenchymal stromal cells (MSC) from patients with acute myeloid leukemia (AML) vs. healthy donors as surrogate niche models to analyze the environment of the leukemic stem cell niche. We compared gene expression profiles of AML samples derived from patients based on the expression levels of aldehyde dehydrogenase (ALDH) activity.

MSC were isolated from human BM samples of healthy donors and AML patients with newly diagnosed AML with low frequencies of ALDH positive blasts (below median value of 1.92% for healthy bone marrow; ALDH-low) or high frequencies of ALDH positive cells (above 1.92%; ALDH-numerous). We have shown that the latter group of patients is associated with high-risk for relapse and poor clinical outcome. Cells were expanded for initial characterization, analysis of CFU-F frequency and long-term culture. RNA of early and senescent passages after long-term culture was used for gene expression profiling.

The majority of AML-MSC displayed a senescent phenotype and shape, decreased CFU-F frequency, and a significantly decreased proliferation capacity, specifically those from ALDH-numerous AML. Comparison of genome profiles of MSC from patients with ALDH-numerous vs. ALDH-low AML showed differential expression of 1031 genes, 369 up- and 662 down-regulated. Of these over 62% could be directly related to senescence upon long-term cultures of MSC from healthy subjects, connected with GO-categories of various malignant disorders, and translation and gene Expression, 203 gene products being localized in the nucleus. Comparison of gene expression profiles of early and senescent cell passages of healthy MSC vs. early passages of AML-MSC showed a specific clustering of MSC derived from patients with ALDH-numerous AML with senescent passages of healthy donors. Profiles of MSC derived from patients with ALDH-low AML were similar to early passages of healthy donors.

These data provide evidence that the microenvironment is involved in the malignant process of AML. The poor prognosis of ALDH-numerous AML is reflected by an accelerated senescence status and a characteristic gene expression profile of MSC derived from such patients. Experiments are ongoing to identify the roles of specific genes for the accelerated aging process in high risk AML.

Disclosure: Patrick Horn: No conflict of interest disclosed. Anthony D. Ho: Advisory Role: Genzyme / Sanofi-Aventis; Financing of Scientific Research: Genzyme / Sanofi-Aventis.

P187 - Therapy related acute promyelocytic leukaemia - presentation of two cases

Hoffmann F.1, Graul K.1, Heyn S.1, Franke G.-N.1, Schwind S.1, Jentzsch M.1, Pönisch W.1, Al-Ali H.K.1, Wang S.Y.1, Niederwieser D.1, Vucinic V.1

1Leipzig University, Hämatologie und Internistische Onkologie, Leipzig, Germany

Introduction: Therapy related acute myeloid leukaemias (t-AML) account for about 10-20% of patients with AML and mostly occurs 5-10 years after treatment with alkylating agents and 2-3 years after treatment with topoisomerase II inhibitors. Here we present two patients with therapy related acute promyelocytic leukaemia (t-APL).

Patients: Two female patients aged 49 and 58 years developed a t-AML after breast cancer treatment with 5-Fluorouracil (5-FU), etoposide and cyclophosphatide followed by two courses of docetaxel and sigma carcinoma treated with 5-FU, leucovorine and oxaliplatine, respectively. Patients presented with clinical symptoms of respiratory or urinary infection in April 2014 and January 2015, respectively. The blood counts in both cases showed a pancytopenia with platelet counts (PLT) above 100 GPT/l, i.e. low risk according to Sanz score. The bone marrow smear showed an infiltration with 40% and 45% blasts, respectively. In the fluorescent in situ hybridization (FISH) the positivity for t(15;17) was shown and confirmed by real time polymerase chain reaction (rt-PCR) method showing the PML-RARA fusion in both cases. The APL developed after 3 and after 5 years from diagnosis of the respective primary tumor. The initial treatment of t-APL in both patients consisted of a combination of all-trans-retinoic acid (ATRA) and an induction chemotherapy with intermediate dose cytarabine on days 1, 3, 5 and 7 and idarubicine on days 1, 2 and 3. Both patients achieved a complete remission (CR). The rt-PCR showed a 3 fold log reduction. In both patients a consolidation chemotherapy, identical to induction, was applied. The first patient showed a prolonged hematological regeneration but hematological CR was confirmed. The rt-PCR analysis showed no sign of disease. The patient is still in complete haematological and molecular remission, 1 year after diagnosis. The second patient is at the moment in neutropenic phase following the consolidation therapy.

Discussion: The t-APL is more common in female patients, and the most common cases are patients with previously treated breast cancer. The t-APL after colonic carcinoma has been described in sporadic cases. In the published cases the t-APL developed shortly after diagnosis and treatment of the primary neoplasia (<3 years). t-AML has an equaly good prognosis as de novo APL, and until today both of the reported patients are in complete hematological remission.

Disclosure: No conflict of interest disclosed.


Der spezielle Fall

P188 - Ruxolitinib as a treatment option for steroid-refractory severe graft-versus-host disease - case report

Paul S.1, Wass M.2, Müller-Tidow C.2, Müller L.P.2

1Universitätsklinikum Halle, Klinik für Innere Medizin IV, Halle, Germany, 2Universitätsklinikum Halle, Klinik für Innere Medizin IV-Hämatologie/Onkologie, Halle, Germany

Introduction: If refractory to steroids, acute as well as chronic GvHD represent a life-threatening condition after allogeneic stem cell transplantation (SCT) for which no treatment standards exist. New data suggest that immunomodulatory drugs like ruxolitinib may be of therapeutic use in this situation.

Case: We report on a 51-year-old male diagnosed with acute myeloid leukaemia with intermediate cytogenetic risk. After standard induction and consolidation therapy complete remission was achieved. In July 2014 myeloablative allogeneic SCT from an HLA-matched related donor was performed. Immunosuppressive therapy comprised cyclosporine and long-course methotrexate without ATG. Upon rapid engraftment the patient developed severe, histologically proven acute GvHD of the skin (stage 4, IV°) on day +30. Therapy with prednisolone (2 mg/kgBW) was initiated without any response seen after 7 days. Therapy was escalated with cyclosporine, mycophenolate mofetil and ATG. Signs of GvHD improved completely but the patient developed pancytopenia, systemic EBV reactivation and septic shock. At day +100 severe GvHD of the skin relapsed. A second course ATG with prednisolone was applied followed by treatment with sirolimus, haploidentical mesenchymal stem cells and PUVA. Again treatment was accompanied by severe neutropenic septicaemia and EBV reactivation. After a short response a progression of cutaneous GvHD ensued. Based on a recent publication (Spoerl et al.: BLOOD 2014;123:3832) we started treatment with ruxolitinib at a dose of 10mg bid. Given the stable blood counts and lack of infections the dose was increased to 25mg bid. Treatment with prednisolone was tapered and on day +180 we observed a clear response with residual signs of moderate overlap GvHD.

Conclusions: Steroid refractory GvHD represents a severe complication after allogeneic SCT for which no effective standard treatment is established and which therefore results in high mortality and morbidity. Our reported observation supports the further exploration of ruxolitinib as an effective treatment with low toxicity profile in such patients.

Disclosure: No conflict of interest disclosed.

P189 - Spectrum of hemophagocytic lymphohistiocytosis (HLH): three cases of acquired HLH with three different triggers

Jung B.1, Hartmann U.2, Jaeschke B.1, Becker S.1, Märker-Hermann E.2, Frickhofen N.1

1HSK, Dr. Horst Schmidt Klinik, Helios Kliniken Gruppe, Innere Medizin III (Onkologie, Hämatologie, Palliativmedizin), Wiesbaden, Germany, 2HSK, Dr. Horst Schmidt Klinik, Helios Kliniken Gruppe, Innere Medizin IV (Rheumatologie, Klinische Immunologie, Nephrologie), Wiesbaden, Germany

Introduction: Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening syndrome of excessive immune activation.The primary form manifests in children with genetic abnormalities of immune effectors. Secondary forms occur in the setting of conditions such as infection, malignancy, autoimmune disease or drugs.

Methods: Case reports of 3 patients with acquired HLH, treated 2013-2015. Diagnostic criteria were based on the HLH-2004 trial (Blood 118:4041;2011).


Patient 1: A 20yo female presented 10 days after tonsillectomy for tonsillitis with fever, erythema nodosum and hepatosplenomegaly. EBV-PCR suggested EBV infection as trigger of HLH.Treatment with dexamethasone 10 mg/m2 and 2 doses of etoposide 75 mg/m2 (dose reduced for impaired liver function) for 14 days overall led to complete resolution of signs and symptoms of HLH.

Patient 2: A 61yo female with diffuse large B-cell lymphoma achieved PR after chemotherapy with cyclophosphamide, doxorubicin, liposomal vincristine, prednisone and rituximab. She experienced early progression and was treated with 2 courses of dexamethasone, high dose cytarabin and cisplatin (R-DHAP). After the second course she developed signs of HLH. It improved during treatment with dexamethasone 10 mg/m2 and 2 doses of etoposide 150 mg/m2, but the lymphoma relapsed. After a course of R-DHAX (cisplatin replaced by oxaliplatin for decreased kidney function) HLH relapsed again, now with severe neurological symptoms. Despite immediate initiation of dexamethasone / cyclosporine A , HLH progressed and the patient died.

Patient 3: A 42yo female with a 15 months history of rheumatoid arthritis had been treated with methotrexate and adalimumab. 55 days after initiating adalimumab she presented with fever, aphasia, mental confusion, hepatosplenomegaly and rash. A diagnosis of HLH was made and she was started on dexamethasone/etoposide. Due to persistent fever etoposide was replaced by the IL1 receptor antagonist anakinra. Coincident with the start of anakinra and with PCR evidence of herpes and EBV viremia, HLH dramatically progressed with an increase of ferritin to 304.000 ng/ml. Anakinra was replaced by cyclosporine A and high dose immunoglobulin and aciclovir was initiated. The patient recovered completely.

Conclusions: These cases illustrate infection, lymphoma chemotherapy and adalimumab as triggers of HLH. Treatment of HLH aims to suppress the exaggerated immune response using corticosteroids, etoposide and cyclosporine A.

Disclosure: No conflict of interest disclosed.

P190 - Imitation of bone metastasis by lymphangiomatosis in a patient with colon cancer

Fund N.1, Nuber J.1, Schwella N.1

1Klinikum, Ludwigsburg, Germany

Introduction: Lymphangiomatosis is a rare type of benign disease of the lymphatic system. In principle it can occur in any lymphatic tissue, but seems to prevalently appear in the region of thorax and neck. Even skeletal involvement might be possible. The disease mainly presents in childhood, but there are also cases described in the literature with occurrence in adults.

Case report: A 38-year old male patient was admitted to the hospital because of anemia, significant weight loss and right abdominal pain. During a colonoscopy a malignant tumor of the colon ascendens was diagnosed. Histological findings revealed an adenocarcinoma. The following computer tomography of the thorax/abdomen/pelvis showed multiples osteolytic lesions in the cervical spine and in the pelvis. However, the skeletal scintigraphy remained without pathological findings. In addition a large cystic lesion of 10 × 14 cm was detected within the left upper mediastinum. The differential diagnosis of the mediastinal mass suspected a dermoid cyst or a pleural effusion. However, because of the multiple osteolytic lesions, we assumed a palliative disease related to a metastasized cancer of the colon.

Due to the stenosing malignant process, the patient underwent a right hemicolectomy (R0-resection), aiming to prevent an ileus. After postoperative recovery a biopsy was taken from a bone lesion of the pelvis, but the histology revealed no malignant tumor cells. Thereafter, another biopsy was done from the mediastinal mass. However, again no malignant involvement could be shown by pathological workup. The findings of the computer tomography and the histology without evidence of adenocarcinoma suggested the existence of a lymphangioma in the mediastinum. As the mediastinal mass resulted in a serious compression of the left lung, we decided to undertake a surgical treatment. The en-bloc surgical resection was successfully and histopathological analysis confirmed the diagnosis of a lymphangiomyoma. In conjunction with the bone lesions the final diagnosis was a generalized lymphangiomatosis and the patient was curatively treated by right hemicolectomy (G2, pT3, pN0 (0/29), pM0). Actually the patient is doing well being in regularly oncological outpatient aftercare.

Conclusion: The diagnosis of lymphangiomatosis should be kept in mind even in adult patients, especially in cases with coexistence of cystic mass and osteolytical lesions.

Disclosure: No conflict of interest disclosed.

P191 - Tandem autologous transplantation with melphalane for refractory necrobiotic xanthogranuloma

Bittenbring J.1, Pfeifer M.1, Held G.1, Pfreundschuh M.1

1UKS Homburg/Saar, Innere Medizin I, Homburg, Germany

We report on a 53 year old female treated for necrobiotic xanthogranuloma with tandem autologous stem cell transplantation.

Our patient had a IgG-kappa paraprotein, a bone marrow infiltration of 50% plasma cells and anemia of 8,0 g/dl fulfilling the criteria for multiple myeloma alone. On admission she had extensive ulcers around the orbita, on her right shoulder and on the port catheter implantation site which exposed the device on the surface. and on multiple other sites. The ulcers were infected with haemophilus influenzae and pseudomonas aeruginosa. She was diagnosed 8 years ago, treated at several institutions with typical myeloma protocols incorporating mainly dexamethason, bortezomib and lenalidomide. After failure of these treatments she had repeated plasmapheresis for 18 months which stabilized the condition but then the disease progressed again.

Background: Necrobiotic xanthogranuloma is a plasma cell dyscrasia and patients develop skin granuloma which progess to necrosis. Treatment is aimed at reducing the paraprotein level.

A stem cell transplantation was not yet attempted due to multiple infectious foci and patients wish. However with further progression and unacceptable high disease burden she decided for treatment. The port catheter was removed, a stem cell mobilization with cylcophosphamid was successful with concurrent antibiotic treatment and daily interdisciplinary wound management. High dose melphalan was started immediately thereafter. 14 days later the ulcers began to heal and newer lesions did not appear. At restaging 6 weeks later partial remission was achieved. A second high dose melphalan was performed and the ulcers healed with extensice scarring causing ectropion on both eyes with the need for reconstructive surgery at the ophthalmology departement and LMU Großhadern. Biochemically she is in very good partial remission without evidence for progression of necrobiotic xanthogranuloma 1 1/2 years after autologuous tandem transplantation.

Conclusion: Treatment of necrobiotic xanthogranuloma is aimed at reducing the causative paraprotein and wound healing with combined hematological treatment and surgical & nursing wound management.

Disclosure: No conflict of interest disclosed.

P192 - Pulmonary Marasmius aliaceus infection mimicking aspergillosis

Bittenbring J.1, Klotz M.2, Pfeifer M.1, Held G.1, Pfreundschuh M.1

1UKS Homburg/Saar, Innere Medizin I, Homburg, Germany, 2UKS Homburg/Saar, Medizinische Mikrobiologie, Homburg/Saar, Germany

We report on 32 year old female treated for acute myeloid leukemia (AML with maturation; NPM-1 and FLT3-ITD positive). She received prophylactic posaconazole starting on day 8 after 7+3 induction. In neutropenia on day 18 after induction she had persistent fever > 72 h and a thoracic CT scan showed a round pulmonary nodule in the right upper lobe with halo very suggestive of aspergillosis. Galactomannan screening which was performed twice a week was negative. However treatment for possible aspergillosis was promptly initiated with voriconazole iv. The low-grade fever and infection markers did not resolve the following 10 days. Meanwhile the white blood count resolved and a complete remission of leukemia could be demonstrated. As the aspergillosis did not resolve with normal neutrophil count we switched to liposomal amphothericin B for another 14 days. Radiographic follow up detected cavern formation. Another try with caspofungin resulted in no response for 7 days either. The patients sister was a HLA-match and further delay of consolidative treatment and transplant was not acceptable. An atypical resection of the right upper lobe was performed. As a surprise Marasmius aliaceus (garlic parachute) could be detected by PCR in the surgical specimen. This species was never reported to be a human pathogen. However this could be a reason for the missing response to common antifungals.

After quick recovery our patient had a myeloablative conditioning and siblling transplant and is now alive and well on day 250 after transplant.

Conclusion: First description of marasmius aliaceus as a human pathogen and a note for the option of surgical treatment of pulmonary fungal infections in selected clinical circumstances.

Disclosure: Jörg Bittenbring: Advisory Role: Gilead Michael Pfreundschuh: No conflict of interest disclosed.

P193 - Adulthood Wiskott-Aldrich-Syndrome

Wittke K.1, Hanitsch L.G.1, Grabowski P.1, Volk H.-D.1, Scheibenbogen C.1

1Medizinische Immunologie/ Charite Berlin, Immundefektambulanz für Erwachsene, Berlin, Germany

Introduction: Wiskott-Aldrich-Syndrome (WAS) is a rare x-linked primary immunodeficiency disease (PID) characterized by the clinical triad thrombocytopenia, ekzema and recurrent infections with an average age at diagnosis younger than two years. Due to advances in the molecular understanding of the disease, improvements in diagnostic tools, supportive care and allogenic hematopoetic cell transplantation (HCT) the morbidity and mortality of WAS could be reduced and more patients reach adulthood. Here we present the case of a 28 year old patient with WAS under supportive therapy.

Case: The patient developed petechiae and ekzema as a newborn and had several pulmonary infections during infancy. At the age of 2 years WAS was diagnosed and the patient was taken care of at a specialized pediatric center for PID. Thrombocyte counts fluctuated around 50/nl and he suffered from an Immunoglobuline (Ig)-G- and IgM-deficiency. As the disease-causing genetic alteration a splice site mutation at position +5 of intron 6 of the WAS gene was identified resulting in a truncated WAS protein. In the 1990s the patient's mother decided against an allogenic HCT. Instead a therapy with IgG was initiated. Under this therapy the patient had minor pulmonary but no life threatening infections. At the age of 24 the patient decided to stop IgG therapy and did not see his pediatrician anymore. After two years of increasing rate of infections including at least one pneumonia and worsening ekzema the patient presented to our adult center for PID. After reintroduction of subcutanous IgG therapy, consequent antiinfectious treatment, pneumocystis jiroveci prophylaxis and intensive dermatologic care the patient improved clinically and has been stable with no major infections for almost two years.

Discussion: Retrospective studies show a median survival of 14.5 years in patients with classic WAS unless a hematologic and immune reconstitution is reached by allogenic HCT which is today the treatment of choice and is ideally performed under the age of 5 years. The major causes of non HCT associated deaths are infections, bleeding and malignancies. Still there is a growing number of patients reaching adulthood even under supportive therapy only. A smooth transition between pediatric and adult medical care has to be ensured. There are several unanswered questions concerning the optimal medical care for adult WAS patients with virtually no scientific data, e.g. the role of allogenic HCT.

Disclosure: No conflict of interest disclosed.

P194 - Eculizumab therapy for gemcitabine-induced atypical hemolytic uremic syndrome in a patient suffering from adenocarcinoma of the common bile duct: a case report

Schneider C.1, Lestin M.1, Diwok C.1, Schneider-Koriath S.1, Steiner R.1, Krammer-Steiner B.1

1Klinikum Südstadt Rostock, Rostock, Germany

Introduction: Atypical hemolytic syndrome (aHUS) is a rare complication of chemotherapy with gemcitabine presenting with hemolytic anemia, thrombocytopenia, acute kidney failure and arterial hypertension. In most cases aHUS takes a fatal clinical course. We describe the successful treatment of a gemcitabine-induced aHUS with eculizumab, a humanized monoclonal antibody that prevents the production of the terminal complement complex.

Methods: A 44 year old male patient suffering from adenocarcinoma of the common bile duct was treated with postsurgical adjuvant chemotherapy with gemcitabine (1000 mg/m² day 1,8,15, new cycle day 29). After the fifth cycle he developed arterial hypertension and a Coombs negative hemolytic anemia. In addition, thrombocytopenia and renal failure with proteinuria occurred. Diagnostic evaluation excluded a gastrointestinal infection and revealed schistocytes in the peripheral blood smear and a normal plasma ADAMTS13 activity. Thus, a gemcitabine-induced aHUS was suggested.

Coincidental to the aHUS and complicating the clinical course, the patient developed an adhesive ileus, which had to be surgically revised. Immediately after successful surgery the therapy with corticoids (1g/kg body weight) and plasma exchange was initiated leading to the patient`s clinical stabilisation. Two days later the therapy could be changed to eculizumab (900mg/m² weekly for the first four weeks, thereafter 1200mg/m² every two weeks). Vaccination against Neisseria meningitidis was administered as recommended and accompanying antibiotic prophylaxis was performed for a period of 14 days.

Results: Within one week after the first application of eculizumab, the platelet count as the most sensitive parameter of therapy response normalized. Renal function recovered to near normal and proteinuria resolved. Hemodialysis could be prevented. Eventually, under continued eculizumab therapy over the following weeks all blood counts normalized and the patient is now in excellent clinical condition. Arterial hypertension remained and is currently under medical treatment. Despite abandoning the therapy with gemcitabine the patient is still in complete remission.

Conclusion: Atypical HUS as a life threatening complication of chemotherapy with gemcitabine was successfully treated with eculizumab. The early adoption of therapy allowed to restore renal function and the prevention of hemodialysis.

Disclosure: No conflict of interest disclosed.

P195 - Splenomegaly and cytopenia after surgical resection and chemotherapy for colorectal cancer: An interdisciplinary case of hematology and oncology

Haas M.1, Rexrodt P.2, Schlitt H.J.3, Vehling-Kaiser U.1

1Praxis für Hämatologie, Onkologie und Palliativmedizin, Landshut, Germany, 2Praxis für Radiologie und Strahlentherapie, Landshut, Germany, 3Universitätsklinikum Regensburg, Klinik und Poliklinik für Chirurgie, Regensburg, Germany

Introduction: The current report from an oncologic practice in Germany describes a cross-link between hematology, oncology and surgery

Methods: In 2001, a woman then aged 38 years underwent R0 resection for adenocarcinoma of the ascending colon. Afterwards an adjuvant chemotherapy with 5-fluorouracil/folinic acid/oxaliplatin was administered. 4 years later the patient sufferd from relapse with metasis in the right liver lobe and right hemihepatectomy was done. One year later splenomegaly of 18cm x 5cm became evident, but did not cause any symptoms. The blood count then showed a slight tri-cytopenia possibly due to previous chemotherapy. The patient was fully active without any problems due to splenomegaly. In 2013, the patient increasingly suffered from left-sided abdominal pain and fatigue, the spleen then was 20cm in size. Thrombocyte count had dropped below 50 G/L. Bone-marrow puncture did not show signs of dysplastic syndrome or other malignancy. In 2014, the spleen was 23 cm in longest diameter. A CT-scan showed signs of portal hypertension with pronounced collaterals, possibly due to constriction of the inferior vena cava and/or left hepatic vein, possibly due to the liver-resection in 2005. A gastroscopy confirmed oesophageal varices. The patient was transferred to a university hospital for further diagnostics and potential placement of a portosystemic shunt. Left hepatic vein stenosis could not be substantiated by venous catheter pressure measurements. As a definite reason for portal hypertension was not clear with no evidence for liver cirrhosis or fibrosis, the patient was advised for splenectomy for symptom relief.

Results: To avoid splenectomy, as preferred by the patient and the treating hemato-oncologist, the case was again discussed with the surgeon. It was concluded to initially perform a side-to-side spleno-renal shunt. After constructing the shunt, a significant reduction of blood-flow through the portal vein directly after surgery was observed. Only three months after the intervention, the spleen had shrunk from 24cm to 16cm accompanied by an increase in thrombocytes above 100 G/L and significant improvement of symptoms.

Conclusions: The patient achieved a significant improvement of symptoms after construction of the shunt. Close networking between different disciplines and especially the treating physicians in rural areas and specialists at highly specialized centers were crucial to avoid splenectomy in this case.

Disclosure: No conflict of interest disclosed.

P196 - Chemotherapy and pregnancy: 2 case reports from clinical practice

Lestin M.1, Krammer-Steiner B.1

1Klinikum Südstadt Rostock, Rostock, Germany

Prospectively, the number of pregnant women suffering from cancer will rise due to delayed childbearing and the observed increase in non-Hogdkins lymphoma in the last decades. Oncologists and gynecologists have to face the challenging issue to obtain a good long-term outcome for mother and fetus.

Case 1: A 22-year old pregnant woman (25th week of gestation) presented with an inguinal swelling, hepatosplenomegaly, bone lesions, pancytopenia and elevated LDH. Lymph node biopsy revealed diffuse large B-cell lymphoma. We administered 4 courses of standard dose R-CHOP-14. After 3 courses she underwent surgergy because of pathological femur fracture. In 37+2 week of gestation an eutrophic healthy girl was born via normal vaginal delivery. The mother completed therapy with 4 courses R-CHOEP-14 and 4 single doses Rituximab.

Case 2: A 33-year old woman presented with breast cancer in 21+0 week of gestation. There was no evidence of metastasis. After breast conservative surgery she received 4 courses adjuvant chemotherapy with epirubicine and cyclophosphamide. In 37+1 week of gestation our patient gave birth to a healthy eutrophic boy via normal vaginal delivery. Her therapy has been completed by 12 courses paclitaxel and radiation.

As reported from other authors chemotherapy administered during 2nd and 3rd trimesters of pregnancy seems to be relatively safe. The teratogenicity of chemotherapeutic agents depends on the timing of exposure. Nevertheless chemotherapy during 2nd and 3rd trimesters increases the risk of intrauterine growth restriction.

CHOP regimen is considered safe in the 2nd and 3rd trimesters. Rituximab is essential in treatment of aggressive lymphoma. Notably, Rituximab can cross the placenta. As reported in literature neonates exposed to Rituximab presented haematological abnormalities (B-cell depletion, cytopenia) but none had corresponding infections.

In pregnant women with breast cancer the decision to recommend mastectomy or breast conservative surgery and also the indication for systemic therapy should follow standard practice. Anthracycline-based regimens remain first choice. Due to risk of fetal complications radiation, trastuzumab and endocrine therapy sholud be initiated after delivery.

Disclosure: No conflict of interest disclosed.

P197 - Simultaneous manifestation of Hodgkin lymphoma and follicular Non-Hodgkin lymphoma

Nuber J.1, Fund N.1, Schwella N.1

1Klinikum, Ludwigsburg, Germany

Introduction: The two main groups of malignant lymphoma are Hodgkin lymphoma (HL) and Non-Hodgkin lymphoma (NHL). In general they are considered as two different entities of malignant lymphoma, but reported cases showed the occurrence of both in the same patient. Most of the studies and case reports describe the development of an NHL after the successful treatment of initially diagnosed HL. This may be related to late treatment-toxicity of chemotherapy and/or radiotherapy.

A rare number of cases have shown the possibility of simultaneous manifestation of HL and NHL in the same patient. Especially, the rare occurrence of two histological different lymphomas in the same organ is defined as a composite lymphoma.

As the pathogenesis of the occurrence of two histological distinct malignant lymphoma variants in the same patient is not clarified, a possible genetical relation is object of discussion.

Case report: A 73-year old male patient was diagnosed with weight loss, night sweat and enlarged axillary and inguinal lymph nodes. The biopsy was performed on one of the two palpable inguinal lymph nodes on the right side. Histological findings showed a classical non-EBV associated HL. Computer tomography showed a splenomegaly without further abdominal and thoracic lymphoma. Furthermore, bone marrow examination showed a 20% infiltration of HL (stage IV B/S).

The patient was initially treated with doxorubicin, bleomycin, vinblastine and dacarbazine (ABVD). A follow up computer tomography after 4 courses of ABVD showed in general a partial response. The clinical examination however, surprisingly showed one of the inguinal lymph nodes of constant size, where no biopsy was taken before. Therefore, another biopsy was performed on the non-regressive inguinal lymph node, also located on the right side. The biopsy revealed again Hodgkin and Sternberg-Reed cells, as well as follicular lymphoma cells. After this interesting results we analyzed again the initial biopsy particularly with regard to follicular lymphoma cells, but gained no additional information.

This case shows the possible coexistence of a HL and a follicular NHL grade 1/2 already occurring from the beginning on.

Conclusion: Although the simultaneous manifestation of HL and NHL in the same patient is rare, this possibility and a second biopsy should be considered, especially of non- responding areas following treatment against HL.

Disclosure: No conflict of interest disclosed.

P198 - Severe skin complication after implantation of a subcutaneous port system in a patient with leukaemia AML

Heißner K.1, Kopp H.-G.1, Ghoreschi K.2, Metzler G.2, Horger M.3, Kanz L.1, Baur R.1, Vogel W.1

1Universitätsklinikum Tübingen, Medizinische Klinik 2 Onkologie, Hämatologie, Klinische Immunologie, Rheumatologie und Pulmologie, Tübingen, Germany, 2Universitätsklinikum Tübingen, Universitäts-Hautklinik, Tübingen, Germany, 3Universitätsklinikum Tübingen, Abteilung für Diagnostische und Interventionelle Radiologie, Tübingen, Germany

Case report: A 76-year-old man with acute myeloid leukemia that had evolved from previously diagnosed myelodyspblastic syndrome (MDS) was regularly admitted to our outpatient unit for treatment with 5-Azacytidin s.c. The patient required packed red blood cells on a regular basis and remained chronically neutropenic. Because the patient had lost his right upper arm in early childhood in an accident, repeated left-sided venous access became increasingly difficult. Therefore an implantable venous access device (port) was implanted.

The patient was admitted to our emergency department the day after surgery with local erythema, swelling and overheated skin surrounding the scar. The port system was promptly removed and systemic antibiotic treatment was applied. Microbiological smears identified sensitive natural skin flora. Despite excessive use of different antibiotics, reddish-livid skin destruction enlarged with multiple painful ulcerations. Furthermore, distant ulcerations localized around the left elbow emerged. Necrotizing subcutaneous tissue involvement was ruled out by MRI. Deep skin biopsies interestingly showed accumulation of neutrophils despite of persisting peripheral blood °IV neutropenia, consistent with a diagnosis of pyoderma gangraenosum (PG). Treatment with corticosteroids and immune globulins resulted in rapid and ongoing wound healing, confirming the diagnosis.

Conclusion: This is a case of PG after implantation of a subcutaneous port devide, resulting in progressive local tissue ulceration not primarily caused by bacterial infection. PG typically occurs in patients with chronic disease, including myelodyplasia. Our case highlights the importance of considering non-infectious causes of severe cutaneous complications after surgical procedures in haematologic patients. PG is a diagnosis of exclusion, skin biopsies are strongly recommended to confirm the diagnosis.

Disclosure: No conflict of interest disclosed.

P199 - Long-lasting remission of metastatic breast cancer with Eribulin

Dobbie M.1

1Unité d‘Oncologie, Hôpital du Jura, Porrentruy, Switzerland

Introduction: Eribulin has been approved to treat patients with metastatic breast cancer who have received at least two prior chemotherapy regimens for late-stage disease, including both anthracycline- and taxane-based chemotherapies.

Methods: Case report of a 48 year old patient with metastatic breast cancer treated with Eribulin.

Results: This at diagnosis premenopausal patient was diagnosed with ER pos, Her-2 negative, metastatic breast cancer (ulcerated breast lesion with multiple satelite skin metastasis, axillary lymph node metastasis and a malignant pleural effusion) in 2005. Treatment with 6 cycles of palliative chemotherapy with Docetaxel and Epirubicin) from 9/05-1/06 (partial remission). Antihormonal treatment followed with Letrozole from 2/06-1/07 (stopped because of local progression), Treatment was continued with Tamoxifen with ovarian suppression (recovery of ovarian function) from 1/07-4/12.

In 9/2010 pulmonary embolism and heart failure occurred and standard treatment was given with clinical improvement.

A right sided tuboovarian abscess (12/10) was treated with antibiotics (patient inoperable) Gynecological follow up showed a growing tumor in the left adnexis and in 2/12 after cardiac recompensation, a staging laparoscopy was performed with left adnexectomy and multiple biopsies of peritoneal implants. The pathological diagnosis was serous adenocarcinoma of the ovary.

From 1/12 to 4/12 3 cycles of Carboplatin AUC 5 were given, restaging with CT showed progressive disease. Mechanical ileus developed and right sided adnexectomy with resection of 40 cm of ileum and an ileo-ascendostomy was performed. The second pathological report led to the diagnosis of metastasis of the invasive breast cancer with a typical immunohistochemical profile.

After the operation, base line PET/CT showed FDG positive lesions in both breasts, the right axilla, the liver and the right sided peritoneum.

Palliative Chemotherapy with Eribulin 1,4 mg/m2 i.v. day 1 and 8, every 3 weeks, was started in 10/12 and the patient is still on treatment after 35 cycles (at time point 5/15). Treatment was well tolerated (no hematological toxicity > grade 1) and the general condition has improved. Follow up radiological examinations show a continous partial remission (in 2/15 no signs of metastasis in lung or liver).

Conclusions: The patient tolerated Eribulin very well despite the important comorbidities with an excellent effect on progression free survival.

Disclosure: Michael Dobbie: Employment or Leadership Position: Leitender Arzt Onkologie, Hôpital du Jura; Financing of Scientific Research: Eisei.



P200 - High intratumoral FOXP3+ T regulatory cells (Tregs) density in colorectal cancer is associated with improved survival but unrelated to tumor localization

Marx A.H.1, Sauter G.1, Simon R.1, Bokemeyer C.2, Terracciano L.3, Izbicki J.R.4, Melling N.4

1University Medical Center Hamburg-Eppendorf, Hamburg, Pathology, Hamburg, Germany, 2Hubertus Wald Cancer Center, University Medical Center Hamburg-Eppendorf, Department of Oncology, Hematology, BMT with section Pneumology, Hamburg, Germany, 3University Hospital Basel, Pathology, Basel, Switzerland, 4University Medical Center Hamburg-Eppendorf, Hamburg, Surgery, Hamburg, Germany

Introduction: To determine the density and prognostic significance of FOXP3+ T regulatory cells (Tregs) in colorectal cancer of different localizations (right versus left colon) compared with conventional histopathologic features and with CD3+ and CD8+ lymphocyte densities.

Methods: Tissue microarrays (TMA) and immunohistochemistry (IHC) were used to examine the densities of CD3+, CD8+ and FOXP3+ lymphocytes in normal colonic mucosa and tumor tissue from 1800 colorectal cancers. These densities were evaluated for associations with histopathologic features and patient survival.

Results: FOXP3+ Tregs density was low in 547 (50.5%), high in 120 cases (11.1%). FOXP3+ Tregs density was associated with a high CD8+ (p = 0.018) and CD3+ density (p < 0.0001) in tumor tissue. FOXP3+ Tregs density was higher in tumor tissue compared to normal tissue (p < 0.0001). Low FOXP3+ Tregs density was significantly associated with high tumor grade (p < 0.0001), high tumor stage (p < 0.0001) and nodal status (p = 0.023), but there was no correlation to histological subtypes (mucinous, signet cell; p = 0.42) or tumor localization (right versus left; p = 0.17). High density of FOXP3+ Tregs, CD3+ and CD8+ cells in tumor tissue was related to a better patient survival (p < 0.0001, p = 0.02 and p = 0.01 respectively; Figure 3). In a multivariate analysis including tumor stage, tumor grade, tumor location, nodal status and FOXP3+ Treg content only pT (p < 0.0001) and pN (p < 0.0001) had significance, but not tumor grade (p = 0.42), tumor localization (p = 0.1) or FOXP3+ Treg infiltration (p = 0.07).

Conclusion: FOXP3+ Treg cell density in CRC may help as a predictive biomarker for the risk assessment and therapy management of early and middle stage colorectal cancer.

Disclosure: No conflict of interest disclosed.

P201 - Molecular subtypes and outcomes in regorafenib-treated patients with metastatic colorectal cancer (mCRC) enrolled in the CORRECT trial

Karthaus M.1, Schwenke S.2, Seidel H.3, Beckmann G.2, Reischl J.2, Vonk R.2, Lenz H.-J.4, Tebernero J.5, Siena S.6, Grothey A.7, van Cutsem E.8, Jeffers M.9, Wagner A.2, Laurent D.2, Kobina S.9, Rutstein M.9, Guinney J.10, Tejpar S.11

1Klinikum Harlaching, Klinik für Hämatologie, Onkologie und Palliativmedizin, München, Germany, 2Bayer Pharma AG, Berlin, Germany, 3Bayer HealthCare Pharmaceuticals, Berlin, Germany, 4University of Southern California/Norris Comprehensive Cancer Center, Los Angeles, United States, 5Vall d'Hebron University Hospital, Barcelona, Spain, 6Ospedale Niguarda Ca' Granda, Milano, Italy, 7Mayo Clinic, Rochester, United States, 8University Hospitals Gasthuisberg, Leuven, Belgium, 9Bayer HealthCare Pharmaceuticals, Whippany, United States, 10Sage Bionetworks, Seattle, United States, 11Molecular Digestive Oncology, Leuven, Belgium

Introduction: In the CORRECT Ph3 trial (NCT01103323), regorafenib improved overall survival (OS) and progression-free survival (PFS) vs placebo in patients with mCRC who progressed on standard therapies (Grothey 2013). Initial biomarker analyses suggested that regorafenib provides a clinical benefit in various mutational subgroups (Jeffers 2013). Here we present additional exploratory analyses to evaluate clinical benefit in CRC subgroups defined by gene expression.

Methods: Gene expression analysis (Affymetrix ST1.0 array) was conducted on archival tumor tissue from 281 of the 760 patients (37%) enrolled in CORRECT. Next-generation sequencing (NGS) was done on 239 specimens (FoundationONE). Gene expression data were subjected to hierarchical molecular tumor classification (Marisa 2013). Cox proportional hazards models were used to identify potential prognostic or predictive biomarkers.

Results: The distribution of the 6 different CRC subtypes characterized by gene expression clusters originally defined by Marisa and the classification of the 281 patients from CORRECT are shown in the table.


The six Marisa subtypes derived a similar OS benefit from regorafenib. Although the numbers of patients in the subgroups are small, a greater PFS benefit for regorafenib was seen in patients in the ‘high-risk' subgroup (C4 + C6, n = 26; HR = 0.10; 95% CI 0.02-0.35; p = 0.0009) than the ‘low-risk' subgroup (C1+C2+C3+C5, n = 255; HR = 0.58; 95%CI 0.44-0.77; p = 0.002). NGS analyses suggested that the chromosomal instability group (CIN) was the predominant subgroup in CORRECT.

Conclusion: Molecular characterization by gene expression analysis may allow the identification of CRC subgroups that correlate with regorafenib clinical benefit. Data suggest this does not depend on the presence of mutations. Additional exploratory analyses, including the use of the recently defined Consensus molecular subtypes (Dienstmann ASCO 2014), to validate these results are ongoing.

Disclosure: Meinolf Karthaus: Expert Testimony: Bayer Sabine Tejpar: No conflict of interest disclosed.

P202 - Molecular targeting of EGFR-downstream effectors in colorectal cancer cells by antibody-mediated siRNA treatment

Appel N.1, Bäumer N.1, Wardelmann E.2, Buchholz F.3, Müller-Tidow C.4, Berdel W.E.1, Bäumer S.1

1Universitätsklinikum Münster, Medizinische Klinik A, Labor für Molekulare Hämatologie und Onkologie, Münster, Germany, 2Universitätsklinikum Münster, Gerhard-Domagk Institut für Pathologie, Münster, Germany, 3TU Dresden, UCC, Medical Systems Biology, Medizinische Fakultät, Dresden, Germany, 4Universitätsklinikum Halle, Innere Medizin IV, Hämatologie und Onkologie, Halle, Germany

Introduction: RAS-mutated carcinomas are resistant against EGFR-antibody therapies such as Cetuximab. We overcame this therapy resistance in a proof-of-principle study ADDIN RW.CITE{{168 Bäumer,S. 2015}}[1]. In this study, we coupled the anti-EGFR-antibody Cetuximab chemically to protamine via the linker sulfo-SMCC. This complex carries siRNA into EGFR-positive cells such as the colorectal cancer cells lines HCT116, LoVo, DLD-1, SW480 and HT29. The siRNA-mediated downregulation of KRAS leads to silencing of MAPK pathway signal transduction, decreased proliferation and enhanced apopotosis. Since tumor growth in vivo was inhibited but still detectable, we combined KRAS downregulation with siRNA treatments against additional EGFR-downstream factors that might be operative in MAPK independent tumor cell growth.

Methods: We used the Cetuximab-sulfo-SMCC-protamine complex to bind siRNAs e.g. against PI3CA, which is mutated or overexpressed in many tumors. We treated colorectal cancer cell lines with KRAS-siRNA-, PI3CA-siRNA- or combined Cetuximab-complexes. Cells were then subjected to colony formation assays in soft agar, MTS assays to determine vitality and proliferation. Also, we injected cells subcutaneously into the flank of CD1-nude mice and treated these mice with the Cetuximab-siRNA complexes.

Results: Downregulation of PI3CA in colorectal cancer cells leads to significantly decreased colony formation. Moreover, combination of siRNA against KRAS and PI3CA coupled to Cetuximab-complexes dramatically slowed down tumor growth in vivo.

Conclusions: Antibody-mediated knockdown of KRAS in KRAS-mutated tumors restores EGFR-sensitivity. Downregulation of additional signaling pathways leads to a cooperative treatment success. This indicates that complete molecular characterization of multiple signaling pathways driving malignant behaviour of the respective cell type enables us to tailor the siRNA delivery system to the specifically expressed oncogenes and consequently in a given cell line or tumor yields optimal therapeutic activity.


1 Bäumer S, Bäumer N, Appel N, et al.: Antibody-mediated delivery of anti-KRAS-siRNA in vivo overcomes therapy resistance in colon cancer. Clin Cancer Res 2015;21:1383-1389.

Disclosure: No conflict of interest disclosed.

P203 - Colorectal carcinoma patients with wild-type PIK3CA tumors show improved overall survival

Gebauer L.1, Stabla K.1, Nist A.2, Mernberger M.2, Wischmann V.3, Stiewe T.2, Barckhausen C.1, Moll R.3, Brendel C.1, Neubauer A.1

1Philipps University of Marburg and University Hospital of Giessen and Marburg, Department of Hematology, Oncology and Immunology, Marburg, Germany, 2Philipps University of Marburg, Molecular Oncology, Marburg, Germany, 3Philipps University of Marburg, Institute of Pathology, Germany

Introduction: Specific gene mutations can influence the response of tumors to anticancer drugs. Thus, molecular methods gain in importance for clinical diagnostics to create a decisional base for the administration of a given drug. In the past, there have been conflicting results with regard to aspirin use and survival in correlation with PIK3CA mutations in colorectal cancer patients. This retrospective study was done to understand a potential relation between PIK3CA mutation status and aspirin use in colorectal cancer.

Patients and methods: This was a retrospective analysis of paraffin-embedded tumor tissue of 153 patients suffering from colorectal carcinoma stage I-III (UICC classification) or IV with only a few resectable liver metastases. Genomic DNA was subjected to analysis of PIK3CA and KRAS mutation status by pyrosequencing or semiconductor sequencing, respectively. Mutation status and regular aspirin use were correlated to 5-year and 10-year overall survival (OS).

Results: With regard to the entire patient cohort, aspirin use per se did not affect 10-year-OS significantly. However, wild-type PIK3CA status was associated with a significantly higher rate of 10-year OS (48% vs. 21%, p = 0.01, Fisher`s exact test). Detailed subgroup analyses revealed that especially aspirin users with wild-type PIK3CA tumors show significantly improved 5-year OS (75% vs. 25%, p = 0.01, Fisher's exact test) and 10-year OS (57% vs. 13%, p = 0.05, Fisher's exact test). KRAS mutation did not impact OS in our CRC patient cohort.

Conclusions: Our data indicate that post-diagnosis aspirin intake could be beneficial especially for aspirin users with PIK3CA mutation while KRAS status is irrelevant for clinical diagnostics with regard to a decision for or against aspirin prescription.

Disclosure: No conflict of interest disclosed.

P204 - Comparison of therapeutic efficacy of single VEGF- versus combined VEGF/Angiopoietin-2 inhibition in combination with chemotherapy in a colorectal carcinoma xenograft model

Freystein J.1, Reipsch F.1, Caysa H.1, Schmoll H.-J.2, Müller-Tidow C.1, Müller T.1

1Martin-Luther-Universität Halle-Wittenberg, Universitätsklinik für Innere Medizin IV, Hämatologie/Onkologie, Halle (Saale), Germany, 2Martin-Luther-Universität Halle-Wittenberg, AG Klinische Studien in der Onkologie, Halle (Saale), Germany

VEGF inhibition by addition of Bevacizumab to chemotherapy regimen of metastatic colorectal cancer leads to improved outcome. However, anti-angiogenic tumor therapy targeting a single factor may be limited by complementary mechanisms. It was shown that high Angiopoietin-2 (Ang-2) serum levels are associated with poor clinical outcome of colorectal cancer patients treated with Bevacizumab-containing therapy. The aim of the present study was to compare therapeutic efficacy of single VEGF- vs. combined VEGF/Ang-2 inhibition using a novel bi-specific antibody (CrossMab) in combination with chemotherapy in a colorectal cancer xenograft model.

Subcutaneous xenograft tumors were generated in athymic nude mice using the human colorectal cancer cell line DLD-1. Monitoring of tumor growth was performed by caliper measurement. Mean tumor volume per group (n = 10) at start of treatment was around 130 mm3. Chemotherapy consisted of a combination of 5-fluorouracil and irinotecan. Following antibodies provided by Roche were used: anti-VEGF (B20.4.1), CrossMab (B20.4.1/LC06). Single and combination treatments were performed by i.p. injections once weekly.

Mean tumor volume of control group reached 1400 mm3 on d18 after start of treatment with 4 of 10 tumors exceeded 1500 mm3 and treatment was stopped. Chemotherapy and anti-VEGF treatment inhibited tumor growth to some extent and had to be completed on d27 when 4 tumors within each group exceeded 1500 mm3. An efficient tumor growth inhibition could be achieved by single treatment with the CrossMab and both combination regimen. The study was completed on d49 when 2 tumors within the CrossMab- and anti-VEGF/chemotherapy groups had exceeded 1500 mm3. Mean tumor volumes at this time point were 1018 mm3, 829 mm3 and 296 mm3 for anti-VEGF/chemotherapy, CrossMab and CrossMab/chemotherapy, respectively. Analysis of single tumors revealed that 3 of 10 tumors in the anti-VEGF/chemotherapy group, 5 of 10 tumors in the CrossMab group and 10 of 10 tumors in the CrossMab/chemotherapy group had shown complete growth retardations. Statistical analysis on d18 showed that CrossMab alone and both combination treatments led to a significant inhibition of tumor growth compared to PBS control. The difference between CrossMab/chemotherapy and anti-VEGF/chemotherapy on d49 was highly significant.

In conclusion, the therapy regimen comprising combined VEGF/Ang-2 inhibition was clearly superior to combination therapy with single VEGF inhibition.

Disclosure: Juana Freystein: Expert Testimony: Die Studie wurde durch Drittmittel der Firma Roche unterstützt. Thomas Müller: Expert Testimony: Die Studie wurde durch Drittmittel der Firma Roche unterstützt.

P205 - Heterogeneity of colorectal cancer liver metastasis - analysis of WNT pathway patterns via High-throughput and bioinformatics profiling

Bleckmann A.1,2, Wachter A.2, Conradi L.-C.3, Wolff A.2, Hoppenau C.4, Korf U.5, Schildhaus H.-U.4, Homayounfar K.3, Pukrop T.1, Beissbarth T.2

1Dept. of Hematology and Medical Oncology, Göttingen, Germany, 2Dept. of Medical Statistics, Göttingen, Germany, 3Dept. of General, Visceral and Pediatric Surgery, Göttingen, Germany, 4Dept. of Pathology, Göttingen, Germany, 5German Cancer Research Center (DKFZ), Heidelberg, Germany

Introduction: In cancer progression the development of distant metastases is the crucial adverse event. Focusing on colorectal cancer, the majority (70%) of patients develops metastasis in the liver. Within the BMBF-founded consortium MetastaSys the role of the mutational activation in the WNT pathway are investigated in these metastases. In order to address the phenomenon of intermetastatic heterogeneity the established metastasis profiling was applied to several metastases of the same patient.

Methods: Two pilot cases including four and two liver metastases, respectively as well as corresponding normal liver tissue were comprehensively characterized in terms of clinical annotations as well as standardized histopathological and molecular subtyping. All tumor samples were subtyped according to the specific mutational status of KRAS, NRAS, BRAF and PIK3CA as well as alterations in DNA mismatch repair enzymes and microsatellite instability. Fresh frozen samples for transcriptome analyses were characterized by an experienced pathologist (tumor cell content, amount of stroma, inflammatory infiltrate and necrosis) to assure high quality of tissue and to obtain valid data. After characterization, samples were profiled via subsequent transcriptome sequencing and proteomics profiling via Reverse-Phase-Protein-Arrays (RPPA).

Results: To access the clinical and genetical characterization of the samples a standardized data analysis pipeline for processing the RNA-Seq data was established. The principle-component-analysis revealed a good separation between normal tissue and tumor samples. Interestingly, the correlation between healthy samples among different patients was higher than between the metastatic samples of the same patient. Furthermore, relevant WNT markers such as AXIN2, Lef1 and CDX2 were within the top 25 upregulated genes discriminating between normal and metastatic samples.

Conclusion: RNA-Seq analysis allows a clear discrimination of normal and metastatic liver tissue. In different metastases of the same patient heterogeneous expression patterns in the WNT pathway are identified with relevant implication for clinical treatment approaches. Based on these findings and the positive validation of the established methodology including innovative High-throughput profiling and comprehensive bioinformatics approaches we started analyzing a large number of metastatic patient samples within the consortium.

Disclosure: No conflict of interest disclosed.

P206 - Loss of H2Bub1 expression is linked to poor prognosis in nodal negative colorectal cancers

Marx A.H.1, Simon R.1, Bokemeyer C.2, Sauter G.1, Terracciano L.3, Izbicki J.R.4, Melling N.4

1University Medical Center Hamburg-Eppendorf, Pathology, Hamburg, Germany, 2Hubertus Wald Cancer Center, University Medical Center Hamburg-Eppendorf, Department of Oncology, Hematology, BMT with section Pneumology, Hamburg, Germany, 3University Hospital Basel, Pathology, Basel, Switzerland, 4University Medical Center Hamburg-Eppendorf, Surgery, Hamburg, Germany

Introduction: To correlate H2Bub1 expression with outcome in colorectal cancer.

Methods: H2Bub1 expression was analyzed by immunohistochemistry on a tissue microarray containing 1800 colorectal cancers. Results were compared to clinicopathological parameters.

Results: H2Bub1 IHC was seen in 1256 (79.3%) of 1584 interpretable CRC and was considered weak in 26.2% and strong in 53.1% of cancers. H2Bub1 expression was completely lost in 20.7% of the cases. Loss of H2Bub1 expression was associated with high tumor grade (p = 0.0211), high tumor stage (p = 0.0003), positive nodal status (p = 0.0139) and histological tumor type (p = 0.0202). No link was found between H2Bub1 expression and tumor localization (p = 0.1262), peritumoral lymphocytic infiltration (p = 0.2523) or vascular invasion (p = 0.5970).

Loss of H2Bub1 expression in CRC was strongly associated with poor patient survival (p = 0.0006). This observation held true also in a subset survival analysis of nodal negative (N0) and nodal positive (N1) cancers (p = 0.0296 and p = 0.0197, respectively). In the subgroup of p53 negative cancers no prognostic impact of H2Bub1 staining was seen (p = 0.1924), whereas in p53 positive CRC H2Bub1 expression loss was associated with poor prognosis (p = 0.0031). Strikingly worsened outcome was found for nodal negative cancers presenting with accumulation of p53 when H2Bub1 expression was lost (p = 0.0006).

Conclusion: Our data demonstrate that a reduced H2Bub1 expression is a strong prognostic biomarker both in nodal negative and nodal positive CRC. H2Bub1 expression measurement might help to select nodal negative CRC patients that may benefit from adjuvant therapy.

Disclosure: No conflict of interest disclosed.

P207 - High Ki67 expression in colorectal cancer is an independent prognostic marker associated with good prognosis and left-sided tumor localization

Marx A.H.1, Simon R.1, Bokemeyer C.2, Terracciano L.3, Simon G.1, Izbicki J.R.4, Melling N.4

1University Medical Center Hamburg-Eppendorf, Pathology, Hamburg, Germany, 2Hubertus Wald Cancer Center, University Medical Center Hamburg-Eppendorf, Department of Oncology, Hematology, BMT with section Pneumology, Hamburg, Germany, 3University Hospital Basel, Pathology, Basel, Switzerland, 4University Medical Center Hamburg-Eppendorf, Surgery, Hamburg, Germany

Introduction: To correlate Ki67 expression with outcome and other molecular parameter in colorectal cancer.

Methods: Ki67 labeling index (Ki67LI) was analyzed by immunohistochemistry on a tissue microarray containing 1800 colorectal cancers. Results were compared to clinicopathological and molecular parameters.

Results: Ki67LI was considered low in 26.3%, moderate in 56.7% and high in 17.0% of 1653 interpretable CRCs. High Ki67 expression was associated with low tumor stage (p < 0.0001) and left-sided tumor localization (p = 0.0280), but not with tumor grade (p = 0.8948), nodal status (p = 0.0702) or histological tumor type (p = 0.7274) and was an independent prognosticator of favorable survival (p = 0.0004). High Ki67 expression was also significantly linked to high-level nuclear β-catenin and p53 expression (p < 0.0001 and p = 0.0288, respectively).

Conclusion: Our data show that high Ki67 expression in colorectal cancers is associated with good clinical outcome and has a preference for left-sided tumors. Ki67, p53 and β-catenin overexpression seem to be linked in CRC and indicate a cellular state of high proliferative activity. Finally, our findings strongly argue for a clinical utility of Ki67 immunostaining as an independent prognostic biomarker in colorectal cancer enabling to select patients for adjuvant treatment and contributing to the prognostic evaluation in these patients.

Disclosure: No conflict of interest disclosed.

P208 - Expression of phospho-mTOR kinase is abundant in colorectal cancer and associated with left-sided tumor localization - a potential therapy target?

Marx A.H.1, Simon R.1, Izbicki J.R.2, Bokemeyer C.3, Terracciano L.4, Sauter G.1, Melling N.2

1University Medical Center Hamburg-Eppendorf, Pathology, Hamburg, Germany, 2University Medical Center Hamburg-Eppendorf, Surgery, Hamburg, Germany, 3Hubertus Wald Cancer Center, University Medical Center Hamburg-Eppendorf, Department of Oncology, Hematology, BMT with section Pneumology, Hamburg, Germany, 4University Hospital Basel, Pathology, Basel, Switzerland

Introduction: The mammalian target of rapamycin (mTOR) has been suggested as a prognostic biomarker and therapeutic target in an array of human cancers.

Methods: phospho-mTOR (p-mTOR) expression was analyzed by immunohistochemistry (IHC) on a tissue microarray containing 1800 colorectal cancers (CRC). Clinical follow-up data were available from all cancer patients.

Results: Positive p-mTOR immunostaining was seen in 83.5% of 1640 interpretable CRC and was considered weak in 862 (52.5%) and strong in 508 cases (31.0%). Matching clinico-pathological parameters were available in 1580 cases. p-mTOR staining was more frequent in tubular adenocarcinomas than in the less common histological subtypes (mucinous, medullary, signet cell; p = 0.0163) and significantly linked to carcinomas of the left-sided colon and rectum as compared to right-sided CRC (p = 0.0066). There was no significant association between p-mTOR expression and patients' gender, tumor stage, tumor grade or nodal status. In a survival analysis, p-mTOR IHC status of all CRC was unrelated to patient survival (p = 0.702). In a multivariate analysis including pT, pN, tumor grade, tumor localization and p-mTOR expression, only pT, pN (both p < 0.0001) and grade (p = 0.0001) showed prognostic impact, but not tumor localization (p = 0.9472) or p-mTOR expression (p = 0.8879).

Conclusion: Our observations indicate that p-mTOR overexpression is abundant in CRC, and linked to left-sided tumor localization. The high frequency and overexpression of p-mTOR is providing further rationale for targeting this pathway therapeutically in CRC patients. However, a prognostic role of p-mTOR overexpression in CRC could not be confirmed.

Disclosure: No conflict of interest disclosed.

P209 - RBM3 expression loss is associated with right-sided localization and poor prognosis in colorectal cancer

Marx A.H.1, Simon R.1, Izbicki J.R.2, Terracciano L.3, Bokemeyer C.4, Sauter G.1, Melling N.2

1University Medical Center Hamburg-Eppendorf, Pathology, Hamburg, Germany, 2University Medical Center Hamburg-Eppendorf, Surgery, Hamburg, Germany, 3University Hospital Basel, Pathology, Basel, Switzerland, 4Hubertus Wald Cancer Center, University Medical Center Hamburg-Eppendorf, Department of Oncology, Hematology, BMT with section Pneumology, Hamburg, Germany

Introduction: The RNA-binding motif protein 3 (RBM3) has recently been suggested as a prognostic biomarker in an array of human cancers.

Methods: RBM3 expression was analyzed by immunohistochemistry on a tissue microarray containing 1800 colorectal cancers (CRC).

Results: Nuclear RBM3 immunohistochemical staining was found in 95.9% of all interpretable colorectal cancers. Loss of RBM3 expression was linked to advanced tumor stage (p < 0.0001), right-sided tumor localization (p < 0.0001) and poor prognosis (p = 0.0003). In a multivariate analysis, including RBM3 staining, tumor grade, tumor stage and nodal status, only tumor stage and nodal status proved to be independent prognostic markers (p < 0.0001 each), whereas the prognostic impact of RBM3 staining was not significant (p = 0.2655).

Conclusion: Our observations indicate that loss of RBM3 expression is an unfavorable prognostic marker in CRC and linked to right-sided tumor localization.

Disclosure: No conflict of interest disclosed.

P210 - βIII-tubulin overexpression is linked to left-sided tumor localization and nuclear β-catenin expression in colorectal cancer

Marx A.H.1, Simon R.1, Bokemeyer C.2, Terracciano L.3, Sauter G.1, Izbicki J.R.4, Melling N.4

1University Medical Center Hamburg-Eppendorf, Pathology, Hamburg, Germany, 2Hubertus Wald Cancer Center, University Medical Center Hamburg-Eppendorf, Department of Oncology, Hematology, BMT with section Pneumology, Hamburg, Germany, 3University Hospital Basel, Pathology, Basel, Switzerland, 4University Medical Center Hamburg-Eppendorf, Surgery, Hamburg, Germany

Introduction: βIII-tubulin expression correlates with poor outcome in various malignancies.

Materials and methods: βIII-tubulin expression was analyzed by immunohistochemistry on a tissue microarray containing 1800 colorectal cancers. Results were compared to clinic-pathological and molecular parameters.

Results: βIII-tubulin expression was detectable in 79.2% of 1619 interpretable colorectal cancers. High βIII-tubulin expression was associated with left-sided tumor localization (p = 0.0303) and nuclear β-catenin expression (p = 0.003). High βIII-tubulin expression was not linked to the gender of the patient (p = 0.5842).

When all tumors were analyzed the prognostic role of βIII-tubulin expression was not independent of pT stage, pN stage, tumor grade or tumor localization (p = 0.0517).

Conclusion: βIII-tubulin expression is not an independent prognostic parameter in colorectal cancer. The significant association with left-sided tumor localization and a key genomic alteration of colorectal cancer such as β-catenin suggest interaction with important pathways involved in colorectal cancer.

Disclosure: No conflict of interest disclosed.

P211 - Subgroup analysis of patients with metastatic colorectal cancer (mCRC) treated with regorafenib (REG) in the CORRECT trial who had progressionfree survival (PFS) longer than 4 months

Stein A.1, Grothey A.2, Sobrero A.3, Siena S.4, Falcone A.5, Ychou M.6, Humblet Y.7, Bouché O.8, Mineur L.9, Barone C.10, Adenis A.11, Tabernero J.12, Yoshino T.13, Lenz H.-J.14, Goldberg R.M.15, Xu L.16, Wagner A.17, van Cutsem E.18

1Universitätsklinikum Hamburg-Eppendorf, II. Medizinische Klinik und Poliklinik, Hamburg, Germany, 2Mayo Clinic, Rochester, United States, 3San Martino Hospital, Genova, Italy, 4Ospedale Niguarda Ca' Granda, Milan, Italy, 5Ospedaliero-Universitaria Pisana, Pisa, Italy, 6ICM Val d'Aurelle, Montpellier, France, 7Cliniques Universitaires Saint-Luc, Brussels, Belgium, 8Centre Hospitalier Universitaire Robert Debré, Reims, France, 9Institut Sainte-Catherine, Avignon, France, 10Catholic University of Sacred Heart, Roma, Italy, 11Centre Oscar Lambret, Lille, France, 12Vall d'Hebron University Hospital, Barcelona, Spain, 13National Cancer Center Hospital East, Kashiwa, Japan, 14University of Southern California/Norris Comprehensive Cancer Center, Los Angeles, United States, 15Ohio State University Comprehensive Cancer Center, Columbus, United States, 16Bayer HealthCare Pharmaceuticals, Whippany, United States, 17Bayer Pharma AG, Berlin, Germany, 18University Hospital Gasthuisberg, Leuven, Belgium

Introduction: In the CORRECT phase III trial (NCT01103323), the multikinase inhibitor REG significantly improved overall survival (OS) and PFS vs placebo in patients with mCRC who had disease progression after other standard therapies (HR for OS: 0.77; 1-sided p = 0.0052; Grothey, 2013). A post hoc exploratory subgroup analysis was conducted on patients in the REG treatment group who had a PFS longer than 4 months (long PFS).

Methods: PFS was defined as the time from randomization to disease progression (radiological or clinical) or death from any cause (whichever occurs earlier). Of the 505 patients randomized to REG in CORRECT, 98 (19%) were classified as having long PFS benefit, defined as a PFS event >4 months after randomization. Baseline patient characteristics, safety, and dosing parameters were analyzed.

Results: The long PFS subpopulation was representative of the overall REG-treated population (table). Long PFS patients received a median of 6 cycles of REG (range 1-12); 92% received ≥5 cycles, and 20% had >8 cycles. Adverse events (AEs) of any grade were experienced by all long PFS patients, and the most common grade ≥3 AEs were hand-foot skin reaction (20%), hypertension (17%), diarrhea (17%), and fatigue (16%).

Conclusion: A subset of 98 (19%) patients treated with REG in the CORRECT study had a PFS >4 months, confirming the clinical benefit and tolerability of REG as a treatment option for patients with mCRC. Patients with long PFS generally had better ECOG PS and fewer tumor sites than the overall population. Prospective validation of these findings in conjunction with biomarker analysis from real-life clinical experience is needed.


Disclosure: Alexander Stein: Financing of Scientific Research: Bayer, Roche, Sanofi, Merck Serono, Amgen; Expert Testimony: Roche, Sanofi. Eric van Cutsem: Expert Testimony: Bayer.

P212 - Tp53-dependent response to chemo- and radiotherapy in colorectal cancer (CRC) cell lines

Milazzo A.1, Kanz L.1, Schittenhelm M.M.1, Kampa-Schittenhelm K.1

1Universitätsklinikum, Tübingen, Germany

Background: Colorectal cancer is one of the most common malignancies with high prevalence and low 5-year survival in the advanced stages. Inactivating mutations of the tp53 gene are associated with resistance to therapy in several tumor models, conferring poor prognosis. In CRC, where tp53 mutations occur in nearly half of the cases, the role of tp53 remains unclear. The objective of our study was to evaluate the role of tp53 signaling for response to standard chemotherapy. In addition, we tested whether combination approaches with radiotherapy might overcome therapy resistance in mutant-tp53 CRC - thus potentially providing a novel molecular-defined therapeutic rationale in locally advanced disease.

Methods: Three independent isogenic CRC cell line models (HCT116, RKO, DLD-1), harboring a genetically modified tp53-null variant or tp53 wild type (wt) were generously provided by Dr. Vogelstein (JHU, Baltimore, MD). Cells were treated with 5-Fluoruracil (5-Fu), Oxaliplatin (Ox) or Irinotecan (Ir) with escalating doses and/or gamma irradiation (5Gy) and effects on cellular proliferation, viability, induction of apoptosis and cell cycle were assessed.

Results: We show, that proapoptotic effects mediated via 5-Fu, IR and Ox depend on tp53: significantly higher rates of proapoptotic cells were observed in a dose-dependent manner for the tp53 wt cell strains compared to the corresponding tp53 null cells (with the exception of Ir in DLD-1 cells). In contrast, exposure of tp53 wt vs. tp53 null cell strains tp gamma irradiation at 5 Gy did reveal equipotent p53-independent induction of apoptosis - providing the rational to combine radiation therapy with chemotherapy in mutant-tp53 CRC. Indeed and in particular the tp53 null strains benefited when radiotherapy was combined with either of the chemotherapeutics. Importantly, in the tp53 wt cell strains, we observed adverse effects: Consistent for all wt cell lines, addition of radiotherapy to either agent decreased the proapoptotic effect seen for chemotherapy alone - partly due to unfavorable cell cycle arrest phenomena.

Conclusion: Our findings demonstrate a role of intact p53 signaling for the efficacy of Ox, IR and 5Fu in CRC models. Radiochemotherapy is only of benefit in mutant-tp53 cancer, which provides a rationale for tp53 mutation screening as a helpful tool for assessment of therapeutic options.

Disclosure: No conflict of interest disclosed.

P213 - Analysis of the quality of life in patients treated with Aflibercept and FOLFIRI for metastatic colorectal cancer (mCRC) - Interim results from the non-interventional QoLiTrap study

Derigs H.-G.1, Scholten F.1, Losem C.2, Kröning H.3, Windemuth-Kieselbach C.4, Hofheinz R.-D.5

1Klinikum Frankfurt Höchst, Klinik für Innere Medizin 3, Frankfurt a. M., Germany, 2Johanna-Etienne Krankenhaus, Praxis für Hämatologie und Onkologie, Neuss, Germany, 3Schwerpunktpraxis für Hämatologie und Onkologie, Magdeburg, Germany, 4Alcedis GmbH, Gießen, Germany, 5Universitätsmedizin Mannheim, Interdisziplinäres Tumorzentrum, Mannheim, Germany

Introduction: Aflibercept is an anti-angiogenic fusion protein targeting VEGF-A, VEGF-B and PlGF. It is approved in combination with FOLFIRI for patients with mCRC, who have already received an Oxaliplatin containing therapy.

Methods: Patients with mCRC receiving Aflibercept according to label are included in QoLiTrap, an ongoing non-interventional study to evaluate quality-of-life in patients (AIO-LQ-0113). The study aims to enroll 1500 patients in Germany, Austria and Switzerland. The primary aim is to assess health related quality of life in clinical practice. EORTC-QLQ C30 questionnaires are filled in by the patients at baseline and in every other cycle.

Results: This is the interim analysis of the first 348 included patients who have received at least one cycle of Aflibercept. 329 patients have completed EORTC-QLQ C30 at baseline and 218 patients completed at least baseline and 2 additional questionnaires. These 218 patients received Aflibercept therapy in 1th (10%), 2nd (48%), 3rd (19%) and later lines (22%) with a median of 6 (and up to 35) cycles. Median age was 66 years with good PS (ECOG 0: 38%; ECOG 1: 45%). Complete or partial remission was observed in 12% of all evaluable patients over all lines, SD in 27% and PD in 14% (response not yet documented for 47%). At baseline, patients presented a median EORTC global health score of 58.3 (n = 213). Differences between baseline and mean values after 12 weeks were observed in global health status as well as in physical-, emotional-, cognitive-, role- and social functioning. Changes in physical functionality were mainly driven by an increase in fatigue, whereas in other symptom scales such as constipation and financial impact the value declined. This first analysis from the QoLiTrap study has identified no new safety signals.

Conclusion: QoLiTrap is a large observational study conducted to analyze the quality of life in patients with mCRC treated with FOLFIRI & Aflibercept. This interim analysis shows preliminary, yet encouraging results with PR or SD adding to a tumor control rate of 74% within the cohort of patients with response documentation. The decline in global health status was moderate. Updated results will be presented at the meeting.

This study is supported by Sanofi.

Disclosure: Hans-Günter Derigs: No conflict of interest disclosed. Ralf-Dieter Hofheinz: Financing of Scientific Research: Ja, Firmen Sanofi, Roche, Merck, Amgen, Bayer, medac; Expert Testimony: Ja, Firmen Sanofi, Roche, Merck, Amgen, medac.

P214 - Intravenous ferric carboxymaltose vs. oral iron substitution in patients with metastatic colorectal cancer (mCRC) and iron deficiency anemia: a randomized multicenter treatment optimization study (a study in progress report)

zur Hausen G.1, Rötzer I.2, Reichart A.1, Pauligk C.1, Hunfeld K.-P.3, Hozaeel W.1, Quidde J.4, Hofheinz R.-D.5, Al-Batran S.-E.1

1Krankenhaus Nordwest gGmbH, Institut für Klinisch-Onkologische Forschung, Frankfurt am Main, Germany, 2Krankenhaus Nordwest gGmbH, Klinik für Onkologie und Hämatologie, Frankfurt am Main, Germany, 3Krankenhaus Nordwest gGmbH, Labormedizin, Mikrobiologie und Krankenhaushygiene, Frankfurt am Main, Germany, 4Universitätsklinikum Hamburg-Eppendorf, Hubertus Wald Tumorzentrum, Hamburg, Germany, 5Universitätsmedizin Mannheim, III. Medizinische Klinik, Mannheim, Germany

Introduction: Iron deficiency has a high prevalence in colorectal cancer patients ranging at ca. 60%. About 70% of these patients suffer from iron deficiency anemia (IDA) which adds both physical and cognitive impediments to an already straining chemotherapy. Moreover, a chronic disease like cancer often results in a reduced availability of iron for the body. In clinical practice iron substitution is usually administered orally. Due to low resorption rates, frequent gastric side effects and thus poor patient compliance a parenteral substitution seems to be a better option in terms of efficacy. In the framework of a randomized multicenter clinical trial (‘FerInject') a comparison of efficacy parameters of parenteral vs. oral iron substitution will now be conducted in order to identify the best treatment form for clinical practice in oncology. Furthermore detailed quality of life-data (QoL) will be collected in both treatment arms for effect comparison.

Methods: Randomized explorative multicenter phase II trial: mCRC patients with IDA and a concomitant palliative chemotherapy will be randomly assigned to one of the treatment arms (parenteral vs. oral). Parenteral patients receive up to 1.000 mg ferric carboxymaltose intravenously per week for a maximum of 2 weeks. Oral patients take 200 mg iron per day for a total of 12 weeks.

Course of therapy and QoL-data will be collected. Primary endpoint is the increase or normalization of hemoglobin. Secondary endpoints are fatigue; QoL; handgrip strength; number of allogenic blood transfusions; time until rise or normalization of hemoglobin; genesis of the IDA; number, dose and duration of therapy with recombinant erythropoietin; inflammatory parameters; influence of nutritional status on IDA and therapy success; tolerance and toxicity; dropout rate and overall survival. 64 patients shall be enrolled at currently 12 study sites.

Data will be evaluated exploratively due to lacking reference data. The development of hemoglobin status in the treatment arms will serve as an indicator for a trend towards a better treatment option if a level of significance p = 0.2 (Fisher's exact test) can be detected.

Results: Recruitment started in 04/2015.

Conclusions: FerInject is a trial in progress with high relevance for the clinical management of mCRC patients with IDA. Furthermore it will return additional information to both efficacy and QoL issues of these patients under a concomitant palliative chemotherapy.

Disclosure: Gerrit zur Hausen: No conflict of interest disclosed. Salah-Eddin Al-Batran: Expert Testimony: Research grant (Vifor)


Lymphome aggressiv

P215 - Systemic relapses in primary CNS lymphoma (PCNSL)

Korfel A.1, Fischer L.2, Hummel M.3, Lenze D.4, Hommel A.1, Weller M.5, Roth P.5, Kreher S.1

1Medizinische Klinik m.S. Hämatologie, Onkologie und Tumorimmunologie, Charité - Universitätsmedizin, Berlin, Germany, 2Zentrum für Onkologie und Urologie, Rostock, Germany, 3Institut für Pathologie - Molekulare Diagnostik, Charité - Universitätsmedizin, Berlin, Germany, 4Institut für Pathologie CBF - Molekulare Diagnostik, Charité - Universitätsmedizin, Berlin, Germany, 5Department of Neurology University Hospital Zurich, Zürich, Switzerland

Introduction: PCNSL is a rare lymphoma confined to the CNS at the time of diagnosis. A strong affinity to the CNS is typical for PCNSL with systemic relapse (SR) occurring very rarely.

Methods: We retrospectively analysed two prospective collectives of immunocompetent adult PCNSL patients: 411 from a multicenter therapy study (G-PCNSL-SG-1; collective 1) and 51 treated for PCNSL at Charité Berlin from 1994 to 2007 (collective 2). Within the collective 1, patients with SR were compared to patients who did not develop an SR during the whole follow-up. When possible, tumor tissue from SR was compared to the primary CNS tumor, and clonality analysis was performed using PCR for immunoglobulin heavy/light chain (IgH/L-PCR, Biomed2 protocol).

Results: SR was found in 24 of 411 patients in collective 1 (5.8%) and in 6 of 51 (11.7%) in collective 2 for a total of 30 SR patients. Median time to the first SR was 25.3 months (range 2-97). SR was found at first relapse in 26 patients; at the second relapse in 6 and at the third relapse in 4 patients for a total of 36 SR. The majority of SR were localised extranodally and was not accompanied by CNS relapse (30/36 = 83% each). Median survival from the first SR was 9.9 months (95% CI 3.0-16.9). No significant differences in initial characteristics were found between SR and non-SR patients within collective 1. The outcome of SR versus non-SR patients was similar: median overall survival from PCNSL diagnosis 30.6 months (95% CI 0-64) versus 34.4 months (95% CI 28.5-40.3; p = 0.960); median progression free survival from PCNSL diagnosis to first relapse 15.5 months (95% CI 0-31.6) versus 12 months (95% CI 8.9-15.1), respectively (p = 0.355). Clonality analysis of SR and corresponding primary CNS tumor was performed in 9 patients. Concordant clonality was detected in 6 (67%), discordant clonality in 3 (33%) patients.

Conclusions: SR is rarely diagnosed in PCNSL when restaging does not routinely include search for systemic disease. Patients who develop a SR do not show differences at the time of PCNSL diagnosis as compared to those who do not develop it. The usually extranodal SR localisation underlines the particular tropism of PCNSL. The prognosis of SR patients is comparable to that of non-SR patients. Most of SR showed the same Ig rearrangement as the primary CNS tumor. However, discordant clonality in one third of analysed patients indicated the systemic lymphoma as second malignancy independent of PCNSL.

Disclosure: No conflict of interest disclosed.

P216 - Cotargeting of PIM, PI3K and mTOR in mantle cell lymphoma

Freysoldt B.1,2, Schnaiter A.1,2, Zimmermann Y.1,2, Hutter G.1,2, Hiddemann W.1,2, Dreyling M.1,2

1Helmholtz Zentrum, München, Germany, 2Medizinische Klinik und Poliklinik III, Klinikum Großhadern, LMU, München, Germany

Introduction: Mantle cell lymphoma (MCL) comprises about 6% of all non-Hodgkin´s lymphoma with a median survival of 3-5 years. The proviral insertion in murine (PIM) lymphoma proteins are serine/threonine kinases which play an important role in cell survival and proliferation. They are overexpressed in multiple human cancers, including haematological malignancies. On the other hand, PI3K inhibition achieves high, but temporary responses in relapsed MCL. In this study we evaluated the efficiency and mode of action of a dual PIM/PI3K and a triple PIM/PI3K/mTOR-Inhibitor in MCL cell lines.

Methods: MCL cell lines (Granta 519, Jeko-1, Rec-1 and Mino) were exposed to a dual PIM kinase/PI3K inhibitor (IBL202) and triple PIM-kinase/PI3K/mTOR inhibitor (IBL301). Cell proliferation (trypan blue staining), apoptosis (Annexin V PE/7-AAD staining) and cell cycle (FACS) were investigated. Protein expression and phosphorylation status of downstream proteins (Akt, GSK-3β, 4EBP1) as well as markers of apoptosis (PARP, Caspase 9) were analysed after 1h, 4h, 8h and 24h.

Results: Both IBL202 and IBL301 appear to be effective inhibitors, with the triple inhibitor IBL301 being superior to the dual inhibitor IBL202. IBL301 had a much higher impact on cell proliferation than IBL202 in all tested MCL cell lines, possibly due to its mTOR inhibiting activity. Both, treatment with IBL202 and IBL301, induced cell death, but cell death rate was almost twice as high in IBL301 treated MCL cell lines. The differential impact of the two inhibitors could be also confirmed based on considerably higher PARP and Caspase 9 cleavage after treatment with IBL301. Both inhibitors led to G1 arrest. In Jeko-1, Granta-519 and Mino dephosphorylation of Akt was detected after treatment with both agents, again with a less prominent effect under treatment with IBL202, supporting the mode of action of both inhibitors through the PI3K-AKT pathway. Accordingly, dephosphorylation of GSK-3β was observed after both agents in all MCL cell lines. The decrease of phosphorylated proteins occurred during the first hour of treatment, while subsequently phosphorylation increased again up to the level of controls.

Conclusions: The triple inhibitor of PIM kinases, PI3K and mTOR shows impressive efficiency in MCL cell lines, superior to the dual inhibition of PIM kinase and PI3K. Cotargeting PIM kinases, PI3K and mTOR represents a promising novel approach in MCL.

Disclosure: No conflict of interest disclosed.

P217 - ABC, GCB, and extra nodal DLBCL: Three sides of a story or three stories with similar sides?

Bohlen J.1, Hallas C.2, Preukschaß M.2, Tiemann M.2

1Semmelweis Universität Medizinische Fakultät, Asklepios Campus Hamburg, Hamburg, Germany, 2Hämatopathologie Hamburg, Molekularpathologie, Hamburg, Germany

Since ABC (activated B-cell type) and GCB (germinal center B-cell type) have been described as separate entities of DLBCL (diffuse large B-cell lymphoma)1,2 efforts have been underway to define these entities on a molecular basis. In ABC lymphoma activation of the NF-kB pathway was described as a key feature and this activation has been linked to mutations in the B-cell receptor pathway and MYD88 mutations. The defining features of GCBs are far less well understood. Less attention has been paid to the primary localization of the tumor and its effect on mutation patterns.

DLBCL were classified as non-GCB or GCB subtype according to the Visco-Young immunohistochemistry algorithm and analyzed for mutations in MYD88, CD79A/B, and CARD11 by Sanger Sequencing and real time PCR. Statistical comparisons were made using Fisher's exact test.

As expected, significantly more non-GCB lymphomas carried mutations in MYD88, CD79A/B, and CARD11 than GCB lymphomas (19/48 vs. 4/31 cases, p < 0.02) when only lymphomas of primary nodal origin were considered. In extra nodal lymphomas no difference in mutation pattern was seen between GCB and non-GCB type (overall mutations: 18/40 vs. 9/23, n.s.). Whereas CARD 11 mutations were evenly distributed among all entities investigated (GCB and non-GCB, nodal and extranodal), CD79 mutations were not detected in primary nodal GCB lymphomas and MYD88 mutations only in one case. Overall, the distribution of mutations in all primary extra nodal lymphomas strongly resembled the mutation pattern in nodal non-GCB lymphomas and was significantly different from GCB (30/70 vs. 4/31, p < 0.005).

Whereas primary nodal DLBCL consist of two molecularly different entities (ABC and GCB) these molecular differences do not exist in primary extra nodal lymphomas. Although it is technically possible to use the Visco-Young algorithm to subtype extra nodal DLBCL the resulting categories do not reflect molecular subtypes. Primary extra nodal DLBCL should be regarded as subtype/subtypes of its own with some features shared with non-GCB DLBCL.


1 Nature. 2000:503-511.

2 Proc Natl Acad Sci USA. 2008:13520-13525.

Disclosure: No conflict of interest disclosed.

P218 - MiR-199a and miR-497 are associated with better overall survival due to increased chemosensitivity in aggressive non-Hodgkin´s lymphoma patients

Troppan K.1, Wenzl K.1, Pichler M.2, Pursche B.1, Schwarzenbacher D.3, Feichtinger J.4, Thallinger G.5, Beham-Schmid C.6, Neumeister P.1, Deutsch A.1

1Medical University Graz, Hematology, Graz, Austria, 2MD Anderson Cancer Center, Department of Experimental Therapeutics, Houston, United States, 3Medical University Graz, Oncology, Graz, Austria, 4University of Technology, Institute for Genomics and Bioinformatics/Institute of Biochemistry, Graz, Austria, 5Unversity of Technology, Institute for Genomics and Bioinformatics/Institute of Biochemistry, Graz, Austria, 6Medical University Graz, Pathology, Graz, Austria

Background: Micro-RNAs (miRNAs) are short non-coding single-stranded RNA molecules regulating gene expression at the post-transcriptional level. MiRNAs are involved in cell development, differentiation, apoptosis, and proliferation. MiRNAs can either function as tumor suppressor genes or oncogenes in various important pathways. The expression of specific miRNAs has been identified to correlate with tumor prognosis.

Methods: For miRNA expression analysis real-time PCR on 81 samples was performed, including 63 diffuse large B-cell lymphoma (DLBCL, 15 of germinal center B-cell like subtype, 17 non germinal center B-cell, 23 transformed, and 8 unclassified) and 18 controls, including 9 peripheral B-cells, 5 germinal-center B-cells (GC), 4 lymphadenitis samples, and 4 lymphoma cell lines (RI-1, SUDHL4, Karpas, U2932). Expression levels of a panel of 11 miRNAs that have been previously involved in other types of cancer (miR-15b_2, miR-16_1*, miR-16_2, miR-16_2*, miR-27a, miR-27a*, miR-98-1, miR-103a, miR-185, miR-199a, and miR-497) were measured and correlated with clinical data. Furthermore, cell lines, lacking miR-199a and miR-497 expression, were electroporated with the two respective miRNAs and treated with standard immunochemotherapy routinely used in patients with DLBCL, followed by functional analyses including cell count and apoptosis assays.

Results: 7 miRNAs (miR-16_1*, miR-16_2*, miR-27a, miR-103, miR-185, miR-199, and miR-497) were statistically significant up-regulated in DLBCL compared to normal GC. However, high expression of miR-497 or miR-199a was associated with better overall survival (p = 0.042 and p = 0.007). Overexpression of miR-199a and miR-497 led to a statistically significant decrease in viable cells in a dose-dependent fashion after exposure to rituximab and various chemotherapeutics relevant in multi-agent lymphoma therapy.

Conclusion: Our data indicate that elevated miR-199a and miR-497 levels are associated with improved survival in aggressive lymphoma patients most likely by modifying drug sensitivity to immunochemotherapy. This functional impairment may serve as a potential novel therapeutic target in future treatment of patients with DLBCL.

Disclosure: No conflict of interest disclosed.

P219 - Outcome of patients with transformed CLL and FL compared to patients with relapsed DLBCL - retrospective analysis of the Freiburg cohort

Bach A.1,2, Greil C.2, Marks R.2

1Charité - Universitätsmedizin Berlin, Radiologie, Berlin, Germany, 2Uniklinik Freiburg / Innere Medizin / Hämatologie, Onkologie und Stammzelltransplantation, Freiburg, Germany

Introduction: Patients with transformed chronic lymphatic leukaemia (CLL) have poor prognosis despite dose-intensified therapies. We determined outcome parameters for all patients transformed from CLL or follicular lymphoma (FL) into diffuse large B-cell lymphoma (DLBCL), using patients relapsed from DLBCL as comparison group.

Methods: All patients with either FL or CLL and subsequent transformation into DLBCL or relapsed DLBCL (rDLBCL) who were treated in the University Hospital Freiburg between 2000 and 2011 were analysed. Overall survival (OS) after transformation/relapse and disease free survival (DFS) after first complete remission (CR) were evaluated.

Results: 98 patients were identified, 16 (16.3%) with transformed FL (tFL), 14 (14.3%) with transformed CLL (tCLL) and 68 (69.4%) with rDLBCL.

At the time of transformation/relapse, patients with tFL, tCLL and rDLBCL were treated with either chemotherapy alone (25%, 64%, 49%), autologous (autoSCT; 56%, 0%, 38%) or allogeneic transplantation (alloSCT; 13%, 29%, 3%).

While tFL and rDLBCL patients experienced a CR in 56%/57% with this first line treatment, only 21% of tCLL patients achieved CR. After a median follow-up of 6.8 years, this inferior response was followed by a five-year OS (5y-OS) of only 14%, compared to 38% for tFL (p = 0.1912) and 27% for rDLBCL patients.

Only 7% of all tCLL patients experienced longterm disease control, exclusively after alloSCT. In contrast, of all tFL patients in CR after initial therapy, to date 67% showed no signs of disease.

A low/low-intermediate International Prognostic Index (IPI) at the time of transformation/relapse resulted in a significantly better median survival - especially for tCLL patients (32 months vs. 1.5 months for patients with high/high-intermediate IPI, p = 0.019).

Regarding therapy, tCLL patients treated with alloSCT had improved median OS compared with those who received chemotherapy only (42 vs. 6 months, p = 0.1132). For tFL patients treatment with autoSCT resulted in a better median OS (24 vs. 6.5 months with chemotherapy; p = 0.3654).

Conclusions: Overall survival of transformed low grade lymphoma is strongly influenced by the underlying disease with more favourable outcome for patients with FL compared to CLL. Dose intense therapy including autoSCT results in the best treatment outcome for patients with tFL or rDLBCL, while longterm disease control of transformed CLL patients can only be achieved with alloSCT.

Disclosure: No conflict of interest disclosed.

P220 - Superantigen recognition is a specific hallmark of mantle cell lymphoma-derived B-cell receptors in a substantial proportion of patients

Fichtner M.1, Spies E.1, Seismann H.1, Dreyling M.2, Binder M.1, Klapper W.3, Trepel M.1,4

1Department of Oncology and Hematology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany, 2Department of Medicine III, University Hospital Grosshadern, Munich, Germany, 3Institute of Pathology, Division of Hematopathology, University Medical Center Schleswig-Holstein, Kiel, Germany, 4Department of Hematology and Oncology, Augsburg Medical Center, Augsburg, Germany

Introduction: Mantle cell lymphoma (MCL) is a B cell Non-Hodgkin's lymphoma with a poor prognosis. The success of agents targeting the B-cell receptor (BCR) signaling suggests a major role for this pathway in mantle cell lymphoma. However, it remains unclear what antigens trigger BCR activation in this disease.

Methods: We used an unbiased PCR approach to amplify and sequence the BCR heavy and light chains of 24 randomly chosen MCL patients. The variable regions of the MCL BCRs were cloned into an IgG or Fab expression vector and expressed using Sf9 cells. In total, we produced eleven different recombinant monoclonal IgG antibodies and five Fab-fragments. With this set of immunoglobulins, we performed phage display library screenings, Western blottings, and immunofluorescence stainings as well as Protein A-ELISAs and Protein A-immunoprecipitations.

Results: In line with previous research, the BCRs of our cohort were strongly biased, showing a preference for the usage of vH3-family genes (11/24). Further, all BCRs bearing the vH3-21 heavy chain were paired with vL3-19 lambda light chain (5/24). In addition, 5 of 24 samples used the vH434-family. Nearly all heavy chain sequences were not or only minimally mutated (≥98% germline identity). In the majority of cases, selections with random peptide phage libraries did not enrich peptides binding specifically to these MCL BCRs. Together, this suggested that classical high-affinity antigen recognition may not play a leading role in BCR function of MCL. Therefore, we explored the hypothesis that MCL BCRs might be activated by low-affinity, complementarity determining regions (CDR)-independent, superantigen-like or glycosylation-mediated interactions. While no BCR acquired novel glycosylation sites, all VH3 family expressing BCRs (11/24 cases) showed sequence features indicative for interaction with the superantigen protein A, which could be confirmed by Fab binding and immunoprecipitation studies. Furthermore all VH4-34 expressing BCRs (5/24 cases) exhibited motifs associated with anti-i/I reactivity.

Conclusion: Our findings suggest that the BCR pathway may be activated by CDR-independent mechanisms in a substantial portion of patients with mantle cell lymphoma.

Disclosure: No conflict of interest disclosed.

P221 - Comparison of the efficiencies of different PI3K-Inhibitors in matle cell lymphoma

Hutter G.1,2, Zimmermann Y.1,2, Zoellner A.1,2, Irrgang P.1,2, Arnd J.1,2, Hiddemann W.1, Dreyling M.1,2

1Universitätsklinikum München-Großhadern, Med III, München, Germany, 2Helmholtz Institute Munich, CCG Leukemia, Munich, Germany

Introduction: Mantle cell lymphoma (MCL), is a distinct lymphoma subtype with an aggressive clinical course and a median survival of 3-5 years. New emerging strategies include inhibitors of the B-cell receptorpathway which is constitutively activated in MCL and plays a critical role in tumor growth and survival. In the present study we investigated different PI3K-Inhibitors (IPI-145, CAL101, A66, TGX221 and Bay236) targeting different isoforms of PI3K in MCL.

Methods: MCL cell lines (Z-138, Mino-1, Granta-519, Jeko-1, Rec-1, Maver-1) were exposed to different PI3K-Inhibitors (IPI-145, CAL101, A66, TGX221, Bay 236) with or without murine feeder layer (M210B4). The effect of drugs was evaluated by cell count (trypan-blue staining), cell metabolism (WST-assay), cell cycle (FACS) and apoptosis (Annexin V PE/7-AAD staining). Subsequently, combinations with other inhibitors of the PI3K/mTOR pathway were analysed. Western blot analyses were performed after exposure to the various inhibitors and were correlated to the sensitivity of cell lines. Finally, efficiency of drug combinations were confirmed in primary patient samples.

Results: The PI3K inhibitor- Bay236 was most effective in MCL cell lines followed by IPI-145, A66, TGX221 and CAL101 suggesting a higher efficiency by inhibiting multiple isoforms of PI3K. In patient samples Bay 239 also revealed the highest cytotoxicity followed by IPI-145, CAL101, TGX221 and A66 (75,1%; 65,9%; 50,6% and 40,3%, respectively). Comparing different combinations of single PI3K-isoform inhibitors the combination of A66 and CAL101 was most efficient resulting in a 65,2% reduction of cell count. Nevertheless a combination of multiple isoform-specific PI3K inhibitors (A66, TGX-221, CAL101) led to an even higher reduction of cell proliferation (33,4-66,6% ) than targeting only single isoforms (0-21,6%) confirming the superiority of targeting multiple PI3K isoforms. The higher cytotoxic effect was accompanied by higher dephosphorylation of phAKT, phRictor and phRSK. Results of the effect of PI3K inhibitors on MCL in the presence of a murine feeder layer will be shown.

Conclusion: Based on the comparative analysis of PI3K inhibitors inhibition of multiple isoforms of PI3K appears to be most efficient in MCL. Even more importantly, the murine feeder layer enhanced the effect of PI3K inhibitors indicating that the microenvironment plays a critical role for the mode of action of such inhibitors of the B-cell receptor pathway.

Disclosure: Grit Hutter: No conflict of interest disclosed. Martin Dreyling: Advisory Role: Scientific advisor, Bayer.

P222 - High dose Cytarabine and Thiotepa plus G-CSF is an effective combined salvage and stem cell mobilization regimen in central nervous- and advanced nodal lymphoma

Schäfer H.S.1, Schlosser T.1, Schorb E.1, Fritsch K.1, Finke J.1, Illerhaus G.2

1Universitätsklinikum Freiburg, Department innere Medizin, Klinik für innere Medizin 1, Freiburg, Germany, 2Klinikum Stuttgart, Klinik für Hämatologie, Onkologie und Palliativmedizin, Stuttgart, Germany

Introduction: For aggressive B-cell lymphoma high-dose chemotherapy (HDC) followed by hematopoietic stem cell transplantation (HSCT) is a common treatment approach. Therefor used induction and conditioning regimens have been extensively analyzed for response rates, survival rates and toxicity. The impact and effectiveness of mobilization regimen are much less investigated.

Methods: AraC (3 g/m2) was administered for 2 subsequent days in combination with thiotepa (40 mg/m2) on the 2nd day. Stem cells were mobilized with subcutaneous G-CSF. Stem cell apharesis was performed at Freiburg medical center according to GMP standard. Response and toxicity data were retrospectively assessed from mediacal record.

Results: We retrospectively analyzed 73 patients with primary (n = 59), secondary (n = 9) central nervous non-Hodgkin lymphoma (PCNSL, SCNSL) and advanced nodal lymphoma (n = 5) (ANL) treated with Cytarabine and Thiotepa (AraC/TT) plus G-CSF chemotherapy as mobilization regimen for peripheral blood stem cell (PBSC) collection at our institution. For all patients, proceeding to high-dose therapy with autologous stem cell rescue was intended.

Patients received 112 cycles of this regimen in total . Stem cell collection was performed after first or second cycle. The number of CD34 positive cells mobilized after AraC/TT was 18,55 × 10e6/kg cells (median; range 86,16 × 106 -1,05 × 106). Dose of G-CSF administered was 3360µg (median; range 1500µg-7200µg) in total. PBSC collection of more than 2 × 106/kg CD34 positive cells was successful in 68 (93,2%) patients, for 57 patients (78,1%) this could be done in a single collection. Toxicity of this regimen was mild. Adverse events were reported in 25 patients (34,7%) including febrile neutropenia (n = 19) and prolonged cytopenia leading to harvest failure (n = 3). All events were manageable.

Response to therapy (defined as SD, PR or CR) was seen in 60 (90,9%) patients, major response (PR, CR ) was seen in 30 (45,5%) of all patients.

Conclusion: We conclude that AraC/TT plus G-CSF has high anti lymphoma activity in PCNSL, SCNSL and ANL respectively with acceptable toxicity and excellent PBSC mobilizing characteristics.

Disclosure: No conflict of interest disclosed.

P223 - The B-cell receptor pathway is highly actived in canine DLBCL cell line CLBL-1

Zoellner A.-K.1, Ruetgen B.2, Hutter G.1,3, Zimmermann Y.1,3, Mohring M.4, Hiddemann W.3, Hirschberger J.5, Dreyling M.1,3

1Klinikum der Universität München, Medizinische Klinik und Poliklinik III, München, Germany, 2University of Veterinary Medicine, Vienna, Austria, 3Clinical Cooperative Group Leukemia, Helmholtz Center Munich, München, Germany, 4University Hospital Klinikum rechts der Isar, Nuclear Medicine, München, Germany, 5Centre for Clinical Veterinary Medicine, LMU, Clinic of Small Animal Medicine, München, Germany

Background: Diffuse large B-cell lymphoma (DLBCL) is the most common subtype of canine lymphoma and shows significant similarities in its clinical and biological presentation to human Activated B-Cell (ABC) subtype of DLBCL.

Aims: Here we demonstrate the expression of hallmark kinases of the B-cell receptor pathway in the canine DLBCL cellline CLBL-1. Further we investigated the treatment with B-cell receptor pathway interacting compounds such as BTK inhibitor Ibrutinib and PI3K inhibitor Idelalisib and substances lately known for their anti-lymphoid activity.

Methods: Established canine DLBCL (CLBL-1) and T-cell lymphoma (CL-1) cell lines were cultivated under standard conditions in RPMI and exposed to ascending doses of Idelalisib, Ibrutinib, Temsirolimus, BX-912, Ku-63764,Enzastaurin and Bortezomib. Cell proliferation was determined after 48 hours based on WST cell proliferation assay. Western blotting was performed after 24h. All experiments were performed at least in triplicates.

Results: In Western blot analysis kinases hallmarking the B-cell receptor- PI3K-AKT pathway (AKT, PDK, PI3K, mTOR) and their phosphorylated isoforms were detected in CLBL-1 cells. Untreated CLBL-1 cells expressed p42/44,p38, MEK, GSK alpha and GSK beta and their phosphorylated isoforms as well as the cyclins CDK2, CDK4, CDK7, CDK9 but no cyclin D1. Significantly, treatment with only 1,25nM Ibrutinib induced in WST analysis a growth reduction to 45%, with 1µM arresting growth thoroughly. The PI3K-delta inhibitor Idelalisib showed dose dependent effects: 0,6µM reduced cell growth to 41%, whereas 5µM reduced proliferation to 13%. The mTor inhibitor Temsirolimus showed high efficacy: 1,25nM Temsirolimus reduced cell proliferation to 38%, while the mtorc1/mtorc2 inhibitor Ku-63794 induced at the dose of 0,25µM a reduction to 49%. CLBL-1 was also sensitive towards other compounds with anti-lymphoid activity such as the PDK-1 inhibitor BX-912 (0,25µM; 20%), the PKC inhibitor Enzastaurin (1,25µM; 52%) and the proteasome inhibitor Bortezomib (5nM; 50%).

Conclusion: The detected activated B-cell receptor pathway in CLBL-1 and the sensitivity towards small molecule inhibitors targeting this pathway indicates the similarity to human ABC DLBCL. This data strongly supports the relevance of canine DLBCL as model for its human counterpart.

Disclosure: Anna-Katharina Zoellner: No conflict of interest disclosed. Martin Dreyling: Financing of Scientific Research: Bayer: Speakers Honoraria; Expert Testimony: Bayer: Support of IITs.

P224 - Xenograft mouse model of primary and secondary CNS lymphoma

Isbell L.K.1, Reinacher P.C.2, Klingner K.3, Doostkam S.4, Tschuch C.3, Schorb E.1, Fritsch K.1, Schaefer H.1, Illerhaus G.5, Schaefer H.-E.6, Duyster J.1, Schueler J.3, von Bubnoff N.1

1Uniklinik Freiburg, Klinik für Innere Medizin I, Hämatologie, Onkologie und Stammzelltransplantation, Freiburg, Germany, 2Uniklinik Freiburg, Abteilung für Stereotaktische und Funktionelle Neurochirurgie, Freiburg, Germany, 3Oncotest GmbH, Freiburg, Germany, 4Uniklinik Freiburg, Institut für Neuropathologie, Freiburg, Germany, 5Klinikum Stuttgart, Klinik für Hämatologie, Onkologie und Palliativmedizin, Freiburg, Germany, 6Uniklinik Freiburg, Institut für Klinische Pathologie, Freiburg, Germany

Background: Extranodal diffuse large B-cell lymphoma confined to the central nervous system defines primary CNS lymphoma (PCNSL). In addition, CNS involvement can occur in systemic diffuse large B-cell lymphoma (secondary central nervous system lymphoma, SCNSL). Mechanisms that commit lymphoma cells to the CNS compartment are poorly understood. Despite intensive treatment, many patients suffer from relapse. The generation of functional and reproducible in vivo models is a prerequisite for a better understanding of cerebral lymphoma and for the development of improved therapeutic strategies.

Methods: We established a central nervous system lymphoma (CNSL) xenograft mouse model. Human PCNSL and SCNSL stereotactic biopsies are suspended into a transportation medium and directly transplanted subcutaneously in recipient NSG/NOG mice. Subcutaneous tumor growth was monitored by caliper measurement and mice were sacrificed with a tumor size of 2000 mm³. One third of the tumor as well as spleen, liver and brain were analyzed by histology and immunohistochemistry. One third of the tumor was snap frozen for subsequent genetic analysis, and one third was serially transplanted into NSG/NOG recipient mice.

Results: We observed growth of transplanted cells from PCNSL samples (n = 4) and SCNSL samples (n = 3) at the subcutaneous site of NSG/NOG mice. Histology and immunohistochemical analysis of the xenograft tumors showed the same morphology and immunophenotype as the primary human CNSL samples. Histopathologic analyses of spleen, liver, bone marrow and CNS samples of the tumor bearing mice are under investigation.

Conclusion: This xenograft mouse model allows in vivo studies on the biology of these rare tumors. Since PCNSL research is hampered by the small size of biopsy specimens, the expansion of the tumor cells in a xenografts model without losing the PCNSL phenotype may overcome this limitation. In addition, this tumor model allows us to compare lymphomas with and without CNS tropism with the goal to identify molecules that are critical for CNS tropism and thus could be potential new targets for therapeutic approaches. Furthermore this mouse model might be used to investigate novel pharmacologic agents to treat primary and secondary CNS lymphomas.

Disclosure: No conflict of interest disclosed.

P225 - The modification of acid ceramidase activity and ceramides level as an indicator of the drug resistance in patients by lymphoma

Abovyan M.1, Sahakyan L.1, Saharyan A.2, Shaljyan A.2

1Haematology Center aft. Prof.R.Yolyan, Yerevan, Armenia, 2Yerevan State Medical University, Yerevan, Armenia

Aim: The involvement of sphingolipids, and their metabolites, such as ceramides in cancerogenesis was demonstrated in solid tumors as well as in hematological malignancies. As is known, the appearance of multidrug resistant phenotype (MDR) - one of the major causes of failures in the treatment of malignant diseases. According to recent years' data, ceramide signaling plays an important role in tumor progression and development of chemoresistance. The present study was to examine the modification of acid ceramidase (aCD) activity and ceramides level as an indicator of the drug resistance in patients by lymphoma.

Methods: The study included 123 patients with B-cell nonHodgkin's lymphoma (NHL), which were admitted to the Hematology Center of Armenia. After obtaining informed consent were evaluated clinical and hematological parameters of all patients. Patients´ blood was taken before and after the treatment. For the normal control was used blood from 31 healthy donors of Hematology center. Ceramide has been determined using high performance liquid chromatography (HPLC). The aCDase activity determination was conducted according to the fluorogenic methods (Bedia C. et al., 2010).

Results: According to obtained data two of 105 newly diagnosed and untreated NHL patients (1.9%) and seven of 18 previously treated patients and drug-resistant (38.9%) had detectable high levels of ceramides and twofold increased activity of aCD. Due to failure of therapy, 7 patients died in the treatment first trimesters, the remaining 2 patients 6 and 8 months ago. Development of MDR in NHL is in part driven by the inherent genetic heterogeneity and instability of the tumor cells. Our results suggest that ceramides level and ceramidase activity is a potential pharmacologic target for the NHL treatment. The inhibition of ceramides expression or ceramidase activity might represent a novel strategy to sensitize B-cell NHL patients to chemotherapy.

Conclusions: Sphingolipids metabolites can be considered promising therapeutic tools alone or in combination with other compounds, as well as valid targets in the attempt to lymphoma treatment and overcome drug resistance.

Disclosure: No conflict of interest disclosed.

P226 - Evaluation of safety, tolerability and efficacy of Temsirolimus in patients with relapsed or refractory mantle cell lymphoma (rel/refr MCL) in routine clinical practice

Krekeler G.1, Neuhof A.1, Dreyling M.2, Hess G.3, Kalanovic D.1

1Pfizer Pharma GmbH, Berlin, Germany, 2Ludwig-Maximilians-Universität, München, Germany, 3Johannes-Gutenberg Universität, Mainz, Germany

Background: Temsirolimus (TEM), an mTOR-inhibitor, is approved in the EU for the treatment of patients (pts) with relapsed or refractory (rel/refr) MCL. A pivotal study demonstrated significantly longer progression free survival with TEM (175 mg weekly for 3 weeks followed by 75 mg weekly) in rel/refr MCL pts compared to investigator´s choice therapy (4.8 mo vs 1.9 mo; P = 0.0009). Yet only limited data is available on TEM in an unselected patient population during clinical routine. To evaluate the safety profile and efficacy of TEM in this rare tumor entity, further collection of data in a post-approval prospective non-interventional trial is useful.

Methods: A German multicenter registry for rel/refr MCL pts treated with TEM was started in Germany in Oct 2009 (NCT00700258) with regulatory and ethic committee´s approval. Objectives are the evaluation of the safety profile, tolerability and anti-tumor activity of TEM as well as the profile, comorbidities, characteristics and the sequence of systemic therapies.

Results: From Oct 2012 to Feb 2015, 27 study sites recruited 48 pts. Baseline characteristics: 72.9% male; median age 74.0 yrs; ECOG PS 0 or 1 in 83.0%, ECOG PS 2 in 17.0% of the pts. According to MIPI score 23.9%, 30.4%, and 45.7% of the pts are classified as low, intermediate and high risk at the time of enrollment. Bone marrow is involved in 41.7% of the pts. Median time between diagnosis and start of treatment with TEM is 3.1 yrs (range 0.4-14.9). The median number of prior therapies is 2.0 (range 1-10) with 45.9% treated in ≥4th line. Most common adverse events (≥15%) are thrombopenia (43.8%), anemia (25.0%), leukopenia (18.8%), general physical health deterioration (18.8%) and diarrhoea (16.7%). Severe adverse events (drug related) are general, metabolic, psychiatric, skin, renal, gastrointestinal, respiratory and blood system disorders (in 1 pt each) and infections (in 3 pts). Preliminary efficacy analyses are available for 31 assessable pts and show an objective response in 9 pts (1CR and 8 PR, 29.0%), a clinical benefit (CR, PR, MR and SD) in 17 pts (54.8%) and PD in 14 pts (45.2%). Median PFS is 4.2 months (range 3.3-5.7).

Conclusions: The registry was started to evaluate the safety and efficacy of TEM in pts with rez/refr MCL in the routine clinical practice. In this here included patient collective with 45.7% high-risk pts, TEM shows a predictable, manageable tolerability profile. Efficacy remains comparable with phase III data.

Disclosure: Gabriele Krekeler: Employment or Leadership Position: employee of Pfizer Pharma GmbH, Germany. Daniel Kalanovic: Employment or Leadership Position: employee of Pfizer Pharma GmbH, Germany.

P227 - Preliminary clinical experience on the efficacy and feasibility of a new combination regimen consisting of Pixantrone, Etoposide, and Bendamustine with or without the addition of Rituximab in patients with relapsed/refractory aggressive non-Hodgkin lymphomas

Panny M.1, d'Amore F.2, Nösslinger T.1, Jørgensen J.2, Silkjaer T.2, Leppä S.3, Zintl P.4, Theocharous P.4, Keil F.1

1Hanusch Krankenhaus, Abteilung für Hämatologie und Onkologie, Wien, Austria, 2University Hospital Aarhus, Department of Hematology and Nuclear Medicine, Aarhus, Denmark, 3University Central Hospital Cancer Center Helsinki, Helsinki, Finland, 4CTI Life Sciences Ltd., London, United Kingdom

Introduction: With currently available therapies, relapsed/refractory aggressive NHL after high-dose therapy or, in not transplant-eligible patients, after first-line chemotherapy represents an unmet clinical need.

Methods: evaluating a combination chemotherapy based on pixantrone (Pix), a novel aza-anthracenedione approved by EMA in multiply relapsed/refractory aggressive NHL patients. Etoposide+bendamustine were selected based on prior combination with Pix, and efficacy in salvage regimens in relapsed/refractory aggressive NHL. Schedule: Pixantrone50 mg/m2 i.v. day1+8, Etoposide100 mg i.v. day1, Bendamustine90 mg i.v. day1 ± Rituximab375 mg/m2i.v. day1 (PREBEN/PEBEN). Each cycle was given at 3-weekly intervals for a maximum of 6 cycles. All patients were assessed for chemosensitivity with PET/CT after cycle 1 or 2. G-CSF support was administered according to guidelines.

Results: 10 patients with relapsed/refractory aggressive NHL were treated according to the PREBEN/PEBEN-schedule and were evaluable in terms of presentation, feasibility and response. The histological diagnoses were distributed as follows: relapsed/refractory DLBCL (N = 6), transformed follicular lymphoma (tFL;N = 2), post-transplant lymphoproliferative disease of DLBCL type (PTLD-DLBCL;N = 1), peripheral T-cell lymphoma (PTCL-NOS;N = 1; without rituximab). Based on PET/CT assessment performed already after 1course of treatment, the PREBEN/PEBEN regimen showed metabolic CR in 4/10patients (DLBCL;N = 4) and good PR in 3patients (DLBCL;N = 1,tFL N = 1, PTCL-NOS = 1) corresponding to ORR = 70% (CR: 40%; PR: 30%). No response was seen in 3/10 pts (DLBCL N = 1, PTLD-DLBCL = 1, tFL = 1) (PD: 30%). In both CR and PR patients, a marked reduction of the lesions was already detectable after the 1course of treatment. RD was in the range 5-14+months. The regimen was feasible and most patients received it as out-patients. The most common grade 3-4toxicity was of hematological type. Grade3-4 infections were seen in 40% of these heavily pretreated patient, but they were manageable and could be successfully treated.

Conclusion: The PREBEN/PEBEN schedule is feasible and profound responses early in the course of therapy are not uncommon. A phase 1/ phase 2(extension)study is in preparation.

Disclosure: Michael Panny: No conflict of interest disclosed. Felix Keil: Financing of Scientific Research: CTI, Roche.

P228 - Burkitt lymphoma treated with a rituximab and methotrexate based regimen (GMALL B-ALL/NHL 2002): A single-center retrospective analysis of efficacy, toxicity and experience with relapsed/refractory disease

Cremer M.1, Schwarzbich M.-A.1, Schöning T.2, Lisenko K.1, Ho A.D.1, Witzens-Harig M.1

1Department of Internal Medicine V, University Hospital of Heidelberg, Heidelberg, Germany, 2Pharmacy, University Hospital of Heidelberg, Heidelberg, Germany

Introduction: Immunochemotherapy with a rituximab and methotrexate based regimen according to the GMALL B-ALL/NHL 2002 protocol has become the standard treatment for Burkitt lymphoma in Germany. Treatment consists of six 6-day cycles with rituximab, high-dose methotrexate, high-dose cytosine arabinoside, cyclophosphamide, etoposide, ifosphamide, corticosteroids and intrathekal chemotherapy, with dose reductions for patients over 55 years of age. Two additional doses of rituximab are administered as maintenance. While this treatment is curative in the majority of patients, there is a paucity of data concerning treatment options for patients with relapsed/refractory disease after treatment with this protocol.

Methods: We retrospectively analyzed data from 38 patients who were treated for newly diagnosed Burkitt lymphoma according to the GMALL B-ALL/NHL 2002 protocol at our institution between 2003 and 2013. The probabilities for overall survival and progression-free survival were calculated by the Kaplan-Meier method. Toxicities were retrospectively assessed and categorized according to the Common Terminology Criteria v3.0. We individually reviewed each case with r/r disease and present an overview of post-induction therapy and outcome in these patients.

Results: Of the 38 patients included in the analysis, 36 received the full protocol as planned. The response rate (CR/PR) was 97,4%. OS at 1 and 3 years was 83,9% and 77,8% respectively, PFS at 1 and 3 years was 78,9% and 76,1% respectively. Toxicities were mainly hematologic, infectious and gastrointestinal. The nine patients who relapsed after induction therapy were treated with individual approaches adapted according to comorbidities, age, duration of remission, and site of relapse. Three patients achieved a second remission (CR/PR). Eight out of nine patients with r/r disease died, seven from progressive disease and one from complications of allogeneic hematopoietic stem cell transplantation, while one patient is alive but suffers from a third relapse at time of writing.

Conclusions: Our findings concerning response rate, OS, PFS and toxicity are consistent with reports from the literature, confirming that the GMALL B-ALL/NHL 2002 protocol is efficient and feasible for treatment of Burkitt lymphoma. The analysis of patients with r/r disease shows the lack of effective salvage strategies and the dismal prognosis of this subgroup, emphasizing the need for improved upfront treatment strategies in this setting.

Disclosure: No conflict of interest disclosed.

P229 - Treatment of high risk aggressive B cell lymphomas with DA EPOCH R- a single center retrospective analysis

Panny M.1, Keil F.1, Möstl M.1, Kornberger T.1, Menschel E.1, Simanek R.1, Nösslinger T.1

1Hanusch Krankenhaus, Abteilung für Hämatologie und Onkologie, Wien, Austria

Introduction: Promising results in patients suffering from high risk DLBCL, Burkitt`s lymphoma (BL), mediastinal gray-zone lymphoma (MGZL) and primary mediastinal B cell lymphoma (PMBCL) treated with DA-EPOCH R have been reported.

In our center high risk DLBCL - defined as double-hit/double hit score 2 or high risk IPI - BL, MGZL and PMCL are treated with DA EPOCH R.

Methods: Retrospective analysis of toxicity and efficacy in DA EPOCH R treated patients. So far 11 previously untreated patients - 8 male, 3 female- with a median age of 60 a (32-75 a) have been treated with a total of 48 cycles of DA EPOCH R: 7 DLBCL, 1 MGZL, 1 PMBCL, 2BL.

Results: Grade III/IV ANC occurred in 71% of all cycles, thrombocytopenia III/IV° in 17%, grade III anemia in 4% of 48 cycles. Dose escalation was possible in 10 cycles (21%) - in none of patients aged > 65a dose escalation was possible. Due to peripheral sensory neuropathy II-III° in 6 patients Vincristine had to be dose reduced in 31% of all cycles. Other CTCAE grade III non-hematopoietic toxicities were: peripheral motor neuropathy, mucositis, colitis, hyperglycemia - each in 1 patient. So far 7 patients finished treatment: 3 in CR, 1 in PR, 1 relapsed early, 1 had treatment failure, 1 patient had to be switched to a less toxic regimen due to repeated febrile neutropenia.

Conclusion: Although limited data DA EPOCH R seems to be a feasible treatment with acceptable toxicity. Only in young rather fit patients dose escalation seems to be possible.

Disclosure: Michael Panny: No conflict of interest disclosed. Thomas Nösslinger: Financing of Scientific Research: Roche, Gilead, Janssen.

P230 - Diffuse large B-cell lymphoma in a patient with acute renal failure

Übner M.1, Spriewald B.1, Mackensen A.1

1Universitätsklinikum Erlangen, Medizinische Klinik 5 - Hämatologie und Internistische Onkologie, Erlangen, Germany

Diffuse large B-cell lymphoma (DLBCL) is the most frequent non-Hodgkin lymphoma which shows extranodal involvement in 30% and involvement of the genitourinary tract in about 2-3% of all cases which might be associated with a worse prognosis compared to nodal lymphoma. Manifestation in kidney is extraordinary and the limited reports indicate a poor prognosis.

We report about a 59 year old female patient who presented with acute renal failure. Except nausea and fatigue the patient complained of no further symptoms. Arterial hypertension, hyperthyroidism and a Struma were already known. Creatinine increased to 3.11 mg/dl.

Since no cause for renal failure was found, a biopsy of the left kidney was performed leading to diagnosis of DLBCL. A FDG PET/CT showed enrichment of the tracer in both kidneys, skeletal system and retroperitoneal lymph nodes, which were almost normally configured. Bone marrow was free of lymphoma.

Renal function and initially elevated LDH have improved after the first application of R-CHOP. Therapy was continued with five cycles R-CHOEP and two cycles HD-MTX. The clinical response was excellent and currently the patient is awaiting re-stagning.

Disclosure: No conflict of interest disclosed.


MDS / Stammzellen

P231 - The erythroid regulator Erythroferrone (ERFE) is differentially regulated in CD71+ erythroprogenitor cells of patients with myelodysplastic syndromes (MDS) and is associated with prognosis

Mossner M.1, Stöhr A.1, Nolte F.2, Jann J.C.1, Fey S.1, Nowak V.1, Obländer J.1, Pressler J.1, Baldus C.D.3, Schulze T.J.4, Neumann M.3, Hofmann W.-K.1, Nowak D.1

1Medizinische Fakultät Mannheim der Universität Heidelberg, Mannheim, Germany, 2St. Hedwig-Krankenhaus, Berlin, Germany, 3Charité Campus Benjamin Franklin, Berlin, Germany, 4Institut für Transfusionsmedizin, Mannheim, Germany

Introduction: Myelodysplastic Syndromes (MDS) are characterized by disturbed iron homeostasis. Recently, Erythroferrone (ERFE) was discovered as a novel regulator of iron homeostasis, which is selectively produced by bone marrow (BM) erythroprogenitor cells during hematopoietic stress and EPO stimulation (Kautz et al., Nature Genetics 2014). We therefore sought to examine the role of ERFE expression in CD71+ erythroprogenitor cells derived from patients with MDS and secondary acute myeloid leukemia (sAML).

Methods: CD71+ erythroprogenitor cells were immunomagnetically isolated from mononuclear BM cells of patients suffering from MDS (n = 86, IPSS-low/int-1-risk n = 69, IPSS-int-2/high-risk n = 17), sAML (n = 18) and age-matched healthy donors (n = 17). In addition to CD71+ cells, CD34+, CD61+, CD15+ selected BM as well as CD3+ selected peripheral blood (PB) cells were collected from three MDS patients as well as two healthy young and two healthy old donors. ERFE expression was quantified by quantitative real time PCR. Patient follow up (FU) data was available for n = 55 MDS and n = 13 sAML samples.

Results: ERFE expression analysis in the above described hematopoietic cellular subfractions revealed almost exclusive expression of ERFE in CD71+ erythroprogenitor cells. ERFE expression profiles acquired in CD71 positive cells revealed a highly significant mean overexpression in MDS patients: IPSS-low/int-1-risk (fold change (FC)=4.1, p < 0.0001), IPSS-int-2/high-risk (FC = 4.6, p = 0.0003) and sAML (FC = 6.5, p < 0.0001) relative to age-matched healthy controls. However, a distinct fraction of patients displayed ERFE expression levels similar or even lower than those measured in healthy donors. Univariate analysis of these opposed groups revealed that low abundance of CD71+ ERFE transcripts was significantly associated with inferior overall survival (OS) in MDS patients (median survival 1.7 years vs. not reached, p = 0.0066) and also sAML (median survival 0.1 vs. 0.8 years, p = 0.031).

Conclusion: The current observation of significantly inferior survival probability for MDS and sAML patients with low ERFE expression levels indicates a potentially important biologic and clinical relevance of this novel regulatory gene in the pathogenesis of MDS. Consequently, regulation of aberrant levels of the erythroid hormone ERFE in MDS erythroprogenitor cells could provide a promising target for novel therapeutic avenues that mechanistically address dysfunctional erythropoiesis in MDS.

Disclosure: No conflict of interest disclosed.

P232 - Telomere length is significantly shortened in patients with aplastic anemia compared to hypoplastic myelodysplastic syndrome

Bouillon A.S.1, Ferreira M.S.1, Panse J.1, Reinecke P.2, Schemenau J.3, Haas R.3, Brümmendorf T.H.1, Germing U.3, Beier F.1

1Uniklinik Aachen, Klinik für Hämatologie, Onkologie and Stammzelltransplantation, Aachen, Germany, 2Institut für Pathologie, Uniklinik, Düsseldorf, Germany, 3Klinik für Hämatologie, Onkologie und Klinische Immunologie, Universitätsklinikum, Düsseldorf, Germany

Introduction: The majority of cases of acquired aplastic anemia (AA) is characterized by an autoimmune attack against the hematopoietic stem cell compartment leading to pancytopenia and bone marrow (BM) failure. Thus, immunosuppressive therapy (IST) represents the standard treatment for AA. Hypoplastic myelodysplastic syndrome (hMDS) frequently mimics the clinical findings of AA and proper clinical discrimination of hMDS from AA sometimes remains difficult. The underlying pathogenesis of hMDS is not fully understood. However, based on the close clinical relationship of AA and hMDS, an immune-mediated mechanism similar to AA is propagated. Supporting this hypothesis, hMDS can respond at least partially to IST and on the other hand, AA can clonally evolve to hMDS. Telomere length (TL) shortens with every cell division and reflects the replicative potential of somatic cells. In contrast to AA, the role of TL for disease pathogenesis in hMDS remains unclear. In this study we aimed to retrospectively investigate TL as possible marker to distinguish hMDS from AA.

Material and methods: TL measurement was carried out using confocal Q-FISH protocol as published previously. TL was retrospectively analyzed from BM biopsies at diagnosis of 12 patients with hMDS and 15 patients with AA treated in the University Hospital of Düsseldorf. TL analysis was performed in single-blinded fashion. Mean age was 45.2 years in AA patients and 65.2 years in patients with hMDS. 28 patients (range 18-80 years) with diagnosed M. Hodgkin without BM affection were used as controls for linear regression and calculation of age-adapted TL difference.

Results: First, we analyzed whether TL in AA and hMDS still correlates with age. In contrast to controls (R2 = 0.16, p = 0.03), TL in AA (R2 = 0.07, p = 0.32) and hMDS (R2 = 0.17, p = 0.15) was not significantly correlated with age. Using the controls to adjust for age, age-adapted TL difference was significantly shortened both in patients with AA (median: -2.96 kb, range -4.21 to 0.26, p = 0.001) and patients with hMDS (median: -2.26, range -3.85 to -0.64, p = 0.005). In direct comparison, telomere shortening was more accelerated in patients with AA as compared to hMDS (p = 0.048).

Conclusion: We provide first data on TL in patients with hMDS using confocal Q-FISH. We observe that age-adapted TL is significantly shorter in patients with AA compared to hMDS. Further data are needed to evaluate the role of TL in the evolution of AA and hMDS.

Disclosure: No conflict of interest disclosed.

P233 - Intensive chemotherapy and long term outcome in patients with myelodysplastic syndromes

Zadrozny N.1, Schuler E.1, Strupp C.1, Hildebrandt B.2, Kündgen A.1, Kondakci M.1, Haas R.1, Kobbe G.1, Germing U.1

1Universitätsklinikum Düsseldorf, Klinik für Hämatologie, Onkologie und Klinische Immunologie, Düsseldorf, Germany, 2Universitätsklinikum Düsseldorf, Klinik für Humangenetik und Anthropologie, Düsseldorf, Germany

Introduction: Patients (pts) with high risk MDS can be treated with intensive chemotherapy (IC). The aim of our study was to identify patients´ characteristics that may have relevance for the probability of complete remission or survival after IC.

Methods: We retrospectively analysed 299 pts diagnosed between 1988 and 2014 at our department, and treated with IC and compared this cohort with 1475 pts with more than 4% medullary blasts (mb), not treated with IC or allogeneic stem cell transplantation (SCT).

Results: Median age at IC was 59 (18-79) years. Treatment schedules were ICE 27%, TAD 26%, Idarubicine and Cytarabine (23%) and others (24%). WHO types at IC were: RAEB I 4.8%, RAEB II 17.5%, CMML II 0.7%, AML (20-29% mb) 24.1% and 52.9% already progressed into AML (>30% mb) (8 pts missing information). 32% patients received IC at the initial diagnosis, 68% after a progress to either higher risk MDS or AML. 67% of all pts with known remission status (n = 240) achieved complete remission (CR), 17% partial remission (PR), and 16% did not respond. 82% of pts induced at first diagnosis reached CR, whereas only 60% of pts induced at progression reached CR (p = 0.001). Age above 60 years and presence of auer rods were associated with lower CR rates (73 vs 59%, p = 0.02; 92% vs 63%, p < 0.001). Pts who received IC showed a survival benefit in comparison with the control group (14 vs 23ms, p < 0.001). 33% of pts received a consolidation chemotherapy and lived significantly longer than those without (32 vs 22ms, p = 0.001). 68 (23%) pts received allogeneic SCT 5 ms (0-99) after IC. Pts lived significantly longer after allogeneic SCT (64 vs 21ms, p < 0.001). After censoring pts who received SCT at time of SCT survival in the induction group was still better than in the control group (14 vs 21ms p < 0.001). In multivariate analysis, age >60 years and high/very high risk karyotypes according to IPSS-R were the most important parameters associated with survival. When ignoring karyotypes auer rods, achievement of CR, as well as age >60 were most important.

Discussion: Induction chemotherapy prolongs survival in higher risk MDS pts, with approximately 20% long term survivors (>60ms). Pts benefit from consolidation therapy and CR rates are higher in pts treated at first diagnoses. Poor risk karyotype, higher age, and non-remission are associated with poor prognosis. Whenever possible pts should receive allogenic transplantation (50% of pts are still alive after 60ms).

Disclosure: No conflict of interest disclosed.

P234 - Establishment of a novel “short tandem repeat” based multiplex-PCR assay for accurate del(5q) quantification using genomic DNA

Jann J.C.1, Mossner M.1, Nolte F.2, Fey S.1, Nowak V.1, Obländer J.1, Pressler J.1, Fabarius A.1, Haferlach C.3, Hofmann W.-K.1, Nowak D.1

1Medizinische Fakultät Mannheim der Universität Heidelberg, Mannheim, Germany, 2St. Hedwig-Krankenhaus, Berlin, Germany, 3MLL Münchner Leukämie Labor, München, Germany

Introduction: Cytogenetic lesions play a key role in diagnosis and prognostication of hematologic malignancies. Such aberrations can be readily assessed via metaphase cytogenetics (MC), FISH and microarrays. However, these methods often depend on the availability of viable cells, large amounts of DNA or are very expensive. For accurate quantification of chromosomal deletions in minute amounts of DNA specimen we present a novel PCR-based assay for interrogation of del(5q) aberrations in myelodysplastic syndromes (MDS) that is based on measuring allelic loss at heterozygous short tandem repeat (STR) loci.

Methods: Genomic DNA was extracted from bone marrow (BM) cells from MDS patients with del(5q). 12 fluorochrome-labelled PCR amplicons covering STR loci between chromosome 5q21 and 5q31 were generated from 10 ng DNA in a single multiplex-PCR reaction. Amplicons were subjected to capillary electrophoresis for allele ratio quantification of heterozygous STR loci. Deviations from equal allelic ratios in heterozygous STR markers in the tumour samples were used to calculate the fraction of cells carrying del(5q).

Results: Using our novel assay for quantification of del(5q) burden, we identified an average of 8 heterozygous 5q STR marker per individual in a total of n = 696 samples from n = 67 MDS patients and n = 136 healthy donors. Our analysis revealed strong inter-marker correlations (average standard deviation (SD) = 2.3%) and high reproducibility in duplicate measurements of n = 328 samples (SD = 0.86%). Moreover, highly concordant results were observed for serial dilution series (r² = 0.96) and paired analyses between interphase-FISH and our assay (r² = 0.94). Finally, screening of patient BM samples during Lenalidomide treatment reliably confirmed molecular remission as confirmed by MC.

Conclusion: With the establishment of a highly multiplexed PCR assay for interrogation of STRs in deleted chromosomal regions we provide a highly adaptable technique for quantification of genomic aberrations. As exemplarily shown for del(5q), our assay provides strong reproducibility and concordance with established techniques. Requiring only minute amounts of DNA, this tool is highly suitable for copy number quantification when only residual archival DNA, e.g fragmented DNA from formalin-fixed samples, and no cells for FISH analysis are available. In summary, our newly developed assay provides a sensitive tool for quantitative copy number analysis for any large scale chromosomal deletion.

Disclosure: No conflict of interest disclosed.

P235 - Piaza: Non-interventional study on efficacy and safety of azacitidine (Vidaza®) in patients with myelodysplastic syndromes (MDS, Int-2 or high risk), AML (WHO 20-30% blasts), or CMML (10-29% bone marrow blasts without myeloproliferative disorder)

Wehmeyer J.1, Zaiss M.2, Losem C.3, Schmitz S.4, Neidig C.5, Teichmann B.5, Harde J.5, Trarbach T.5

1Hämatologisch-onkologische Gemeinschaftspraxis, Münster, Germany, 2Praxis für interdisziplinäre Onkologie & Hämatologie, Freiburg, Germany, 3Praxis für Onkologie u. Hämatologie, Neuss, Germany, 4Gemeinschaftspraxis für Hämatologie und Onkologie, Köln, Germany, 5iOMEDICO AG, Freiburg, Germany

Introduction: Myelodysplastic syndromes (MDS) represent one of the most frequent haematologic diseases of the elderly. Treatment with hypomethylating agents, such as azacitidine (AZA), has become the standard therapy for higher-risk MDS patients (pts) who are ineligible for stem cell transplantation. AZA is associated with improved outcomes in MDS, including delayed AML transformation and prolonged survival in pts with higher-risk MDS and is also approved for CMML and AML pts with 20-30% blasts.

Methods: This prospective non-interventional study collected data on the routine treatment of MDS (Int-2 or high risk), AML or CMML with AZA. Treatment with AZA was documented for one year followed by a one year follow-up phase. Data on demographics, tumour characteristics, treatment duration and reasons for treatment discontinuation was collected. The main endpoints are PFS, ORR, and safety parameters.

Results: A total of 149 pts were enrolled in 31 German sites. Pts were diagnosed with primary MDS (59.7%), AML/CMML (37.6%) or secondary MDS (2.7%). Mean age was 75.5 (SD: ±7.4) years. At inclusion most pts had an ECOG PS of 0 or 1 (73.8%). Median PFS was 10.9 months (95% CI = 8.7-12.8). Parameters found to be relevant to PFS in a multivariate Cox-model were ECOG PS at baseline (P = 0.0018; HR = 2.23) and previous RBC transfusion (P = 0.0362; HR = 1.57). Pts with ECOG 0 had a longer PFS than those with ECOG 1 (18.4 vs. 9.8 months). Further, pts without the requirement for RBC transfusion prior to treatment appeared to have a longer PFS than those who did receive transfusions (12.5 vs. 8.6 months). Median OS was 14.1 months (95% CI 12.3-17.2). The 2-year survival rate was 28.9%. ORR was 41.6% (CR: 10.7%; PR: 30.9% vs. 28.2% SD; 6% PD (remaining 24.2% missing data)). Median TTD was 6.8 months (95% CI 5.4-7.9). Most common AEs with suspected relation to AZA were gastrointestinal disorders (27.5%), such as nausea (20.8%) and vomiting (9.4%), blood and lymphatic disorders (23.5%) such as leukopenia (12.8%), anaemia (10.1%), thrombocytopenia (10.1%).

Conclusions: AZA is an effective and well tolerated therapeutic option for pts with higher-risk MDS. ECOG PS and previous RBC transfusion were found to be a predictor of PFS. The response rate was higher than reported in pivotal trials. Median OS was 14.1 months, which is lower than reported in study AZA 001, the registration trial for AZA. This may be due to the fact that pts were 5 years older than in study AZA 001.

Disclosure: Jürgen Wehmeyer: Advisory Role: AMGEN, Bayer, BMS, Celgene, Roche; Stock Ownership: iOMEDICO AG; Financing of Scientific Research: AMGEN, Bayer, BMS, Celgene, Roche. Tanja Trarbach: Employment or Leadership Position: iOMEDICO AG.

P236 - Identification of somatic mutations by next-generation sequencing in secondary myelodysplastic syndromes of multiple myeloma patients

Murga Penas E.M.1, Pomplun C.1, Paul U.1, Nagel I.1, Becher C.1, Gramatzki M.2, Günther A.2, Siebert R.1

1Institute of Human Genetics, University Hospital Schleswig-Holstein Campus Kiel/Christian-Albrechts University Kiel, Kiel, Germany, 2University Hospital Schleswig-Holstein Campus Kiel/Christian-Albrechts University Kiel, Division of Stem Cell Transplantation and Immunotherapy, 2nd Medical Department, Kiel, Germany

Introduction: Life expectancy of patients with multiple myeloma (MM) has been improved with the advent of novel treatments based on high-dosage chemotherapy combined with autologous stem cell transplantation and novel agents (immunomodulatory drugs and proteasome inhibitors). However, improvement in survival has led to increasing risk of secondary malignancies in MM especially for the development of myelodysplastic syndrome (secondary sMDS).

Methods: Three cases of MM patients diagnosed in addition with a sMDS were analyzed by karyotyping, fluorescence in situ hybridization (FISH), and next-generation sequencing (NGS). NGS was performed on DNA from CD138-negative cells of each patient using the MiSeq benchtop sequencer and the MiSeq Reporter and Illumina VariantStudio 2.2 for data analyses (Illumina, San Diego, USA). In case 1, a 49-years-old female (stage IIIA), and in case 3, a 55-years-old male (stage IIIA), had been treated with melphalan, cyclophosphamide, bendamustine, and lenalidomide. In case 2 a 60-years-old male (stage IIIA) had been treated with melphalan and cyclophosphamide but never had received an immunomodulatory drug. sMDS was diagnosed 7-, 8-, and 4-years after diagnosis of MM in case 1, 2, and 3, respectively.

Results: FISH analyses on CD138-positive cells at time of diagnosis or relapse revealed a complex karyotype with a t(4;14)(p16:q32)/MMSET/FGFR3-IGH in case 1 (non-hyperdiploid MM) and gains of chromosomes 5, 11, and 21 in cases 2 and 3 (hyperdiploid MM). At the time of sMDS diagnosis, standard cytogenetic analyses revealed a monosomy 7 (-7) in all cases. In the two hyperdiploid MM, -7 was part of complex karyotypes with additionally -5/5q- and -12. By NGS, case 1 showed a heterozygous c.408T>A mutation in the RUNX1 gene. Cases 2 and 3 showed both heterozygous mutations in splice sites of the TP53 gene, c.783-2A>G and c.375+2T>G, respectively. All detected mutations by NGS could be validated using Sanger sequencing.

Conclusions: Here, evidence is provided that somatic mutations of RUNX1 and TP53, which are associated with advanced disease, occur recurrently in sMDS of MM patients. Moreover, we show -7 and -5/5q- to be recurrent chromosomal changes in sMDS in both hyperdiploid and non-hyperdiploid MM. Additional studies of a larger group of patients will be required to determine the incidence of these mutations in sMDS occurring in MM patients.

This work was supported by the Medical Faculty of the Christian-Albrechts- University.

Disclosure: No conflict of interest disclosed.

P237 - Is this scleroderma? Skin lesions in patient with profound cytopenia

Ihne S.M.1, Karsten A.1, Schmidt M.1, Kerstan A.2, Einsele H.1, Grigoleit G.U.1, Knop S.1

1Universitätsklinikum Würzburg, Medizinische Klinik II, Hämatologie, Würzburg, Germany, 2Universitätsklinikum Würzburg, Dermatologie, Würzburg, Germany

Accompanying autoimmune phenomena are observed in about 10% of patients with myelodysplastic syndrome (MDS). Manifestations may be unspecific, thereby impeding the diagnosis of the underlying disease. The pathomechanism behind is still rarely understood.

Here we report a case of a 51-year-old man with severe fatigue, asthenia, weight loss (14 kg/10 months) and generalized scleroderma which started from the distal limbs and resulted in progressive limitation of motility. Noteworthy, those signs suggestive for connective tissue disease had been preceded by a seemingly idiopathic deep vein thrombosis. Concomitantly, we observed pancytopenia: leukocytes, 1.9 x 103/ul; neutrophils, 650/ul; eosinophils, 100/ul; hb, 10.2 g/dl; MCV, 99,3 fl; MCH, 34,3 pg; MCHC, 34,6 g/dl; platelets, 9 x 103/ul). Vitamin B12 deficiency was substituted without improvement of the haemogram, the level of folic acid was normal. Spleen was moderately enlarged.

Skin biopsies showed generalized deep morphea with scattered figures of hemophagocytosis.

Bone marrow aspirates revealed reactive changes without manifest signs of dysplasia or lymphoma infiltration. Erythropoiesis was increased with left shift, but megakaryocytes were nearly absent. Viral infections (Parvovirus B19, CMV, VZV, EBV, HHV6, HSV1/2, adenovirus) could be excluded. Astonishingly, augmented phagocytes with hemophagocytosis figures were found suspect for secondary hemophagocytic lymphohistiocytosis (HLH/MAS). According to the criteria for HLH, only 4 of 8 were fulfilled and mutation analysis of perforin showed a wild type gene, making an HLH unlikely. Because of rapidly progressing skin lesions, we decided on an immunosuppression, but neither high-dose steroids nor application of cyclophosphamide improved pancytopenia or generalized morphea. In view of increasing need of transfusion and progressively increased MCV another bone marrow biopsy was performed, then showing unequivocal signs of dysplasia of erythropoiesis and granulopoiesis. As synopsis of our findings, we interpreted the deep morphea as a paraneoplastic event preceding overt MDS. Because of the young age, increasing transfusion frequency and rapid progressive scleroderma with resulting limitations of respiratory movements, the patient underwent allogeneic stem cell transplantation. In the course of treatment skin lesions improved rapidly. Restaging 30 days after allogeneic SCT showed reactive changes without signs of dysplasia.

Disclosure: No conflict of interest disclosed.

P238 - Optic disc swelling and preretinal Hemorrhage in Patient with Myeloblastic Syndrome

Kochkorov A.1, Bauer G.1, Killer H.1

1Kantonsspital Aarau AG, Augenklinik, Aarau, Switzerland

Background: The term Myelodysplastic Syndrome (MDS) is a set of mostly acquired, rare congenital, clonal diseases of the hematopoietic stem cells with heterogeneous clinical presentation.

History and Signs: A 16-year-old patient was presented due to a gray spot in front of his left eye. Otherwise, he had no complaints, especially no dizziness and no fever. There was an optic disc swelling of both eyes, a preretinal haemorrhage in front of the macula with reduced visual acuity (0.2) of the left eye and pancytopenia.

Therapy and Outcome: The patient was assigned into the children´s hospital for further clarification. Until the diagnosis was confirmed he got erythrocyte- and platelet- transfusions. Under this therapy his visual acuity on the left eye begun to improve and the funduscopic abnormalities started to reduce. After the refractory cytopenia type of MDS was confirmed, a stem cell transplant was performed. From that time on and for more than 1 year, he has normal visual acuity and completely normal funduscopic findings.

Conclusions: For unexplained retinal hemorrhages and optic disc swelling an underlying hematological disease should be excluded. Usually in MDS with ocular involvement is only the treatment of the underlying disease sufficient.

Disclosure: No conflict of interest disclosed.

P239 - Additional plerixafor to granulocyte colony-stimulating factors for haematopoietic stem cell mobilisation for malignant lymphoma or multiple myeloma patients: A systematic review with meta-analysis

Hartmann T.1, Hübel K.2, Monsef I.1, Engert A.2, Skoetz N.1

1Uniklinik Köln, Evidenz-basierte Onkologie, Klinik I für Innere Medizin, Köln, Germany, 2Uniklinik Köln, Klinik I für Innere Medizin, Köln, Germany

Introduction: Autologous stem cell transplantation is widely used to restore functioning bone marrow in malignant lymphoma or multiple myeloma patients after myeloablative chemotherapy. Results of clinical trials indicate that plerixafor plus G-CSF leads to an increased mobilisation and release of CD34+ cells, facilitating effective apheresis. To evaluate the efficacy and safety of plerixafor we conducted a systematic review and meta-analysis.

Methods: We searched Cochrane Central Register of Controlled Trials and MEDLINE (1990 to April 2015) for RCTs and included trials evaluating additional plerixafor compared to placebo or no therapy for stem cell mobilisation. Two review authors independently screened the results of the search strategies extracted data, assessed quality and analysed data. Final interpretation was done together with an experienced clinician.

Results: We identified four trials fitting the inclusion criteria. However, one was closed prematurely and did not report results, another was not published at all (completed in 2009). The remaining two trials evaluated 600 participants with multiple myeloma or Non-Hodgkin lymphoma. No evidence for differences between plerixafor and control group regarding mortality at 12 months (RR 1.00; 95% CI 0.59 to 1.69; P = 1.00) and adverse events in study period one emerged in the meta-analysis.

The meta-analysis showed a significant advantage for those patients randomised to plerixafor for achieving a targeted stem cell number in four or less apheresis days (RR 2.42, 95% CI 1.98 to 2.96; P < 0.00001). As there is high heterogeneity between studies for the number of transplanted patients, we did not meta-analyse these data. In AMD3100-3102MM 95.9% in the plerixafor arm and 88.3% in the placebo arm underwent transplantation (RR 1.09, 95% CI 1.02 to 1.16), in AMD3100-3101NHL 90% versus 55.4% (RR 1.62, 95% CI 1.39 to 1.89).

None of the trials reported quality of life.

Conclusion: The two trials, both conducted by Genzyme Corporation, the manufacturer of plerixafor, were published several times. Two other trials are not published at all. Therefore the meta-analysis must be interpreted with caution as there is a high risk for publication bias. There are hints for an increased stem cell collection in a shorter time, without affecting overall survival or adverse events. For a better comparability of GCS-F plus plerixafor and GCS-F alone, all conducted RCTs must be published.

Disclosure: No conflict of interest disclosed.

P240 - Quantification of the time and effort associated with autologous peripheral blood stem cell mobilisation: A European perspective

Hübel K.1, Ostermann H.2, Noppeney R.3, Glaß B.4, Berger K.2, Reitan J.5, Mohty M.6

1Universität Köln, Klinik I für Innere Medizin, Köln, Germany, 2Universitätsklinikum, München, Germany, 3Universitätsklinik, Essen, Germany, 4Asklepios Klinik St. Georg, Hamburg, Germany, 5RJM Group, Crown Point, United States, 6Saint-Antoine Hospital, Paris, France

Introduction: Plerixafor is indicated in combination with G-CSF to enhance mobilisation of hematopoietic stem cells to the peripheral blood (PBSC) for collection. The aim of this non-interventional study was to assess resource utilization, including time, effort and costs to the hospital, associated with PBSC mobilisation and apheresis.

Patients and methods: The study population includes patients with a primary diagnosis of NHL who underwent PBSC mobilisation at European centres. Part I of the study, currently ongoing, is a retrospective medical record review of 200 NHL patients from 9 centers across France and Germany. Selected patients are evenly divided between two eras: 1) prior to approval of plerixafor (until 1 June, 2009), pre-P era, and 2) after approval of plerixafor (1 July, 2010 and onwards), P era. Outcome measures include number of visits for administration of mobilising agents; duration (days) of administration of mobilising agents; agents used as mobilising agents; adverse events (AEs); number and hours of apheresis sessions; attainment of CD34+ target (yes, no) and days until CD 34+ target level was met. Part II of the analysis is an ongoing prospective time and motion evaluation of apheresis performed at each center. Time-motion assessments will be obtained retrospectively (Part I) and prospectively (Part II). Costs will be evaluated and quantified through micro-costing group interviews with local hospital administration.The primary study end point is difference in mean time/effort to perform apheresis (including apheresis related AEs, if any) and total costs associated with mobilisation to the hospital between patients in the “Pre-P” versus “P eras”.

Results: At time of abstract submission, data collection is ongoing at all centers. Final results from four German centers will be presented. Comparisons will be made between the German centers and the overall study results. It is hypothesized that the key findings of this study will demonstrate the favorable impact of novel Interventions on the number of apheresis procedures required to reach a target PBSC, and failure rate of mobilisation, thus translating into reduced total transplant costs without increasing toxicity.

Conclusion: The financial implications for transplant centers could be significant and may lead to further studies aiming to optimize staff time and resource utilization related to apheresis in real-world practice.

Disclosure: Kai Hübel: Advisory Role: Beratung Sanofi; Financing of Scientific Research: Vortragshonorar Sanofi; Expert Testimony: Sanofi; Other Financial Relationships: Reisekostenerstattung Sanofi. Mohamad Mohty: Advisory Role: Beratung Sanofi; Financing of Scientific Research: Vortragshonorar Sanofi; Expert Testimony: Sanofi; Other Financial Relationships: Reisekostenerstattung Sanofi.

P241 - Addition of plerixafor to stem cell mobilization regimens in autologous donors with multiple myeloma and lymphoma can increase the efficacy of stem cell harvesting in poor mobilizers

Stümpel J.-P.1, Jentzsch M.1, Ruschpler E.1, Leiblein S.1, Franke G.-N.1, Schwind S.1, Jäkel N.1, Wang S.-Y.1, Heyn S.1, Pönisch W.1, Niederwieser D.1, Vucinic V.1

1Universitätsklinikum Leipzig, Hämatologie und Internistische Onkologie, Leipzig, Germany

Introduction: Plerixafor is a potent CXCR4 receptor antagonist that is licensed for stem cell mobilization in autologous stem cell donors with multiple myeloma or lymphoma who show poor mobilization after having been stimulated with recombinant granulocyte colony stimulating factor (r-metHuG-CSF, Filgastrim).

Patients and methods: We performed 27 stem cell collections in 21 poor mobilizing autologous donors with multiple myeloma (n = 13) or malignant lymphoma (n = 8) adding plerixafor to the stimulation regimen with recombinant G-CSF from January 2011 till April 2014. At stem cell collection, our patients (9 males, 12 females) had a median age of 61.5 years (y) (range 44-72y). In all cases a large volume leucapheresis (maximal 4 × total blood volume) was performed. Patients received 20 µg/kg body weight (BW) plerixafor after having achieved peripheral reconstitution of white blood cells (WBC, >1 Gpt/l). 6 patients received a second dosage of plerixafor due to insufficient CD34+ cell harvesting.

Results: The median concentration of CD34+ cells in peripheral blood (PB) before and after the first application of plerixafor was 7.8/ µl (range 2.9-14.3 /µl) and 21.2 /µl (range 10.2-54.1 /µl) respectively, resulting in a median 3.3-fold increase (range 1.4-5.3-fold). The median CD 34+ yield for all patients was 3.7 × 10^6/kg BW (range 1.6-12.8 × 10^6/kg BW) with median WBC counts of 857.3 × 10^8 (range 446-2214 ×10^8) in the product. The vitality of the cells after thawing was tested with methan blue and showed excellent in all cases (median 86.9%, range 73.3-92%, standard deviation 4.0%). Until today, 18 patients received the autologous stem cell transplantation. All of them showed good peripheral WBC recovery, in median on day 11 (range 10-15).

Discussion: Our data shows that addition of plerixafor increases the efficacy of stem cell harvesting in poor mobilizers. The produced stem cells fulfilled the required quality control criteria like vitality and building of colony forming units (CFU). We identified a median 3.3 fold increase of CD34+ concentration in peripheral blood after the application of plerixafor in poor mobilizers. Thus, the mobilization outcome of our patients receiving plerixafor shows acceptable efficacy within the expected range.

Disclosure: No conflict of interest disclosed.

P242 - A panel of mass spectrometry based serum protein tests for predicting Graft-versus-Host Disease (GvHD) and its severity

Koldehoff M.1, Roder J.2, Hoffmann A.C.3, Roder H.2

1University of Duisburg-Essen, Department of Bone Marrow Transplantation, Essen, Germany, 2Biodesix, Inc. Steamboat Springs, Colorado, United States, 3University of Duisburg-Essen, Department of Medical Oncology, Essen, Germany

Introduction: Transplants of pluripotent hematopoietic stem cells (PHSC) or bone marrow (BM) are effective therapies against hematological malignancies. However, GvHD remains a major source of morbidity and mortality post-transplant. Identification of patients likely to develop severe forms of acute or chronic GvHD could allow the selection of more aggressive therapeutic regimens for patients assessed to be at high risk.

Methods: Serum samples and clinical data were available from 124 patients (age 18-70) who had received PHSC or BM transplants. Five patients suffered no GvHD, 15 de novo chronic GvHD (cGvHD), 21 acute GvHD (aGvHD) but no cGvHD and 83 both aGvHD and cGvHD. Of patients with aGvHD 51% had grade I disease and of patients with cGvHD 53% had limited disease. Matrix assisted laser desorption/ionization (MALDI) mass spectra were acquired from the samples using the deep MALDI method, allowing a deep probing of the proteome. The spectra were preprocessed and spectral features defined. The integrated intensities of these features were combined with the clinical data using deep learning based machine learning techniques to create classifiers able to stratify patients into groups depending on occurrence and severity of GvHD.

Results: Classifiers could be developed with significant power to predict occurrence and severity of GvHD. The area under the curves (AUCs) obtained for the clinical questions investigated and examples of the sensitivity and specificity achievable are summarized in the table (Occurrence of GvHD).


Conclusions: It is possible to provide information on occurrence and severity of GvHD from mass spectral analysis of post-transplant serum samples. If validated, this panel of tests could provide additional information useful for clinicians choosing treatment regimens for patients following PHSC or BM transplants.

Disclosure: Michael Koldehoff: No conflict of interest disclosed. Heinrich Roder: Employment or Leadership Position: Biodesix, Inc. Steamboat Springs, CO, USA.

P243 - Mobilization and hematopoietic stem cell collection in patients with autoimmune disease

Lisenko K.1, Pavel P.2, Ho A.D.1, Wuchter P.1, Blank N.1

1Universitaetsklinikum Heidelberg, Innere Medizin 5, Heidelberg, Germany, 2Universitaetsklinikum Heidelberg, IKTZ Institut für Klinische Transfusionsmedizin und Zelltherapie, Heidelberg, Germany

Introduction: High-dose chemotherapy followed by autologous hematopoietic stem cell (HSC) transplantation is a promising therapeutic approach in patients with refractory systemic sclerosis (SSc), multiple sclerosis (MS) and other autoimmune diseases (AID). We investigated the feasibility and efficacy of HSC mobilization in AID-patients.

Methods: This is a single center retrospective analysis of HSC mobilization and collection in 33 patients with refractory SSc (n = 15), MS (n = 11) and other AID like vasculitis, connective tissue disease or rheumatoid arthritis (n = 7). HSC mobilization with cyclophosphamide + G-CSF and HSC collection was performed between 1999 and 2015 at our institution. The impact of age, body weight, diagnosis, disease duration, skin sclerosis and previous cyclophosphamide treatment upon HSC collection parameters was investigated. The efficacy of two different mobilization regimens (cyclophosphamide 2 × 2 g/m2 versus 2 × 1 g/m2) was analyzed.

Results: HSC collection was successful in all 33 patients. The median number of collected HSCs was 12.2, 8.0 and 8.2 CD34+ cells/kg ×106in SSc, MS and other AID. Twenty-five of 33 (76%) patients achieved a sufficient collection during one leukapheresis session, while 5 of 33 (15%) required two and 3 of 33 (9%) required three or more leukapheresis sessions. No correlation of the collected HSC number was observed regarding age, body weight, diagnosis, disease duration, skin sclerosis or previous cyclophosphamide treatment. Mobilization chemotherapy with cyclophosphamide 2 × 2 g/m2 (n = 16), 2 × 1 g/m2 (n = 15) or 2 × 0.5 g/m2 (n = 1) resulted in equally high numbers of collected HSCs by leukapheresis on day 13 or 14. One patient collected HSC without mobilization chemotherapy.

Conclusions: We could demonstrate that HSC collection is safe and feasible in patients with AID irrespective of age, body weight, diagnosis, disease duration and previous cycles of therapy. Mobilization chemotherapy with cyclophosphamide 2 × 1 g/m2 or 2 × 2 g/m2 and HSC sampling efficacy is equally effective in those patients.

Disclosure: No conflict of interest disclosed.

P244 - Donor evaluation before syngeneic hematopoietic stem cell transplantation for high-risk B-cell chronic lymphocytic leukemia

Klink A.1, Eigendorff E.1, Schmidt V.1, Dornaus S.1, Hilgendorf I.1, Hochhaus A.1, Sayer H.G.1,2

1Universitätsklinikum Jena, Klinik für Innere Medizin II, Hämatologie und Onkologie, Jena, Germany, 2HELIOS Klinikum Erfurt, 4. Medizinische Klinik, Hämatologie und internistische Onkologie, Hämostaseologie, Erfurt, Germany

Introduction: Allogeneic hematopoietic stem cell transplantation (HSCT) is an established treatment in high-risk B-cell chronic lymphocytic leukemia (B-CLL). Lower relapse rate in lymphoma patients is observed after allogeneic including syngeneic transplantation when compared with autologous transplantation. Few data for syngeneic HSCT in B-CLL show promising results regarding long-term survival. The advantages are the absence of graft rejection and graft versus host disease. Epidemiological data confirmed significant higher disease risk among 1st degree relatives.

Patient and methods: We report on a female patient, who was diagnosed with B-CLL, stage Binet A, in 9/2003. In 2010 the patient required treatment due to progressive leucocytosis and lymphadenopathy. Former treatment with chlorambucil/prednisolone resulted in stable disease parameters. Because of significant disease progression into Binet C with evidence of IGH gene aberration and a deletion (17p)/TP53 mutation in 03/2012 second-line treatment with five courses of Rituximab (R) and fludarabine/cyclophosphamide was initiated. In the presence of poor-prognostic marker and partial remission of CLL the patient was transferred to our center in order to evaluate allogeneic HSCT. The HSCT-comorbidity score revealed 3 points (infection [Lyme disease], asthma bronchiale) for the 64-year-old patient. Syngeneic transplantation was favored instead of matched unrelated HSCT.

Results: Immunocytology of the twin sisters bone marrow excluded a monoclonal cell population since no cytogenetic evidence was found for the presence of malignant hematological disease, particularly B-CLL. The patient received 1.7 × 106 CD34+cells/kg after conditioning with R-BEAM from her identical twin sister in 09/2013. Early transplantation complications were stomatitis °I on day +3 and CMV-reactivation on day +15. Nineteen months after HSCT, the patient is still in complete remission of B-CLL with a Karnofsky index of 90-100%. The donor remains without any signs of a hematological disorder.

Conclusion: Our case shows a favorable outcome for syngeneic HSCT for advanced high-risk B-CLL. To achieve long-term disease control a careful assessment of genetically identical twins regarding leukemic donor cell clone is advised.

Disclosure: No conflict of interest disclosed.


Supportive Therapie

P245 - Efficacy of antibiotic prophylaxis in AML patients treated with low-intensity therapeutic regimens

Bainschab A.1, Quehenberger F.2, Greinix H.T.1, Krause R.3, Wölfler A.1, Sill H.1, Zebisch A.1

1Medical University Graz, Division of Hematology, Graz, Austria, 2Medical University Graz, Institute of Medical Informatics, Statistics and Documentation, Graz, Austria, 3Medical University Graz, Section of Infectious Diseases and Tropical Medicine, Department of Internal Medicine, Graz, Austria

Introduction: Survival of older patients with acute myeloid leukemia (AML) has significantly improved with the advent of low-intensity therapeutic regimens (LITR, comprising decitabine, azacitidine, and low-dose cytarabine). Yet, infectious complications are common during LITR treatment and might hamper the beneficial effect of these drugs. Therefore, we aimed to evaluate the incidence of and predisposing risk factors for infectious complications, as well as the value of antibiotic prophylaxis during LITR treatment of AML.

Methods: We retrospectively analyzed 39 AML patients, who were treated with 215 cycles of LITR at the Medical University Graz. LITR was administered first-line in 29 and after failure of conventional high-dose approaches in ten cases. Infection was defined as any event necessitating appropriate diagnostic measures and anti-infective treatment. Variables tested as risk factors were assessed at the beginning of each LITR cycle and comprised hemoglobin and transfusion dependence, neutrophil and platelet counts, lactate dehydrogenase (LDH), c-reactive protein (CRP), creatinine and glomerular filtration rate, precedent high-dose chemotherapy as well as antibiotic prophylaxis (fluoroquinolones in most cases). Risk factors for the occurrence of infectious complications were assessed by multivariate logistic regression.

Results: Infectious complications occurred in 29/39 patients (74%) resulting in death in six cases (15%). Of the 215 LITR cycles administered, 53 (25%) were complicated by infections. A clinically diagnosed infection was reported in 27/53 cases (51%) with pneumonia being the predominant event. A microbiologically diagnosed infection was reported in 17/53 cases (32%), most frequently gram-negative bacteria. In a multivariate analysis of parameters assessed at the start of each LITR cycle, low hemoglobin (P = 0.009) and transfusion dependence (P = 0.001), as well as increased LDH (P = 0.006) and CRP (P = 0.048) levels independently predicted the occurrence of infection. Most importantly, antibiotic prophylaxis was associated to a decreased rate of infections (P = 0.010).

Conclusions: Low hemoglobin and transfusion dependence, as well as high levels of LDH and CRP are valuable markers to predict infection during LITR treatment in AML. Additionally, antibiotic prophylaxis might help to reduce the incidence of infectious complications. These data suggest evaluation of antibiotic prophylaxis in this setting within a prospective clinical trial.

Disclosure: Antonia Bainschab: No conflict of interest disclosed. Armin Zebisch: Financing of Scientific Research: Celgene.

P246 - Retrospective evaluation of the safety and efficacy of therapy with Vancomycin versus Linezolid in patients with febrile neutropenia following myelosupressive chemotherapy for hematologic malignancies in a real life setting

Hegge N.1, Hahn-Ast C.2, Brägelmann J.2, Schwab K.2, Molitor E.2, Brossart P.2, Wolf D.2, Mayer K.2

1Universitätsklinik, Bonn, Germany, 2Universitätsklinik Bonn, Medizinische Klinik 3, Bonn, Germany

Introduction: In patients with hematological malignancies and febrile neutropenia as a consequence of previous chemotherapy, broad-spectrum antibiotics are considered 1st line treatment. If fever persists and gram-positive infection is suspected Vancomycin (VAN) or Linezolid (LIN) represent potential options for escalation of antibiotic therapy. This single center retrospective analysis compared the efficacy and safety of adding VAN or LIN to an existing broad-spectrum antibiotic therapy.

Methods: All patients with febrile neutropenia and normal renal function, in which VAN or LIN were part of the antibiotic escalation regimen treated in a tertiary care university hospital between 2010-2014 were identified and analyzed retrospectively.

Results: 87 cases were identified (VAN: n = 62, LIN: n = 25). Baseline characteristics (gender, malignancy, comorbidity, age, neutropenia-duration, type of chemotherapy and renal function) were comparable between the groups. Median age was 56 years (46-62). No significant difference was observed in distribution of gram (+), gram (-) and mixed gram (±) pathogens. VAN or LIN was used as second escalation in the antibiotic regimen in more than 70% of patients. Because of persisting fever, antibiotic strategy had to be modified more often in the VAN-group as compared to the LIN-group (VAN 51.6%, LIN 24.0%, p = 0.03). The median days (d) with persisting fever after escalation to VAN or LIN was slightly shorter in the LIN-group (VAN = 3d, LIN = 2d, p = 0.13). 92.2% of all detected gram(+)-pathogens were glycopeptide-sensitive. Regarding toxicities there was no significant difference in renal function (glomerular filtration rate/GFR) during therapy (VAN 78.35 ml/min, LIN 90.20 ml/min, p = 0.8). After escalation to VAN/LIN the LIN-group showed a trend towards shorter median hospital stay (VAN = 16 days (11-23), LIN = 12 (8-18) p = 0.09). In-hospital mortality was not significantly different between the groups (VAN 10.9%, LIN 8.7%, p = 1).

Conclusion: Antibiotic escalation to Linezolid in febrile neutropenia following chemotherapy required significantly less additional antibiotic switches when compared to Vancomycin. Considering the good tolerability (especially the absence of significant renal side effects) and a comparable efficacy in this cohort, Vancomycin still represents a potent and more cost-effective option for suspected gram+ infection in patients with neutropenic fever.

Disclosure: No conflict of interest disclosed.

P247 - A retrospective analysis of the impact of VRE colonization on patients with hematological malignancies

Bodden G.1, Kondakci M.1, MacKenzie C.2, Kolbe-Busch S.2, Schulze-Röbbecke R.2, Germing U.1, Kobbe G.1, Fenk R.1, Schroeder T.1, Haas R.1

1Uniklinik Düsseldorf, Klinik für Hämatologie, Onkologie und klinische Immunologie, Düsseldorf, Germany, 2Uniklinik Düsseldorf, Institut für Medizinische Mikrobiologie und Krankenhaushygiene, Düsseldorf, Germany

Introduction: The spread of vancomycin-resistant enterococci (VRE) strains causing infections may be associated with greater morbidity, mortality and healthcare cost. In this study, we evaluated the rates of VRE in patients with hematological malignancies on the basis of routine screening cultures obtained by rectal swabs.

Methods: We routinely perform VRE screenings since 2009 with rectal swabs aiming to detect all VRE-colonized hematological patients treated in our Department of Hematology and Oncology. For this study, baseline data, rates of VRE infections and outcome data of colonized patients were collected. Analyzing the impact of colonization on patients' clinical outcomes, we matched VRE-colonized patients with AML with AML patients not being found VRE-positive.

Results: Between 2009-2013, a total of 1433 samples were examined, 330 (23,02%) of these were detected as VRE-positive according to 223 hematological patients, 133 (59,64%) male, 90 (40,36%) female, median age was 60 years (range 18-83 years), median length of hospital stay was 25 days (1-194) and median survival after detection of VRE was 291 (1-2093) days. The diagnoses were 86 cases of AML/MDS (38,57%), 71 NHL/Hodgkin´s lymphomas (27,36%), 38 multiple myelomas (MM) (17,04%), 14 ALL (6,28%) and 24 other malignancies (10,78%). We compared the outcome of patients with VRE-findings and no VRE-findings in the subgroup of AML patients. One year survival rates were 77,5% (55 out of 71) and 74,6% (53 out of 71) respectively (p = 0,839). The rates of discontinuation of AML therapy were 16,41% and 21,09% respectively (p = 0.273). A total of 5 patients out of 223 (2,41%) with VRE developed VRE blood stream infections (BSI). Four of these were suffering from advanced hematological malignancies refractory to chemotherapy (1x AML, 1x AML + melanoma, 2 x NHL). These four patients died 1-15 days after VRE detection in blood cultures. One patient was in first line treatment of a MM. In this case VRE-BSI occurred in neutropenia and resolved without complications after treatment with Linezolid.

Conclusions: In our single-center study the baseline characteristics of patients with VRE colonisation show a representative cross section of all patients with hematological malignancies. VRE colonisation did not show any significant influence on the outcome of patients with AML. VRE-BSI mainly occur in patients with end stage malignant diseases.

Disclosure: No conflict of interest disclosed.

P248 - Computerized routine screening for malnutrition in cancer patients enables early nutritional interventions

Stangl W.M.1, Stimpfl I.2, Guger-Halper U.3, Huber M.1, Szedlak G.1, Stangl T.A.4, Amtmann R.2

1KH Oberwart, Hämato-/Onkologie und Palliativmedizin, Oberwart, Austria, 2KH Oberwart, Diätologie, Oberwart, Austria, 3KH Oberwart, Anstaltsapotheke, Zentrale Zytostatika-Aufbereitung, Oberwart, Austria, 4VU University, Amsterdam, Netherlands

Introduction: The prevalence of malnutrition in hospital patients in Europe lies between 20-50%, but in cancer patients this number goes up to 85%. About 20% of cancer patients die from the consequences of malnutrition and not from the tumour itself. According to a global survey conducted in 2012, 59% of cancer patients experienced weight loss >5% in the last 3 months, but only 8% received dietary counselling.

Methods: Based on the recommendations of the European Society for Nutrition (ESPEN), the parameters of body weight, height and weight loss in the last 3 months were entered as a mandatory field in the record sheet of cancer patients. When a threshold is exceeded by 3 points, the underlying disease and the presence of additional stressors such as surgery, pressure ulcers or concurrent chemotherapy/radiotherapy or the presence of infection must be entered too. This means that with a total number of 3-5 points there is a risk of malnutrition, if the number is >5 a manifest malnutrition exists. In both cases, dietary counselling and specific therapy is automatically requested.The programming of the individual steps were made in i.s.h.med (SIEMENS AG).

Results: During the observation period of 12 months, 237 patients were screened repeatedly for malnutrition (1488 patient cases). In 17 patients (7.17%) there was a score of 3-5 points (risk for malnutrition), and in 35 patients (14.77%), a score of 6 points or more (overt malnutrition). All patients with a score of 3 points or more( n = 52 (21,9%) received an initial consultation on the same day by the dietician and further nutrition- therapy planning.

Conclusion: Our pilot study shows that all cancer patients treated in our institution are recognized as malnourished, or at risk for it, with the establishment of a computerized screening program.

The risk or presence of malnutrition was detected according to the current ESPEN guidelines and targeted nutrition counselling and nutritional intervention were initiated promptly.

Disclosure: No conflict of interest disclosed.

P249 - A retrospective analysis on the complications of intravenous port catheters in haemato-oncological patients

Georgoula L.1, Schlicht E.2, Hebart H.3

1Stauferklinikum Schwäbisch Gmünd, Klinik für Kinder- und Jugendmedizin, Mutlangen, Germany, 2Stauferklinikum Schwäbisch Gmünd, Klinik für Gynäkologie, Mutlangen, Germany, 3Stauferklinikum Schwäbisch Gmünd, Zentrum für Innere Medizin, Mutlangen, Germany

Introduction: Totally implantable port systems allow a simple venous access and are frequently used in haemato-oncological patients. The port chamber can be connected either to an open ended or a valved Groshong catheter.

Methods: In a retrospective analysis we enrolled 294 consecutive patients who underwent a central venous port implantation between 2008-2011. The implanted catheter type and the frequency of catheter-related complications were analysed. In this analysis we compared the type of complications according to the type of implanted catheter.

Results: Overall 310 port systems were implanted in 294 consecutive patients (162 female, 132 male) with an average age of 64,2 years (31-91 years). A system with an open-ended catheter was used in 207 patients (70,4%) whereas a Groshong catheter was used in 87 (29,6%) cases.

Open-ended catheters were inserted into 193 patients suffering from solid tumours, 13 patients from haematological malignancies, and one patient with a benign disease. Groshong catheters were inserted exclusively in female patients suffering from breast or gynaecological cancers.

The total number of catheter-indwelling-days was 123.483 days. Systems with an open-ended catheter remained in situ for a total of 69.593 days (average 325 days), Groshong catheters for 53.890 days (average 561 days).

Overall, complications related directly to the intravenous device occurred in 45 events (14,56%) in 36 patients. In the group of open-ended catheters we observed adverse events in 12,5% (n = 26) of the patients consisting of infectious complications (5,8%, n = 12), thrombotic complications in 1,9% (n = 4), postoperative wound bleeding in 1,4% (n = 3) and dislocation of the catheter in 1% (n = 2). Paravasation injury, pneumothorax and loosening of the port chamber occurred in 0,5% (n = 1) each. In 2 (1%) patients the implantation of the device was unsuccessful. In the Groshong-group we observed 19 complications (21,8%) consisting of thrombosis (6,9%, n = 6), dislocations of the catheter (5,7%, n = 5), infectious complications (3,45%, n = 3) and paravasation injury (3,45%, n = 3). Pneumothorax and wound bleeding were observed in 1,1% (n = 1) each.

Conclusions: Totally implantable port-systems are safe and complication rates are low. In this retrospective study we could not observe any definite advantage of the Groshong-catheter when compared to open-ended catheters.

Disclosure: No conflict of interest disclosed.

P250 - Development and validation of a novel real-time PCR assay for the rapid detection of fungal infections

Rahn S.1, Schuck A.2, Kondakci M.2, Haas R.2, Pfeffer K.1, Henrich B.1

1University of Düsseldorf, Department of Medical Microbiology and Hospital Hygiene, Düsseldorf, Germany, 2University of Düsseldorf, Department of Hematology, Oncology and Clinical Immunology, Düsseldorf, Germany

Introduction: Fungal infections are a major problem in immunocompromised patients, especially as they are difficult to diagnose due to low sensitivity and specificity of available tests. Detection of fungal pathogens by polymerase chain reaction (PCR) has recently become a promising diagnostic tool, although most of them specialize only on few selected fungal species. In this study, a real-time PCR assay was developed consisting of eight multiplex reactions for the standardized, broad range detection and differentiation of fungal pathogens in human specimens.

Methods: The PCR targets the ITS1-ITS2 region of the fungal rDNA gene. Branch-specific sequence regions were identified in multiple sequence alignments and used to design primers for seven real-time PCRs covering distinct fungal branches, in addition to a pan fungal reaction. Positive reactions were detected by subsequent melting curve analysis and the fungal species were identified by Sanger sequencing of the respective PCR-product. Each Real time PCR reaction was validated by testing DNA of respective ATCC strains.121 throat swabs, 119 EDTA and 128 serum samples obtained from 51 patients with hematological malignancies during neutropenia were evaluated prospectively with the PCR assay and the results were viewed in the context of clinical findings.

Results: Using this novel set of PCR assays, 368 samples of immune-compromised patients were analyzed.


Overall, 16 different fungal species were detected with 49.4% (41/83) of the fungi belonging to Candida spp., 19 to Saccharomyces spp., 11 to Cladosporium spp. and 11 to other fungi of clinical relevance. Six of eight patients with highly suggestive signs of fungal pneumonia on high resolution CT had positive findings of Candida sp., Cladosporium sp. and two rarely reported fungi such as Pleosporales and Microdochium.

Conclusion: In this study, a standardized broad range Real time PCR was validated to improve the diagnosis of invasive fungal infections in terms of speed and specificity. At this point, it is possible to identify the fungal genus of a patient sample in a single PCR run. Different methods of DNA extraction will be assessed in further studies to increase the sensitivity of the PCR in patients with hematological malignancies.

Disclosure: No conflict of interest disclosed.

P251 - Clostridium difficile infections after autologous and allogeneic blood stem cell transplantation - a single center experience

Kobbe G.1, Nagorny N.1, Mackenzie C.2, Rachlis E.1, Dienst A.1, Lopez Niedenhoff D.1, Schroeder T.1, Fenk R.1, Pfeffer K.2, Haas R.1, Kondakci M.1

1Uniklinik Düsseldorf, Klinik für Hämatologie, Onkologie und klinische Immunologie, Düsseldorf, Germany, 2Uniklinik Düsseldorf, Institut für medizinische Mikrobiologie und Krankenaushygiene, Düsseldorf, Germany

Patients after blood stem cell transplantation (BSCT) are at risk of opportunistic and nosocomial infections. Among these, symptomatic Clostridium Difficile Infection (CDI) is of special importance because it is the most common nosocomial, antibiotic-associated infection.

Between 1/2006 and 10/2014 a total number of 4797 stool specimens were analyzed for C.-diff. toxin from 751 patients with diarrhea who had previously received an auto or allo BSCT. Sixty-five individuals (MM 25, AML 23, Lymphoma 9, MDS/MPS 7 and 1 germ cell tumor) tested positive, reflecting an infection rate of 8.7% among all BSCT recipients with diarrhea (4,01% for autoTx and 8,16% for alloTx). Five patients (8%) had a history of CDI before Tx. The detection rate varied between 4.1% in 2012 and 10% in 2008 (mean 6.3%). When considering only the 1st year after BSCT, infection occurred earlier after autoTx (median day +37, range 1-331) than after alloTx (median day +85, range 3-364, p = 0.014) and in MM patients (median day +19, range 1-331) than in Lymphoma patients (median day +93, range 59-159, p = 0.03) which may be the result of metronidazole prophylaxis in allografted patients until day +20 after transplant and different previous antibiotic exposure. Of 29 autoTx patients with CDI 21(72%) relapsed with their primary malignancy (11 before, 9 after CDI). Of 36 alloTx patients 18(50%) relapsed (9 before, 9 after CDI). As therapy for CDI 40 patients (61.5%) received metronidazole, 12 (18.5%) vancomycin, 2 (3.1%) fidaxomycin, 7 (10.8%) unknown and 3 (4.6%) no therapy. No patient died of CDI but 16 (24.6%) had at least one recurrence (12 after metronidazole, 1 after vancomycin, 1 after fidoxomycin and 2 after unknown therapy). At day +30 after the 1st positive CDI test 93.1% and 94.5% of patients were alive after autoTx and alloTx, respectively. While overall mortality in CDI patients was similar after autoTX (55%, median FU 202 days, 4-2361) and alloTx (53%, median FU 365 days, 1-3223), causes of death differed (TRM 7%, DRM 48% after autoTX vs TRM 17%, DRM 36% after alloTx).

Between 2006 and 2013 the rate of CDI among patients with diarrhea after BSCT varied from 4.1 to 10% and was related to diagnosis and type of transplant. CDI recurred frequently, especially after metronidazole. While CDI itself was not fatal, patients who developed CDI after blood stem cell transplantation resembled a high risk population with high treatment and disease related mortality.

Disclosure: No conflict of interest disclosed.

P252 - Management of chemotherapy-induced symptomatic anemia with epoetin theta as documented in the multicentre, non-interventional study EPO-amb

Link H.1, Lück A.2, Rauh J.3, Bòrquez D.4, Maas C.5, Hesse J.6, Koenigsmann M.7, Schulze M.8, Tölg M.9, Scheuerlein R.W.10

1Westpfalz-Klinikum Kaiserslautern, Klinik für Innere Medizin I, Kaiserslautern, Germany, 2Zentrum für Urologie und Onkologie, Rostock, Germany, 3Fachinternistische Gemeinschaftspraxis und Therapiezentrum, Witten, Germany, 4Praxis für Onkologie und Hämatologie, Bergisch Gladbach, Germany, 5Gemeinschaftspraxis für Hämatologie und internistische Onkologie, Halberstadt, Germany, 6Gemeinschaftspraxis, Parchim, Germany, 7Onkologisches Ambulanzzentrum OAZ, Hannover, Germany, 8Praxis für Urologie, Andrologie und Onkologie, Markkleeberg, Germany, 9Mediveritas GmbH, München, Germany, 10Teva GmbH, Berlin, Germany

Introduction: Chemotherapy-induced anemia (CIA) in cancer patients correlates with poor performance status and decreased quality of life. Currently by guidelines recommended therapies are erythropoiesis-stimulating agents, Fe substitution or a combination of both; RBC transfusions in case Hb-levels below 8-7g/dl.

Material and methods: NIS EPO-amb is a multicenter, prospective observational study in Germany. 500 adult cancer patients (pts) with CIA treated by epoetin theta (Epo) were planned to be enrolled in 60 sites. We report on an interim analysis of the study.

Results: 310 pts were enrolled by 07/2014. Mean (SD) age of pts was 63.6 (11.3) years with 64% females. The majority of pts had breast cancer (33%), followed by lung cancer (12%) and ovarian cancer (9%). Almost 2/3 of pts underwent chemotherapy with palliative intent. The following results are based on 162 pts with completed CIA treatment (max. over 12 weeks).

CIA management: In addition to Epo 66 pts (40.7%) received transfusions, 28 pts (17.3%) iv Fe and 8 pts (4.9%) oral Fe substitution, resulting in the following CIA-management:


Hb-level: Median (Q1 / Q3) hemoglobin level at baseline (week 1) was 9.7 (9.0/10.3) g/dl . Overall, 93.8% of pts had Hb levels below 12.0 g/dl, 69.8% below 10.0 g/dl. Median Hb-level (Q1 / Q3) increased to 10.3 (9.5/11.2) in week 5 and 10.6 (9.8/11.7) in the last week of Epo treatment (or last Hb-value available).

Epo dosage: Starting dose was 20.000 I.U. in 86.4% of all pts (10.000 I.U: 3.9%, 30.000 I.U: 9.3%, 40.000 I.U.: 0.4%). Only for 23 pts (14.2%) dosage was increased during the observational period. Mean (SD) duration of Epo treatment was 7.9 (3.6) weeks (median: 8.0 weeks).

RBC transfusions: For 66 pts (40.7%) a total of 99 RBC transfusions was reported. Overall, in 78.3% of transfusions the Hb-value at baseline was >8.0 g/dl.

Conclusions: EPO-amb data show that epoetin theta is mainly used according to the labeled recommendation to start with 20.000 I.U. A high proportion of patients received transfusions while this was not justified. Use of guidelines for treatment of CIA should be improved.

Disclosure: Hartmut Link: Advisory Role: Teva, Amgen, Hexal, Vifor-Pharma; Financing of Scientific Research: Teva, Amgen, Hexal, Vifor-Pharma; Expert Testimony: Amgen. Robert Willy Scheuerlein: Employment or Leadership Position: TEVA GmbH.

P253 - Privigen® in secondary immunodeficiencies - interim analysis of a multicenter non-interventional study in Germany

Plath M.1, Slawik H.R.1, Reiser M.2, Otremba B.3, Pfründer D.4

1Onkologische Schwerpunktpraxis, Augsburg, Germany, 2Praxis internistischer Onkologie und Hämatologie, Köln, Germany, 3Onkologische Praxis, Oldenburg, Germany, 4CSL Behring, Hattersheim, Germany

Introduction: Privigen® is a 10% liquid preparation of polyvalent human IgG for intravenous administration. The use of the stabiliser L-proline fully preserves IgG functional activity without refrigeration, making Privigen® ready-to-use. Privigen® is licenced as a maintenance therapy in primary and secondary immunodeficiencies, and as an immunomodulatory therapy in autoimmune and inflammatory diseases.

Methods: This is an interim analysis of an ongoing multicenter non-interventional study to evaluate the efficacy and tolerability of Privigen®, focussing on secondary immunodeficiencies. The cut-off date was March 8, 2015.

Results: 1,554 patients (824 males, 730 females; mean age 67 years) in 145 centers received a total of 18,128 Privigen® infusions. The mean observation period was 12.5 months. The average monthly dose was 14 g (0.2 g/kg body weight). Underlying diseases with n≥5 were chronic lymphocytic leukemia (n = 766), acute lymphocytic leukemia (n = 8), chronic myeloid leukemia (n = 5), acute myeloid leukemia (n = 22), hairy-cell leukemia (n = 5), Hodgkin lymphoma (n = 19), non-Hodgkin lymphoma (n = 373), myeloma (n = 282), macroglobulinemia Waldenström (n = 42), monoclonal gammopathy (n = 21), myelodysplastic syndrome (n = 11) and solid tumor (n = 22). The efficacy was judged as very good or good in 94.4%, moderate in 4.7% and insufficient in 0.9% of evaluable cases (n = 1,439). Patients who had not received any IVIG treatment prior to study entry and who received ≥6 infusions of Privigen® for ≥120 days (infusion intervals ≥20 d and ≤60 d) experienced significantly fewer infections during the study than before: The mean annualized infection rate dropped from 5.4 to 1.6 (n = 143; p < 0.0001). The tolerability was judged as very good or good in 92.7%, moderate in 4.8% and insufficient in 1.9% of all cases (0.6% missing data). Adverse events possibly or probably related to Privigen® were reported for 243 of the 18,128 infusions (1.3%); 17 events were considered serious (0.1%).

Conclusions: Privigen® significantly reduced infection rates in secondary immunodeficiencies. The tolerability was very good or good in the majority of patients.

Disclosure: Margarete Plath: Financing of Scientific Research: Ja. Dietmar Pfründer: Employment or Leadership Position: Ja.

P254 - Fatal outcome of human coronavirus infection despite successful viral elimination by IFN-alpha in a patient with newly diagnosed ALL

Mayer K.1, Nellessen C.1, Wallau A.1, Schumacher M.1, Pietzonka S.2, Drosten C.2, Brossart P.1, Wolf D.1

1Universitätsklinik Bonn, Medizinische Klinik III, Bonn, Germany, 2Universitätsklinik Bonn, Institut für Virologie, Bonn, Germany

Introduction: Human Coronavirus NL63 (HCoV-NL63) is one of four common respiratory CoV. Despite high incidence and options for treatment with interferon, HCoV infections are grossly understudied. We here report a case of HCoV infection in a leukemia patient with fatal ARDS despite successful virus elimination by pegylated Interferon-alpha (PEG-IFN-α).

Case: The 27-year old female pre-T-ALL patient was treated according to the German-Multicenter Trial for Adult ALL (GMALL) 07-03 protocol. No relevant infectious complications were seen until day 35 when the neutropenic patient developed fever without any clinical infectious focus. Antibiotic prophylaxis was switched to meropenem. Two hours later the patient collapsed, a positive shock index immediately triggered ICU referral. Anti-infectious therapy was escalated to additional linezolid and liposomal amphotericine B. The patient deteriorated rapidly with respiratory failure due to ARDS. BAL revealed a significant viral load of HCoV-NL63 (Ct = 30 in real-time RT-PCR, no other respiratory viruses detected). Based on successful application of pegylated IFN against the related SARS-coronavirus in animal experiments, a single injection of 180µg PEG-IFN-α2b was applied. Intravenous immunoglobulins were given because of the wide prevalence of anti-HCoV-NL63 in the population. Steroids were provided to limit inflammation, based on experience with SARS-CoV. Despite immediate initiation of treatment and elimination of virus in subsequent tests, septic shock with progressive lung failure led to death 7 days after onset of fever with massive lung bleeding as a consequence of diffuse alveolar haemorrhage.

Conclusion: This report emphasizes the fatal consequences of common respiratory virus infections in immunocompromized patients. HCoV which in total account for 5-10% of adult upper respiratory tract infections can be blocked by treatment with peg-IFN if diagnosed early.

Disclosure: No conflict of interest disclosed.

P255 - Prophylactic treatment of chemotherapy-induced neutropenia with Lipegfilgrastim in patients with breast cancer: subgroup analysis of the non-interventional study NADIR

Kurbacher C.M.1, Fietz T.2, Salat C.3, Zaiss M.4, Gazawi N.5, Steffens C.-C.6, Egert M.7, Graffunder G.8, Papke J.9, Weißenborn G.10, Bückner U.11, Illmer T.12, Jungberg P.13, Lorenz A.14, Weide R.15, Klare P.16, Tesch H.17, Oskay-Özcelik G.16, Teichmann B.18, Harde J.18, Scheuerlein R.19

1Gynäkologisches Zentrum Bonn-Friedensplatz - Schwerpunkt gynäkologische Onkologie, Bonn, Germany, 2Schwerpunktpraxis für Innere Medizin, Hämatologie, Onkologie und Gastroenterologie, Singen, Germany, 3Hämato-Onkologische Schwerpunktpraxis, München, Germany, 4Praxis für interdisziplinäre Onkologie & Hämatologie, Freiburg, Germany, 5Klinik für gynäkologische Onkologie, Leipzig, Germany, 6MVZ für Hämatologie/Onkologie Klinik Dr. Hancken, Stade, Germany, 7Praxis für Innere Medizin, Hämatologie, Intern. Onkologie, Werdau, Germany, 8Frauenarzt-Zentrum-Zehlendorf, Berlin, Germany, 9Internistische Praxis und Tagesklinik, Neustadt i.S., Germany, 10Praxis für Hämatologie und Onkologie, Twistringen, Germany, 11Hämatologisch-onkologische Schwerpunktpraxis, Bochum, Germany, 12Gemeinschaftspraxis Hämatologie - Onkologie, Dresden, Germany, 13Gynäkologische Praxis, Chemnitz, Germany, 14Frauenarztpraxis, Hildburghausen, Germany, 15Praxis für Hämatologie und Onkologie, Koblenz, Germany, 16Praxisklinik Krebsheilkunde für Frauen / Brustzentrum, Berlin, Germany, 17Hämatologisch-Onkologische Gemeinschaftspraxis am Bethanien-Krankenhaus, Frankfurt a. M., Germany, 18iOMEDICO AG, Freiburg, Germany, 19TEVA GmbH, Berlin, Germany

Introduction: Anthracycline and/or taxane-based chemotherapy regimen are the most effective therapies in breast cancer. Chemotherapy-induced neutropenia (CIN) and febrile neutropenia (FN) are two of the most common and life-threatening toxicities of myelosuppressive chemotherapy regimen leading to dose reductions and delay of cycle intervals. Corresponding guidelines recommend primary prophylaxis with granulocyte colony-stimulating factors (G-CSF) in patients receiving chemotherapy with a high risk for febrile neutropenia (FN). Lipegfilgrastim is a glyco-pegylated r-metHuG-CSF approved to reduce the duration of neutropenia and the incidence of FN. We report on NIS NADIR looking at systemic treatment and use of Lipegfilgrastim in breast cancer pts.

Methods: This prospective NIS was initiated to collect data on prophylactic use of lipegfilgrastim in patients at risk for CIN according to the corresponding guidelines. As of 12/2013 a total 120 outpatient centers and hospitals in Germany are planned to recruit 1.500 patients (pts). Data on demographics, tumour characteristics, comorbidities, previous and ongoing systemic treatments, and Lipegfilgrastim treatment are collected. Pts complete a customized questionnaire focusing on self-injection and handling of the syringe. Main objectives are the incidence of severe neutropenia (NCI CTCAE grade 3-4), FN and resulting complications.

Results: At time of the interim analysis (04/2015) 80 sites enrolled 835 pts. 717 pts were evaluable thereof 317 pts with breast cancer. Mean age was 54.8 (SD: ±11.2) years. At inclusion 89.9% had an ECOG status of 0 or 1. Chemotherapy was mostly given in (neo) adjuvant setting (80.8%). Overall 94.6% pts were treated with taxane and/or anthracycline combinations. 23.3% were treated with dose-dense regimen (q2w regimen including epirubicine and/or cyclophosphamide and/or taxanes). Lipegfilgrastim was mostly given within 3 days after chemotherapy application (93.9%). Nearly all pts treated with dose-dense chemotherapy received Lipegfilgrastim within 3 days after chemotherapy (96%). 1.3% (n = 4) developed FN during the first Lipegfilgrastim supported cycle, thereof one pt (0.3%) received a dose-dense regimen.

Conclusions: These data show, that in a primary (neo) adjuvant setting with mostly anthracycline and/or taxane-based chemotherapy, application and efficacy of Lipegfilgrastim was as expected. The low incidence of FN was comparable to literature.

Disclosure: Christian Martin Kurbacher: Advisory Role: TEVA GmbH; Financing of Scientific Research: TEVA GmbH. Robert Scheuerlein: Employment or Leadership Position: TEVA GmbH.

P256 - Prophylactic treatment of chemotherapy-induced neutropenia with Lipegfilgrastim in patients with NHL: Subgroup analysis of the non-interventional study NADIR

Fietz T.1, Wolff T.2, Sandner R.3, Janssen J.4, Hurtz H.-J.5, Heits F.6, Weide R.7, Weißenborn G.8, Losem C.9, Lück A.10, Kurbacher C.M.11, Teichmann B.12, Harde J.12, Scheuerlein R.W.13

1Schwerpunktpraxis für Innere Medizin, Hämatologie, Onkologie und Gastroenterologie, Singen, Germany, 2OncoResearch Lerchenfeld GmbH, Hamburg, Germany, 3Gemeinschaftspraxis für Hämatologie und Internistische Onkologie, Passau, Germany, 4Gemeinschaftspraxis für Hämatologie und Onkologie, Westerstede, Germany, 5Onkologische Gemeinschaftspraxis, Halle (Saale), Germany, 6Agaplesion Diakonieklinikum Rotenburg gGmbH, Rotenburg (Wümme), Germany, 7Praxis für Hämatologie und Onkologie, Koblenz, Germany, 8Praxis für Hämatologie und Onkologie, Twistringen, Germany, 9Praxis für Onkologie und Hämatologie, Neuss, Germany, 10Zentrum für Urologie und Onkologie, Rostock, Germany, 11Medizinisches Zentrum Bonn-Friedensplatz - Schwerpunkt gynäkologische Onkologie, Bonn, Germany, 12iOMEDICO AG, Freiburg, Germany, 13TEVA GmbH, Berlin, Germany

Introduction: Chemotherapy-induced neutropenia (CIN) and febrile neutropenia (FN) are serious dose-limiting toxicities. Lymphoma patients who develop FN are typically hospitalized and treated with intravenous antibiotics. As a consequence the dose of chemotherapy is often reduced and/or the cycle intervals are prolonged, always associated with poor outcomes. This is why existing guidelines recommend the use of G-CSF in primary prophylaxis for patients (pts) at high FN risk or for pts at intermediate risk in the presence of additional risk factors. Lipegfilgrastim is a glyco-pegylated r-metHuG-CSF approved to reduce the duration of neutropenia and the incidence of FN. We report on interim data obtained during the course of the non-interventional study (NIS) NADIR in NHL patients.

Methods: This prospective NIS NADIR was initiated to collect data on prophylactic use of Lipegfilgrastim in patients with high risk for CIN according to the corresponding guidelines. A total of 120 outpatient centers and hospitals in Germany are planned to recruit 1.500 pts. Data on demographics, tumour characteristics, comorbidities, previous and ongoing systemic treatments, and Lipegfilgrastim treatment are collected. Main endpoint is the incidence of severe neutropenia (NCI CTCAE grade 3-4), FN and resulting complications.

Results: At time of the interim analysis (04/2015) 80 sites in Germany enrolled 835 pts. 717 pts were evaluable with at least one completely documented Lipegfilgrastim cycle. The 91 pts with NHL are discussed here: Mean age was 64.9 (SD ± 12) years, with 67% males. At inclusion 81.4% of all pts had an ECOG status of 0 or 1 (with 16.5% missing data). Overall 39.6% pts were treated with (R)-CHOP, (R)-CHOP-like (24.2%) and (R)-Bendamustin (13.2%). Lipegfilgrastim was mostly given within 3 days after chemotherapy application (88%). For 7.7% (n = 7; CI 3.1-15.2%) dose modification was reported after the first Lipegfilgrastim supported cycle, thereof 2 cases (2.2%) due to neutropenia. No pts developed FN during the first Lipegfilgrastim supported cycle. Twelve pts (13.2% [95%-CI: 7.0-21.9%]) developed NCI CTCAE grade 3 or 4 neutropenia in the first Lipegfilgrastim supported cycle.

Conclusions: Here for the first time data to the application of Lipegfilgrastim in hematological diseases are shown. In a mostly R-CHOP-based chemotherapy setting, application and efficacy of Lipegfilgrastim was as expected. The low incidence of FN is comparable to literature.

Disclosure: Thomas Fietz: Advisory Role: TEVA GmbH; Financing of Scientific Research: TEVA GmbH; Other Financial Relationships: TEVA GmbH Robert Willy Scheuerlein: Employment or Leadership Position: TEVA GmbH.

P257 - Fosfomycin in febrile neutropenic patients with haematological malignancies: A retrospective case documentation

Schwab K.S.1, Mayer K.1, Brossart P.1, Hahn-Ast C.1

1Universitätsklinik Bonn, Medizinische Klinik III, Bonn, Germany

Introduction: Fosfomycin (FOS) is a phosphonic acid derivative with bactericidal activity and an expanded spectrum of activity. There is no published experience of treating invasive infections in onco-hematological patients. We observed that occasionally critically ill neutropenic patients unresponsive to other antibiotics were treated with FOS in our departments. Aim of our study was to analyse effectiveness and toxicity of FOS in febrile neutropenia (FN).

Methods: Data of febrile neutropenic episodes treated with FOS were retrospectively collected. Baseline data of the patients, therapy of haematological malignancy, treatment of infection and adverse events were documented. Success was defervescence (>/= 7d) in absence of any sign of continuing infection.

Results: Data of eighteen patients (12 female, 6 male) were collected so far. Median age was 57 years (range 26-75 years). All patients (pts.) had haematological malignancies (12 AML, 2 ALL, 2 NHL, 2 Multiple Myeloma). All pts. were neutropenic at the onset of fever with a median duration of 19 days (d) (range 5-40 d). The type of infection was pneumonia in 3 pts., 14 Bacteriaemia, 1 FUO. Before initiation of FOS, 8 pts. (44%) received two lines or more of antibiotic therapy. FOS (15g/d) was given for a median of 10 d (range 3-25 d). In 6/16 pts. an invasive fungal infection was considered as possible and in two as probable. A viral infection was never assumed. Treatment was successful in 10/16 pts. (56%), in two patients success of FOS could not be evaluated. Four pts. (22%) died due to infection. Grade 3-4 toxicity did not occur.

Conclusions: Although this analysis included very ill patients, our results showed promising response rates to FOS. Since success rates of escalated antibiotic therapy in FN are generally very low, we consider FOS a promising alternative for salvage antibiotic therapy in FN. This case documentation will be continued to get a more precise idea of the efficacy of FOS in FN.

Disclosure: No conflict of interest disclosed.

P258 - Multiprofessional cancer medication management at the center for integrated oncology

Jansen C.1, Zipfel M.2, Koch L.2, Becker G.2, Schulze I.3, Kuhn W.4, Schmidt-Wolf I.G.H.2, Jaehde U.1

1Institute of Pharmacy, Clinical Pharmacy, University of Bonn, Bonn, Germany, 2Center for Integrated Oncology Cologne/Bonn, Department of Internal Medicine III, University Hospital, Bonn, Germany, 3Hospital Pharmacy, University Hospital, Bonn, Germany, 4Department of Obstetrics and Gynecology, Center for Integrated Oncology Cologne/Bonn, University Hospital, Bonn, Germany

Introduction: Due to the high toxicity of anti-cancer drugs and the complexity of the medication, patients are at high risk of experiencing adverse events. Patient safety may be enhanced by providing a structured multiprofessional medication management for cancer patients.

Methods: The medication management consists of four multiprofessional care modules composed of a basic module (medication review, interaction check) and specific modules for the management of three common adverse events (nausea and emesis, mucositis, fatigue). All care modules contain a care algorithm, evidence-based recommendations for supportive care and patient information brochures. The medication management was tested in a single-arm pilot study by measuring patient-reported symptom load, quality of life and patient satisfaction with information.

Results: For the pilot study 21 cancer patients with any solid tumor were recruited. The results show the feasibility of the medication management and acceptance by the patients and the multiprofessional care team. The most frequently applied adverse event module was nausea and emesis (100%) followed by mucositis (91%) and fatigue (62%). Mainly patients with head and neck cancer receiving concomitant radiochemotherapy showed early toxic symptoms of high severity.

Conclusion: Our pilot study confirmed the feasibility and acceptance of structured multiprofessional care. Assuming that patients with head and neck cancer may particularly benefit from medication management, a randomized two-arm study will be conducted by focusing on this patient cohort. Primary endpoint of the study will be the frequency and severity of treatment-associated toxicity.

Disclosure: No conflict of interest disclosed.


Urogenitale Malignome

P259 - FNTB promoter polymorphisms influence survival in prostate cancer

Virchow I.1,2, Schmid K.W.3, Rübben H.4, Heukamp L.C.5,6, Siffert W.1, Bachmann H.S.1

1Institute of Pharmacogenetics, Medical Faculty, University of Duisburg-Essen, Essen, Germany, 2West German Cancer Center, University Hospital Essen, Department of Medical Oncology, Essen, Germany, 3Institute of Pathology and Neuropathology,Medical Faculty, University of Duisburg-Essen, Essen, Germany, 4Department of Urology, Medical Faculty, University of Duisburg-Essen, Essen, Germany, 5NEO New Oncology, Cologne, Germany, 6Hämatopathologie, Hamburg, Germany

Introduction: Farnesyltransferase inhibitors (FTIs) revealed promising results in vitro as antitumoral therapeutic strategy. They inhibit activation of different tumor related proteins. The reason why FTIs (Lonafarnib, Tipifarnib) failed in large clinical trials remains unclear. Predictive biomarkers to identify patients, who most likely benefit from FTIs would be highly useful. The primary target of FTIs is the farnesyltransferase β-subunit (FNTB). Therefore we functionally analysed polymorphisms in the promoter region of the FNTB gene. In addition the prognostic impact of these polymorphisms were evaluated in patients with prostate cancer.

Methods: Gene sequencing including promoter regions were performed using the method by Sanger. Functional analysis of detected polymorphisms were evaluated by electrophoretic mobility shift (EMSA) and reporter assays. Two independent prostate cancer patient populations (n = 222, n = 179) were retrospectively genotyped for the polymorphisms using restriction enzymes and pyrosequencing. Association of genotypes as well as haplotypes with risk for cancer and disease progression were analysed using Cox regression and Kaplan- Meyer analysis.

Results: Three promoter polymorphisms were detected, -609G/C, -179T/A, and -173delG. EMSA analysis for the polymorphisms -609 and -173 revealed different binding properties of transcription factors, genotype- and haplotype-dependent differences of promoter activity and altered FNTB expression dependent on genotype. Kaplan Meyer analysis showed significant association of polymorphisms and haplotypes with survival in the combined analysis of the -609 and -173 polymorphisms (p = 0.021) in one population. Patients with the -173 del/del genotype combined with the -609 CC genotype show the worst survival, but the best survival if they carry at least one -609 G allele. In the second population the -609 CC genotype independently influenced disease free survival till metastasis. These patients had the worst overall survival (p = 0.006).

Conclusions: In patients with prostate cancer functional FNTB promoter polymorphisms influence FNTB expression and survival. Our results suggest the FNTB polymorphisms as promising target for further evaluation with regard to their prognostic as well as predictive impact.

Disclosure: No conflict of interest disclosed.

P260 - Quality of life (QoL) and patterns of care in second line treatment of metastatic castration resistant prostate cancer (mCRPC) after prior chemotherapy docetaxel chemotherapy: Results of a prospective Swiss observational treatment registry (SEQOND)

Stenner F.1, Betticher D.2, Rothschild S.1, Caspar C.3, Morant R.4, Popescu R.A.5, Rauch D.6, Huber U.S.7, Zenhäusern R.8, Rentsch C.9, Cathomas R.10

1Universitätsspital Basel, Onkologie, Basel, Switzerland, 2HFR Fribourg - Hôpital cantonal, Oncologie, Fribourg, Switzerland, 3Kantonsspital Baden, Medizinische Klinik/ Onkologie, Baden, Switzerland, 4ZeTuP Rapperswil-Jona, Tumorzentrum, Rapperswil, Switzerland, 5Hirslanden Medical Center, Tumor Zentrum, Aarau, Switzerland, 6Spital STS AG, Medizinische Onkologie, Thun, Switzerland, 7Onko-Zentrum, Zürich, Switzerland, 8Spitalzentrum Oberwallis, Onkologie, Brig, Switzerland, 9Universitätsspital Basel, Urologie, Basel, Switzerland, 10Kantonsspital Graubünden, Abteilung Onkologie und Hämatologie, Chur, Switzerland

Introduction: Several new treatment options have demonstrated improved overall survival in the treatment of mCRPC after prior chemotherapy with docetaxel including the cytotoxic agent cabazitaxel. In the pivotal phase III trial of cabazitaxel (TROPIC) no evaluation of QoL on treatment with cabazitaxel was performed. With a variety of second line options the optimal treatment sequence after prior docetaxel remains elusive. We prospectively evaluated quality of life of different treatments at progression after prior docetaxel and describe the patterns of care of second line treatment in Switzerland.

Methods: Patients (pts) with mCRPC and disease progression after having received >225 mg/m2 docetaxel as first line treatment were eligible. Pts were enrolled consecutively in participating centers. Second line treatment consisted of cabazitaxel (25 mg/m2 every 3 weeks) or any other treatment including abiraterone acetate. Choice of second line treatment was at the discretion of the investigator. Primary objective was assessment of QoL maintenance under second line treatment. Secondary objectives included improvement of QoL, evolution of QoL as well as assessment of fatigue and pain during second line therapy. QoL was measured using the FACT-P questionnaire. QoL maintenance was defined as having at maximum a decrease in two functional domains of the FACT-P at any time point compared to baseline. Fatigue was evaluated with the BFI and pain with the MPQ-SF pain questionnaire. Questionnaires were completed at baseline and then monthly for four months. All pts gave their informed consent and an ethical approval was received for all participating centers.

Results: A total of 165 pts were included in 35 Swiss centers between November 2011 and December 2014. 72 pts were treated with cabazitaxel and 93 pts with other second line treatments. The data presented on the primary and secondary endpoints at the time of the meeting will give a detailed overview of the treatment arms and the outcomes. Currently, the data are not mature since not all pts have completed their evaluation period at this time point.

Conclusions: We will be able to show prospectively collected QoL data including fatigue and pain assessments on treatment with cabazitaxel and other second line treatment options. Moreover our data should give insight into the different usage of second line treatment options for pts with mCRPC and describe the patterns of care in Switzerland.

The study was funded by Sanofi/CH.

Disclosure: Frank Stenner: Advisory Role: Nicht regelmässig, aber siehe 5.; Financing of Scientific Research: Im Rahmen von ad hoc advisory boards von Sanofi Honare für Beratertätigkeiten erhalten; Expert Testimony: Finanzielle Unterstützung für wissenschaftliche Projekte durch Sanofi; Other Financial Relationships: Reisekostenerstattungen und Kongressgebühren für Kongresse. Richard Cathomas: Advisory Role: Nicht regelmässig, aber siehe 5.; Financing of Scientific Research: Im Rahmen von ad hoc advisory boards von Sanofi Honare für Beratertätigkeiten erhalten; Expert Testimony: Finanzielle Unterstützung für wissenschaftliche Projekte durch Sanofi; Other Financial Relationships: Reisekostenerstattungen und Kongressgebühren für Kongresse.

P261 - Impact of prior docetaxel, Extent of Disease (EOD), and prior Bisphosphonates (Bp) on Hematologic (Heme) safety of radium-223 Dichloride (Ra-223) from ALSYMPCA

Strauss A.1, Michalski J.M.2, Parker C.3, Sartor O.4, Vogelzang N.J.5, Haugen I.6, Garcia-Vargas J.7, Nilsson S.8

1Universitätsmedizin Göttingen, Urologische Klinik und Poliklinik, Göttingen, Germany, 2Washington University School of Medicine, St Louis, United States, 3The Royal Marsden NHS Foundation Trust and Institute of Cancer Research, Sutton, United Kingdom, 4Tulane Cancer Center, New Orleans, United States, 5Comprehensive Cancer Centers of Nevada, Las Vegas, United States, 6Algeta ASA, Oslo, Norway, 7Bayer HealthCare, Whippany, United States, 8Karolinska University Hospital, Stockholm, Sweden

Objective: Ra-223, a first-in-class α-emitter, reduced risk of death by 30% in phase 3 ALSYMPCA (Parker et al. NEJM 2013; ALSYMPCA) and is approved in 34 countries to date for treatment (tx) of CRPC with symptomatic bone mets and no known visceral disease. In ALSYMPCA, Gr 3 and 4 heme adverse events (AEs) incidence was low, with no clinically meaningful differences vs placebo (Pbo). This post hoc subgroup analysis evaluated whether prior docetaxel, prior Bp, or EOD predisposes pts with Ra-223 tx to myelosuppression.

Methods: ALSYMPCA pts had progressive CRPC with ≥ 2 symptomatic bone mets and no known visceral mets, were receiving best standard of care, and previously received docetaxel or were unfit for or declined docetaxel. Pts (2:1) had 6 injections of Ra-223 (50 kBq/kg IV; q 4 wk) or matching Pbo. Subgroup analyses of Gr 3/4 anemia, neutropenia, and thrombocytopenia by prior Bp and EOD at baseline used Cox regression; a prespecified analysis determined heme AE incidences in docetaxel subgroups.

Results: No tx differences were observed for Gr 3/4 anemia across all subgroups. Gr 3/4 thrombocytopenia was higher in Ra-223 vs Pbo pts for prior docetaxel, EOD > 20 mets/superscan, and prior Bp. For Ra-223, Gr 3/4 neutropenia was higher with prior vs no prior docetaxel; Gr 3/4 thrombocytopenia was higher with prior vs no prior Bp, EOD >20 mets/superscan vs ≤20 mets, and prior vs no prior docetaxel. For Pbo, Gr 3/4 anemia was higher for >20 mets/superscan vs ≤20 mets; Gr 3/4 thrombocytopenia was higher with prior vs no prior docetaxel.

Conclusion: As in ALSYMPCA overall analyses, Gr 3/4 anemia showed no differences for Ra-223 vs Pbo across all subgroups of CRPC pts. Gr 3/4 neutropenia was higher for Ra-223 with prior vs no prior docetaxel. Thrombocytopenia was higher with Ra-223, prior docetaxel, greater EOD, and prior Bp.

Disclosure: Arne Strauss: Financing of Scientific Research: Bayer, Pfizer, Novartis, Amgen. Sten Nilsson: Advisory Role: Bayer.

P262 - Quality-of-life (QoL) of patients with metastatic castration resistant prostate cancer (mCRPC) treated with Cabazitaxel - results from the non-interventional study QoLiTime

Al-Batran S.-E.1, Lange C.2, Ecke T.3, Windemuth-Kieselbach C.4, Kloss S.5, Hammerer P.6, Hofheinz R.-D.7

1Hospital Nordwest, Institute of Clinical Research, Frankfurt, Germany, 2Practice for Urology and Uro-Oncology, Bernburg, Germany, 3Helios Hospital, Department of Urology, Bad Saarow, Germany, 4Alcedis GmbH, Biometry, Gießen, Germany, 5DRK Hospital, Center for Prostate Cancer, Luckenwalde, Germany, 6Academic Hospital Braunschweig, Department of Urology and Uro-oncology, Braunschweig, Germany, 7University Hospital Mannheim, Interdisciplinary Tumor Center, Mannheim, Germany

Background: Cabazitaxel (Caba) combined with Prednis(ol)one is approved for 2nd-line treatment of mCRPC after Docetaxel. Potential toxicity of chemotherapy may impact on patient´s QoL and counterbalance treatment benefits. Thus QoL data are important from physicians´, patients' and regulatory perspective.

Methods: Patients with mCRPC receiving Caba were included in the non-interventional study QoLiTime. EORTC QLQ C30 was handed out at baseline and each cycle. We present QoL results and the correlation of QoL with biochemical response (PSA decrease >50%) of 238 patients from baseline (BL) and after 4 cycles (C4) of Caba.

Results: The total population consists of 527 patients, 406 received 4 cycles of Caba, 266 of those had PSA measurements at BL and C4 and 238 completed the EORTC QLQ 30 at BL and C4. Median age was 72 years with good performance status (ECOG 0:36%; ECOG 1:54%). No significant change in QoL was observed between BL and C4, independent of PSA response. At C4, there was an improvement in physical and role functioning (-5.21; p < 0.0001 and -8.79; p < 0.0001) while there was a worsening of fatigue (6.49; p = 0.0003), dyspnoea (6.36; p = 0.004) and diarrhoea (9.16; p < 0.0001). Other scales did not vary significantly from BL to C4. Improvement in physical functioning and pain decrease (-7.61) were associated with a PSA decrease >50%.

Conclusions: This study is the largest prospective analysis of QoL in patients receiving Cabazitaxel for mCRPC. Symptom increases were seen in typical areas of chemo toxicity such as fatigue and diarrhoea but QoL was maintained during the 12-week observation period. Improvement in physical functioning and pain was associated with a good PSA response.

This study is funded by Sanofi.

Disclosure: Salah-Eddin Al-Batran: Financing of Scientific Research: Sanofi. Ralf-Dieter Hofheinz: Financing of Scientific Research: Sanofi; Expert Testimony: Sanofi.

P263 - Trials in progress: The influence of the therapy sequence in patients with metastatic castration resistant prostate cancer (mCRPC) treated with Cabazitaxel - the international, prospective non-interventional trial SCOPE

Bokemeyer C.1, Stoiber F.2, Amram M.-L.3, Gschwend J.E.4

1University Hospital Eppendorf (UKE), Department of Oncology, Hematology with section Pneumology, Hamburg, Germany, 2LKH Vöcklabruck, Department of Urology and Andrology, Vöcklabruck, Austria, 3Geneva University Hospital, Department of Oncology, Geneva, Switzerland, 4Klinikum rechts der Isar, Technische Universität München, Department of Urology, Munich, Germany

Introduction: In the treatment of patients with metastatic castration resistant prostate cancer (mCRPC) different options at progression after Docetaxel are available. However, clear evidence to decide which drug to use for which patient at what time point is not yet available. Retrospective data have led to the hypothesis that post-docetaxel, the sequence of Cabazitaxel followed by an androgen receptor (AR) targeted agent may be associated with a longer survival than the inverse sequence. The prospective, international, multicenter, non-interventional study SCOPE will evaluate the influence of the sequence of different drugs on the therapeutic outcome by comparison of results based on different Cabazitaxel treatment time points as used in daily clinical practice.

Methods: The SCOPE study will be conducted in 300 centers in Germany, Austria and Switzerland with a total of 900 planned patients, starting in summer 2015. Patients who are started on Cabazitaxel treatment and who have received prior Docetaxel therapy will be consecutively enrolled. The primary objective is to evaluate the impact of treatment sequence on progression free survival (PFS) with cabazitaxel, as assessed by the investigator. The analysis will focus mainly on the following two sequences: 1. Docetaxel pre-treatment followed directly by Cabazitaxel (continuous taxane-therapy sequence), 2. Docetaxel followed by an AR-targeted or other therapy prior to Cabazitaxel (intermittent taxane-therapy sequence). Secondary endpoints include PSA response, time to PSA progression with each therapy, number of Cabazitaxel cycles received, overall survival and safety. Data of each patient will be collected prospectively from time of first dose of Cabazitaxel until 24 months follow-up after first dose of Cabazitaxel or until death.

Conclusion: An important question for the optimal treatment of patients with mCRPC is the best treatment sequence according to patient and tumor characteristics. Selection of therapies may be influenced by potential resistance between AR-targeted therapies and patient performance status. SCOPE aims to analyze and compare outcomes for different sequences in which Cabazitaxel is used in daily clinical practice to gather insight into this important question.

This study is funded by Sanofi.

Disclosure: Carsten Bokemeyer: Advisory Role: Sanofi; Financing of Scientific Research: Sanofi. Jürgen E. Gschwend: Advisory Role: Sanofi; Financing of Scientific Research: Sanofi.

P264 - External-beam radiation therapy (EBRT) use and safety with Radium-223 dichloride (Ra) in patients (pts) with castration-resistant prostate cancer (CRPC) and symptomatic bone metastases (mets) from the ALSYMPCA trial

Strölin P.1, O'Sullivan J.2, Sartor O.3, Parker C.4, Hoskin P.5, Widmark A.6, Mellado B.7, Helle S.8, Aksnes A.9, Garcia-Vargas J.10, Nilsson S.11

1Martini-Klinik am UKE GmbH, Hamburg, Germany, 2Queen's University, Centre for Cancer Research and Cell Biology, Belfast, United Kingdom, 3Department of Medicine: Section of Hematology & Medical Oncology and Department of Urology, Tulane Cancer Center, New Orleans, United States, 4Academic Urology, The Royal Marsden NHS Foundation Trust and Institute of Cancer Research, Sutton, United Kingdom, 5Mount Vernon Hospital Cancer Centre, Middlesex, United Kingdom, 6Radiation Sciences, Oncology, Umeå University, Umeå, Sweden, 7Medical Oncology, Hospital Clinic, Barcelona, Spain, 8Oncology, Haukeland University Hospital, Bergen, Norway, 9Clinical Research, Algeta ASA, Oslo, Norway, 10Oncology Global Clinical Programs, Bayer HealthCare, Whippany, United States, 11Clinical Oncology, Karolinska University Hosiptal, Stockholm, Sweden

Aim: Bone mets in CRPC frequently cause symptomatic skeletal events (SSEs) requiring EBRT for pain. In ALSYMPCA, Ra improved overall survival and delayed time to first SSE (Parker NEJM 2013). This post hoc study analyzed ALSYMPCA pts with EBRT for bone pain before randomization and duringtreatment (tx).

Methods: Eligible pts had symptomatic CRPC with ≥2 bone mets and no known visceral mets; had best standard of care; and had prior docetaxel (pD) or were unfit for, lacked access to, or declined docetaxel (no pD). Pts were stratified by prior docetaxel use (yes/no), baseline total alkaline phosphatase level (ALP; <220 U/L or ≥220 U/L), and current bisphosphonate use (yes/no), and randomized 2:1 to 6 injections of Ra (50 kBq/kg IV q 4 wk) or placebo (pbo). EBRT use for bone pain was assessed. Adverse events were analyzed by concomitant EBRT.

Results: Of 921 ALSYMPCA pts, 30% (186/614) Ra pts and 34% (105/307) pbo pts had EBRT for bone pain. Time to EBRT was significantly longer with Ra vs pbo (HR = 0.67, P = 0.001). Ra vs pbo delayed time to EBRT in pts with <20 mets (HR = 0.49, P < 0.001), current bisphosphonate use (HR = 0.47, P = 0.004), total ALP < 220 (HR = 0.66, P = 0.008), and no pD (HR = 0.65, P = 0.038). In pts with prior EBRT, a higher percentage of pbo vs Ra pts (33% vs 24%) had EBRT for bone pain during the 6 mo of tx. In pts with no prior EBRT, this percentage was also higher for placebo (33% vs 20%), and the difference was even greater at 12 mo after randomization (41% vs 29%). Ra safety profiles were similar with and without concomitant EBRT.

Conclusions: Ra delays the need for EBRT for bone pain vs pbo. Prior EBRT does not appear to affect need for EBRT for bone pain in pts receiving Ra. EBRT use does not affect the Ra favorable safety profile.

Disclosure: Petra Strölin: Other Financial Relationships: Support for travel cost for scientific congresses. SI Nilsson: Advisory Role: has had a consultant or advisory relationship with Algeta and has received remuneration from Bayer HealthCare for travel costs and accommodation for study and writing meetings.

P265 - Effects of radium-223 dichloride (Ra-223) on health-related quality of life (HRQoL) assessed by the EQ-5D utility scores in ALSYMPCA

Strölin P.1, Cislo P.2, Sartor O.3, Reuning-Scherer J.4, Shan M.5, Zhan L.2, Parker C.6

1Martini-Klinik am UKE GmbH, Hamburg, Germany, 2Bayer HealthCare, Global Health Economics and Outcomes Research Department, Whippany, United States, 3Tulane Cancer Center, Departments of Medicine and Urology, New Orleans, United States, 4Yale University, Department of Statistics, New Haven, United States, 5Bayer HealthCare, Department of Statistics, Whippany, United States, 6The Royal Marsden NHS Foundation Trust and Institute of Cancer Research, Academic Urology Unit, Sutton, United Kingdom

Background: Ra-223, a first-in-class α-emitter, significantly improved overall survival versus placebo (pbo) (ALSYMPCA) and had a positive impact on pain and HRQoL in patients (pts) with castration-resistant prostate cancer (CRPC) and symptomatic bone metastases (mets) (Nilsson. ASCO GU 2013). In this HRQoL post hoc analysis, EQ-5D-based utility scores were assessed using treatment (tx), disease progression, and symptomatic skeletal events (SSEs) as predictors.

Methods: EQ-5D, a preferred measure of HRQoL for economic evaluation by the UK National Institute for Health and Clinical Excellence, was used to determine HRQoL utility scores at baseline, weeks 16 and 24, and 8 follow-up visits using UK tariffs. Repeated measures linear regression models were used to estimate tx effects on utility scores for disease states based on measures of disease progression: prostate-specific antigen (PSA), alkaline phosphatase (ALP), and SSEs. Models were adjusted for baseline utility score and trial stratification factors. Least-squares mean (LSMean) utility scores were estimated for each disease state. The relationship between survival and disease states were described with Kaplan-Meier curves and time to death estimates.

Results: LSMean utility score (general measure of HRQoL; score of 1 [full health], 0 [death]) was higher for Ra-223 pts versus pbo pts over the entire trial period and the period prior to an SSE. A tx benefit with Ra-223 was seen in pts with stable disease and prior to an SSE. If a pt experienced ALP progression prior to an SSE, the LSMean utility score decreased (0.62 vs 0.50). However, if progression is defined by PSA, there is little effect of progression on the LSMean utility score (0.61 vs 0.59). There were no tx differences post SSE regardless of ALP or PSA progression.

Conclusion: Ra-223 tx benefit in HRQoL is greatest in stable disease/pre-SSE state. Utility analysis results suggest that ALP and SSE are more appropriate than PSA as measures of progression in pts with CRPC and symptomatic bone mets.

Disclosure: Petra Strölin: Other Financial Relationships: Support for travel cost for scientific congresses. Christopher Parker: Advisory Role: has had a consultant or advisory relationship with Algeta, Bayer, and BNIT. He has also received honoraria from Amgen, Astellas, Bayer, Janssen, Sanofi-Aventis, and Takeda.

P266 - Evaluation of safety, tolerability and activity of Sunitinib in patients (pts) with advanced or metastatic renal cell carcinoma (a/mRCC) in routine clinical practice: the STAR-TOR registry

Woike M.1, Niedtner R.1, Krekeler G.1, Bergmann L.2, Steiner T.3, Goebell P.J.4, Göhler T.5, Harich H.-D.6, Herrmann E.7, Rebmann U.8, Kalanovic D.1

1Pfizer Pharma GmbH, Onkologie, Berlin, Germany, 2Universitätsklinikum Frankfurt, Medizinische Klinik II, Frankfurt, Germany, 3Helios-Klinikum, Urologie, Erfurt, Germany, 4AURONTE (Urologische Klinik und Hämatologie/Onkologie (Med 5)), Universitätsklinikum, Erlangen, Germany, 5Onkozentrum, Dresden, Germany, 6Onkologie Hof MVZ GmbH, Hof, Germany, 7Universitätsklinikum Münster, Urologie, Münster, Germany, 8Diakonissenkrankenhaus Dessau, Urologische Klinik, Dessau, Germany

Introduction: Sunitinib (SUN), an oral tyrosine kinase inhibitor, is approved in the EU for the treatment of pts with a/mRCC. A pivotal study had demonstrated significantly increased progression-free survival (PFS) with SUN compared to the former standard interferon alpha (11 vs. 5 months); p < 0.001). To evaluate the safety profile and efficacy of SUN in a clinical routine setting, data collection in a post-approval non-interventional trial seems to be appropriate.

Methods: A German multicenter registry for pts with a/mRCC (NCT00700258) was amended to include SUN pts in June 2010 with regulatory and ethic committee´s approval. Objectives are the evaluation of the safety profile of SUN, the tolerability and anti-tumor activity of SUN as well as the profile, comorbidity and characteristics of pts and the sequence of systemic therapies in pts with a/mRCC. Inclusion criteria are histologically confirmed a/mRCC treated with SUN and written informed consent.

Results: From July 2010 to February 2015, 108 active study sites recruited 316 pts for this interim analysis. Characteristics: 71.9% male, median age 67.8 years (41.6-87.7), median Karnofsky index 80% (40-100%). Histological subtype: 83.2% clear cell, 16.8% other histologies. In 81.9% of pts SUN is used as first-line therapy. 193 pts were evaluable with regard to MSKCC criteria: 9.3% favorable, 64.2% intermediate, 26.4% poor risk. For all 316 pts, adverse and serious adverse events were observed in 74.7% and 33.9% of the pts (drug related in 60.1% and 14.2% of the pts), respectively. Most common drug-related toxicities (incidence ≥ 5%) of any grade were diarrhea (17.4% of pts), fatigue (13.9%), hand-foot-syndrome (13.0%), thrombocytopenia (11.7%), leukopenia (10.8%), dysgeusia (10.8%), stomatitis (10.4%), nausea (9.8%), anemia (7.6%), mucosal inflammation (7.6%), hypertension (5.7%), and vomiting (5.4%). Median PFS for clear-cell a/mRCC pts (n = 263) was 9.0 months (mo), for other histologies 4.2 mo. The subgroup of clear-cell a/mRCC pts for which SUN was first-line therapy (n = 215) had a PFS of 9.2 mo, for second line clear-cell pts (n = 33) 6.7 mo, ≥ third line (n = 14) 11.5 mo . Overall survival (OS) for the entire study cohort was 18.9 mo.

Conclusions: The patient population in the registry represents the expected pattern in pts with a/mRCC regarding distribution of age, sex, and histology. Efficacy and tolerability of SUN in routine clinical practice confirms published data.

Disclosure: Michael Woike: Employment or Leadership Position: Pfizer Pharma GmbH, Deutschland. Daniel Kalanovic: Employment or Leadership Position: Pfizer Pharma GmbH, Deutschland.

P267 - Evaluation of safety, tolerability and activity of Temsirolimus in patients (pts) with advanced or metastatic renal cell carcinoma (a/mRCC) in routine clinical practice

Woike M.1, Strunz A.M.1, Krekeler G.1, Bergmann L.2, Steiner T.3, Goebell P.J.4, Göhler T.5, Harich H.-D.6, Herrmann E.7, Rebmann U.8, Kalanovic D.1

1Pfizer Pharma GmbH, Onkologie, Berlin, Germany, 2Universitätsklinikum Frankfurt, Medizinische Klinik II, Frankfurt, Germany, 3Helios-Klinikum, Urologie, Erfurt, Germany, 4AURONTE (Urologische Klinik und Hämatologie/Onkologie (Med 5)), Universitätsklinikum, Erlangen, Germany, 5Onkozentrum, Dresden, Germany, 6Onkologie Hof MVZ GmbH, Hof, Germany, 7Universitätsklinikum Münster, Urologie, Münster, Germany, 8Diakonissenkrankenhaus Dessau, Urologie, Dessau, Germany

Introduction: Temsirolimus (TEM), an i.v. mTOR inhibitor, is approved in the EU for the first-line treatment of pts (pts) with a/mRCC who have at least 3 of 6 prognostic risk factors. A pivotal study had demonstrated significantly increased overall survival (OS) with TEM in poor risk pts compared to the former standard interferon alpha (10.9 vs. 7.3 mo; p = 0.0078). To evaluate the safety profile and efficacy of TEM in a clinical routine setting, collection of data in a post-approval non-interventional trial is useful.

Methods: A German multicenter registry for pts with a/mRCC treated with TEM was started in January 2008 (NCT00700258) with regulatory and ethic committee´s approval. Objectives are the evaluation of the safety profile of TEM, the tolerability and anti-tumor activity of TEM as well as the profile, comorbidity and characteristics of pts and the sequence of systemic therapies in pts with a/mRCC. Inclusion criteria are histologically confirmed a/mRCC treated with TEM and written informed consent.

Results: From February 2008 to February 2015, 108 active study sites recruited 535 pts. Characteristics: 68.9% male, median age 66.9 years (34.9-87.0), median Karnofsky index 80% (40-100%). Histological subtype: 75.3% clear cell, 11.6% papillary, and 2.2% chromophobe RCC. 182 pts were evaluable with regard to modified MSKCC criteria. 78.6% of these pts are classified as poor risk. Median number of metastatic sites is 2 (0-6). Adverse and serious adverse events are observed in 72.7% and 43.4% of the pts (drug related in 41.3% and 9.5% of the pts), respectively. Most common drug-related toxicities (incidence ≥ 3%) of any grade are fatigue (7.9% of pts), anemia (7.3%), rash (6.4%), pruritus (6.1%), oedema of any kind (5.9%), mucosal inflammation (4.1%), thrombocytopenia (3.9%), nausea (3.7%), diarrhoea (3.7%), and stomatitis (3.7%). Median progression-free survival for the total patient population is 4.4 months. A subgroup analysis of pts with baseline LDH ≤ 300 U/l (n = 299) vs > 300 U/l (n = 106) shows significant longer OS for the low LDH group (11.4 vs. 7.2 months, p = 0.015). Median OS for all pts is 10.5 months.

Conclusions: The patient population in the registry represents the expected pattern in pts with a/mRCC regarding distribution of age, sex, and histology. Safety and efficacy data of TEM in routine clinical practice confirm current phase III data. Baseline LDH > 300 U/l seems to be a negative prognostic parameter for OS.

Disclosure: Michael Woike: Employment or Leadership Position: Pfizer Pharma Gmbh, Deutschland. Daniel Kalanovic: Employment or Leadership Position: Pfizer Pharma GmbH, Deutschland.

P268 - Decision making in metastatic renal cell cancer - A retrospective analysis of predictive factors for application of 2nd line therapy

Eggers H.1, Ganser A.1, Grünwald V.1

1Medizinische Hochschule Hannover, Hannover, Germany

Introduction: In renal cell cancer (RCC) not eligible for a curative treatment option tyrosinekinase inhibitors (TKI) have become the standard treatment for first line therapy. Thereby treatment options have grown significantly compared to the era of interleukin-therapy. Consequently new prognostic scores, risk stratification and treatment algorithms had to be developed. Nevertheless we are still in need for information to improve personalized treatment pathways. The aim of this analysis was to determine differences between patients who received a second line therapy to patients who did not (for whatever reason).

Material and methods: A random sample of 161 patients treated for RCC between 2005 and 2012 was included. Clinicopathologic data was evaluated Chi-square or Fisher´s exact tests. Continuous parameters were compared applying the Mann-Whitney test. Survival analysis was conducted using Kaplan-Meier analysis, univariate and multivariate cox regression analysis.

Results: There was no significant difference between clinical baseline parameters in patients receiving a 2nd line therapy to patients who did not (table 1). In univariate analysis receiving a 2nd line therapy was associated with a significantly better outcome (HR 1,75; p = 0,008; table 2). 3-year overall survival (OS) was 71% in patients with 2nd line compared to 47,6% without (p = 0.007; Fig 1). Progression within 6m of therapy occurred more often in patients receiving no 2nd line therapy (27% vs. 21%, respectively, p = 0.063). Interestingly, the fraction of patients receiving no 2nd line therapy with progressive disease (PD) in less than 6m was larger than the same fraction in patients with PD in more than 6m (40% vs. 21%, p = 0.063). Due to small sample sizes there was no clinicopathologic attribute at all significantly associated with OS in multivariate analysis.

Conclusion: The major limitations of our analysis are it's retrospective nature, small sample size and missing values. A predictive baseline characteristic could not be identified. 2nd line therapy was given irrespective of MSKCC risk category, but early progression was seen more frequently among patients without 2nd line treatment. Improvement for 3-year-OS was detected in patients who were eligible for a 2nd line therapy, underscoring the relevance of continuous treatment in RCC.

Disclosure: No conflict of interest disclosed.

P269 - Marine compound Frondoside A effectively inhibits proliferation of urothelial carcinoma cell lines independent of p53 activity

Madanchi R.1, Dyshlovoy S.1,2,3, Honecker F.4, Otte K.1, Alsdorf W.1, Schumacher U.1, Hauschild J.1, Bokemeyer C.1, von Amsberg G.1

1UKE, Hamburg, Germany, 2G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Vladivostok, Russian Federation, 3Far Eastern Federal University, Vladivostok, Russian Federation, 4ZeTuP, St. Gallen, Switzerland

Introduction: Despite the progress in the therapy of advanced or metastatic urothelial carcinoma, Vinflunin is the only second line treatment approved after failure of Cisplatin based therapy to date - however, with only moderate activity. Thus, novel therapeutic strategies are urgently needed. We examined the efficacy of Frondoside A - a marine natural compound belonging to the group of triterpene glycosides, in a human urothelial carcinoma model.

Methods: Anticancer activity of Frondoside A (FronA) was investigated using the 6 human urothelial carcinoma cell lines. The effects on cell viability and cell cycle were investigated using trypan blue staining, MTT cytotoxicity assay, and flow cytometry. Mode of action was analyzed by Western blot analyses. The impact of p53 activity for the cytotoxic activity of FronA was analyzed using siRNA transfection and co-treatment with pifithrin-α - a chemical inhibitor of p53 activity. Synergistic effects of FronA in combination with Cisplatin were investigated using the Chou-Talalay method.

Results: FronA significantly inhibited proliferation and reduced viability of 6 human urothelial carcinoma cell lines in a dose dependent manner with IC50 of 0.5~2 µM, while IC50 of Cisplatin were higher ranging from 2~6 µM. The cell lines used for investigation has been the following: RT-112, RT-4, HT-1197, TCC(sup), T-24 and 489p. FronA induced apoptosis by caspase-3, -8, and -9 activation, as well as PARP cleavage. Note, inhibition of caspase-3 activity did not suppress the pro-apoptotic activity of the drug, suggesting caspase-independent character of the induced apoptosis. Inactivation of ERK and p38, and activation of JNK as well as up-regulation of pro-apoptotic Bax were detected. Up-regulation of p21 was observed, although there were no significant effects on cell cycle progression.

Silencing of p53 gene via siRNA transfection or inhibition of p53 activity by pifithrin-α did not abolish the cytotoxic effect of FronA. In contrast, Cisplatin acivity was significantly reduced.

Furthermore synergistic effects of FronA with Cisplatin were detected in different urothelial carcinoma cell lines.

Conclusion: Marine compound Frondoside A is a promising novel therapeutic option for the treatment of advanced or metastatic urothelial carcinoma showing high efficacy in vitro. Inductions of p53- and caspase-3-independent apoptosis are its suggested modes of action. Currently in vivo efficacy and toxicity of FronA are examined.

The first and second author contributed equally.

Disclosure: No conflict of interest disclosed.

P270 - A diagnostic and therapeutic challenge - CD 138 positive cells in bone marrow in a patient with urothelial carcinom of the renal pelvis

Müller L.1, Tiemann M.2, Schulz H.3, Haferlach T.4

1Onkologie UnterEms, Leer-Emden-Papenburg, Leer, Germany, 2Hämatopathologie, Hamburg, Germany, 3Überregionales Institut für Pathologie, Wilhelmshaven, Germany, 4Münchner Leukämielabor MLL, München, Germany

Background: Plasmacytoid urothelial carcinoma (PUC) is a rare and aggressive var. bladder cancer (BC) that mimics plasmacytoma cytologically, that can be confused with hemolymphoid neoplasms secondarily affecting the urinary bladder.

For the first time to our knowledge we describe a pat. with an urothelial carcinoma (UC) of the renal pelvis who presented with a plasmocytoid CD 138+ UC.

Patient: In 01/15 the pat. presented with macrohematuria. In ACT was a left sided renal pelvis mass. In biopsy was a UC G3, nephroureterectomy + lymphadenectomy was performed, no signs of metastatic disease could be detected. The histology showed an UC, G3 left renal pelvis + infiltration in the pararenal adipose tissue, no N+ (0/6).

03/15 anemia, thrombopenia, elevated LDH + diffuse bone pain. In bone scan showed diffuse bone metastasis in skull, total spine, pelvis, ribs, humerus+femura). Retrospectively, preop. CT scans showed no sign for bone metastases.

In BM was a diffuse infiltration of plasmocytoid cells up to 80% with suspicion for multiple myeloma with CD 138+ cells. Myeloma could be ruled out by immune cytology + IH with AE1/3, CK20 +CK7 pos. By FISH + selection for CD138+ cells (purity in cytospin >80% of cells) the cells showed trisomie 1 and trisomie 17. A thoroughful reexamination of the primary showed focal PUC

Conclusions: We described for the first time to our knowledge a PUC of the renal pelvis. We detected this PUC by chance because of the bone metastases mimicing undifferentiated myeloma cells. Until 2012 only 62 cases of PUC have been described in the literature, but only in the bladder[i] [ii]. All published data of the PUC of the bladder showed all a dismal prognosis, much worth than papillary urothelial carcinoma with early metastases, specially in the focal PUC. In older series the 5-years DFS in T3+ tumors ranged from 8%-40%[iii] [iv]. As in our case the PUC was detected mostly in G3 tumors, sometimes after reexamination and they all were CD 138-positive. In FISH we could describe a trisomie 1 and trisomie 17 in a part of the tumor cells as investigated in the bone marrow after CD138 selection.

We think esp. in G3 UC one should investigate esp. for CD138+PUC because of the bad prognosis + develop new therapies.


1 Fritsche HM et al.: J Urol., 180(5);2008:1923-1927.

2 Wang Z et al. Int J Clin Exp Pathol. 5;2012:601-608.

3 Ghoneim MA, 180(1):2008:121-127.

4 Shariat SF et al.: J Urol.,176;2006:2414-2422.

Disclosure: No conflict of interest disclosed.

P271 - Efficacy and safety of sequential high-dose chemotherapy (HDCT) for relapsed or refractory germ cell cancer in a unicentric standardized approach

Niegisch G.1, Henn A.1, Zaum M.1, Kobbe G.2, Haas R.2, Schirren J.3, Albers P.1, Lorch A.1

1Universitätsklinikum Düsseldorf, Urologische Klinik, Düsseldorf, Germany, 2Universitätsklinikum Düsseldorf, Klinik für Hämatologie, Onkologie und Klinische Immunologie, Düsseldorf, Germany, 3Horst-Schmidt-Kliniken, Klinik für Thoraxchirurgie, Wiesbaden, Germany

Introduction: HDCT with carboplatin and etoposide (CE) followed by residual tumor resection (RTR) is a routine regimen for patients (pts) with refractory germ cell cancer (rGCC). We evaluated the results of a unicentric approach with one oncology, one urologic and one thoracic surgeon.

Methods: All 26 pts who underwent HDCT for rGCC between 03/12 and 02/2015 were analyzed. Mobilization chemotherapy (paclitaxel/ifosfamide [TI] or VIP) was followed by 3 cycles of high-dose CE (500 mg/m² each, d 1-3). Whenever feasible, all residual tumors were resected immediately after HDCT. Kaplan-Meier method was used to estimate progression-free (PFS) and overall survival (OS).

Results: Median age of pts was 41 [range 20-57] years. Histology was non-seminoma in 20/26 pts, 24/26 pts had gonadal primaries. HDCT as 1st salvage was performed in 19/26 pts (prognostic score: 5/19 very low/low, 9/19 intermediate, 5/19 high/very high risk) and as 2nd salvage in 7/26 pts

TI for mobilization was used in 24/26 pts, VIP in 2/26 pts. Median time from mobilization to d1 HDCT was 14 [range 8-27] days. Stem cell harvest was sufficient for 3 cycles in all pts.

All HDCT cycles were administered in 22/26 pts. Dose-adjustment due to upfront impaired renal function was required in 4/26 pts. Median time between HDCT cycles was 28 [range 21-36] days. Neither treatment-related deaths nor renal failure requiring hemodialysis was observed. Major toxicities occurred in 5/25 pts (4x sepsis, 1x ileus).

Complete response by HDCT alone was achieved in 5/26 pts, 13/26 pts got one or more RTRs. Median time to first RTR was 41 [range 27-72] days. Major surgical complications were observed in 2/13 pts (1x urinoma, 1x wound dehiscence). Histologically 7/13 pts had viable cancer, 2/13 teratoma only and 4/13 necrosis.

Maximum response to protocol was CR in 14 pts, PRm- in 7 pts, PRm+ in 4 pts and PD in 1 of 26 pts. Median follow-up was 16.3 [range 4.2-34.3] months with 1-year PFS and 1-year OS of 56 [95% CI 39-79] and 78 [95% CI 62-97]%.

Conclusions: A unicentric approach in a high-volume center of expertise provides low complication rates both during HDCT and perioperatively and enables salvage treatment in an optimal timely manner. Even limited by a short follow up outcomes of our series emphasizes the efficacy of HDCT.

Disclosure: No conflict of interest disclosed.

P272 - Efficacy of kinase inhibitors in germ cell cancer cell lines with differential cisplatin sensitivity

Schaffrath J.1, Schmoll H.-J.1, Müller-Tidow C.1, Müller T.1

1Martin-Luther-Universität Halle-Wittenberg, Klinik für Innere Medizin IV - Hämatologie und Onkologie, Halle (Saale), Germany

Introduction: Testicular germ cell tumors (GCTs) are the most common malignancy in young adult men between the age of 15 and 35. Although cisplatin-based chemotherapy is highly effective even in advanced stages, approximately 20% of patients develop cisplatin resistance which is associated with an unfavorable prognosis. Therefore, new therapeutic options capable of overcoming this resistance are urgently needed. We examined the efficacy of kinase inhibitors targeting mTOR (RAD001), EGFR, HER2/neu, VEGFR (AEE788) and IGF-1R (AEW541) and their influence on cisplatin resistance in germ cell cancer cell lines with differential cisplatin sensitivity.

Methods: The in vitro activity of RAD001, AEE788 and AEW541 in the cisplatin sensitive GCT cell lines H12.1 and GCT72F as well as in the resistant cell lines H12.1RA, H12.1ODMneu, 1411HP and 1777NR was investigated using the sulforhodamin-B-(SRB)-cytotoxicity-assay. The agents were applied alone and in combination with cisplatin. To evaluate the activity of the kinase inhibitors, western blot analysis of both the targeted receptors and their phosphorylated state was performed both before and after exposure to each substance.

Results: The investigated kinase inhibitors RAD001 (IC50 0,17 to > 10 µM), AEE788 (IC50 0,13 to > 10 µM) and AEW541 (IC50 0,21 to 1,80 µM) showed cytotoxic activity in all GCT cell lines and significantly inhibited proliferation in both cisplatin sensitive and resistant cell lines. While the examined cell lines showed different expression profiles of the targeted receptors, there was no correlation between their expression and phosphorylation and the effectiveness of the agents. The combination of cisplatin and kinase inhibitors showed both additive and antagonistic effects in the studied cell lines, yet no significant synergistic effect was observed.

Conclusion: According to our data, the investigated kinase inhibitors are effective in germ cell cancer cell lines independent of their cisplatin sensitivity. However, when combined with cisplatin they do not demonstrate any promising ability to overcome cisplatin resistance in testicular GCTs.

Disclosure: No conflict of interest disclosed.

P273 - β-1,4-Galactosyltransferase-I expression in peripheral T-lymphocytes is associated with relapse-free survival in testicular cancer patients

Nilius V.1, Timmesfeld N.2, Neubauer A.1, Brendel C.1

1Uni Marburg, Hämatologie, Marburg, Germany, 2Institut für medizinische Biometrie und Epidemiologie, Marburg, Germany

Outcome of germ cell tumor patients with progression or relapse after a cisplatin-based first line chemotherapy is highly heterogeneous, ranging from 10 to 70% overall survival. As growing evidence suggests that immunosurveillance is important for tumor control we investigated β-1,4-Galactosyltransferase-I (B4GALT1) expression levels in a cohort of 49 testicular cancer patients. High expression levels of B4GALT1 in peripheral blood T-lymphocytes were associated with a better relapse-free survival (RFS) (p ≤ 0.006). B4GALT1 is a cell surface molecule involved in immune cell cross-talk, expression levels proved to be more accurate in predicting RFS than the IGCCCG scoring system in our patient cohort (p ≤ 0.006 versus 0.068). Moreover, interleukin 10 (IL10), a cytokine released by cytotoxic T-cells, was also significantly elevated in T-lymphocytes of non-relapse germ cell tumor patients (p ≤ 0.025).

Our data indicate that T-lymphocyte biology may play a pivotal role in disease control in testicular cancer patients with relapsed or progressive disease state.

Disclosure: No conflict of interest disclosed.


MDS Differentialdiagnose und Therapiemodalitäten

V274 - MDS: Diagnosis and differential diagnoses

Pfeilstöcker M.1

1Hanusch KH, 3. Med Abt, Wien, Austria

Despite recent developments in diagnostic tools and methods diagnosis and differential diagnoses are still a challenge in myelodysplastic syndromes (MDS). One reason is the heterogeneity of the disorder with many different underlying molecular mechanisms, therefore there is still no single test available that defines MDS unambiguously. In addition the slow onset of MDS especially in low risk cases poses diagnostic difficulties in early disease. In principle each cytopenia that cannot be explained by other reasons such as vitamin deficiencies, infections, autoimmune or metabolic disorders or toxic effects on the bone marrow has to be evaluated for the presence of MDS, while other clonal hematopoietic disorders such as leukemias, paroxysmal nocturnal hemoglobinuria, aplastic anemia, myeloproliferative neoplasms (myelofibrosis) or lymphoma (hairy ell leukemia) have to be considered as well. Diagnosis of MDS is made by bone marrow puncture (cytology including iron staining and histology). After exclusion of other reasons evidence of dysplasia according to published diagnostic criteria establishes the diagnosis. Metaphase cytogenetics complemented by FISH analysis is mandatory also for prognostication, FACS and molecular analyses are still optional as their role in classification and prognosis is presently being established. IDUS (idiopathic dysplasia of undetermined significance) and ICUS (idiopathic cytopenia of undetermined significance) designate states where the diagnostic criteria of MDS are not yet fulfilled, recently a partially overlapping state of CHIP (clonal hematopoiesis of indeterminate potential) - with ascertained clonal hematopoiesis but not necessarily cytopenia - has been described with yet unclear clinical consequences. After a diagnosis of MDS has been made subtypes according to the WHO classification should be defined and prognostic risk groups assessed using instruments such as the recently improved international prognostic scoring system IPSS (-R) which uses number and degree of cytopenias, bone marrow blast count and refined cytogenetics as prognostic parameters that may be complemented with differentiating features such as age. Assessment of ferritin, endogeneous erythropoietin levels and optionally HLA typing completes initial work up. Combining these disease related features with patients´ resources (performance status, comorbidities, geriatric assessment) finally provides the tools for clinical decision making.

Disclosure: No conflict of interest disclosed.

V276 - Therapy in MDS

Platzbecker U.1, Deutsche MDS SG

1Uniklinik, Dresden, Germany

The diversity of MDS makes the disease challenging and “truly personalized” not only in terms of diagnostics but also in carrying out clinical decisions. The heterogeneity of MDS manifests in the individual patient as a disease ranging from an indolent condition with a considerable life expectancy to forms approaching the aggressiveness of acute myeloid leukemia (AML). A risk-adapted treatment strategy is therefore mandatory for a disease showing such a highly variable clinical course. Prognostic factors may be subdivided into those related to the general patient's characteristics and health condition and those related to the MDS disease itself. During the past decades treatment has been stratified according to the International Prognostic Scoring System (IPSS) risk score; i.e. into “lower-risk” MDS (low/int-1, LR-MDS) where correction of cytopenia was the main objective and “higher-risk” MDS (int-2/high, HR-MDS) where the reduction or delay of progression or AML evolution and prolonged survival was the objective.

Disclosure: Uwe Platzbecker: Financing of Scientific Research: Celgene, Novartis, Amgen; Expert Testimony: Celgene, Novartis, Amgen.

Wissenschaftliches Symposium

NHL indolent neue Substanzen

V277 - Antibody-drug conjugates in indolent lymphoma

Viardot A.1

1Klinik für Innere Medizin III, Universitätsklinikum, Ulm, Germany

Monoclonal antibodies revolutionized many fields of hemato-oncology. Antibody-drug conjugates (ADCs) realize ideally the Paul Ehrlich´s conception of “magic bullets”. Recently, two ADCs were approved by the European Medical Agency (EMA): brentuximab vedotin in Hodgkin´s disease and anaplastic large cell lymphoma and trastuzumab emantansine in HER2+ metastatic breast cancer. In follicular lymphoma (FL), there is a long-term experience with radioimmunoconjugates (RIC). For example, Yttrium-90 Ibritumomab is approved in relapsed FL and as consolidation therapy after chemoimmunotherapy. In contrast to RICs, the ideal target for ADCs is internalized after binding of the antibody. CD20, which remains stable at the cell surface after binding, is less suitable than the targets CD19, CD22, CD37, CD74 or CD79b. The most frequently used toxins - aurastatins, calicheamicins and maytansinoids - are synthetically derived from natural compounds and too toxic for direct application. Few drugs have already finished phase-I/II studies including patients with indolent lymphomas. Inotuzumab ozogamicin (anti-CD22) has an overall response rate (OR) of 68% in FL at maximum tolerated dose (MTD) (Advani A et al., J Clin Oncol 28:2085, 2010). Polatuzumab vedotin (against CD79b) shows an OR 47% in FL at MTD (MC Palanca-Wessels et al., Lancet Oncology 2045:70128, 2015). Coltuximab-ravtansine (against CD19) has a response rate of 40% in FL (V Ribrag, Clin Cancer Res 20:213, 2014). Addition to Rituximab increased the response rate in inotuzumab ozogamicin (OR 87% in FL, L Fayad et al., J Clin Oncol 31:573, 2013) and polatuzumab vedotin (OR 60% in FL, F Morschhauser et al., J Clin Oncol 32:5s, 2014 (suppl; abstr 8519)). However, the first randomized trial with inotuzumab ozogamicin and rituximab versus the investigator´s choice was terminated due to slow accrual (NCT562965). New promising combinations of ADCs with other drugs will be tested in ongoing clinical trials. Antibody-drug conjugates will complement the therapeutic options in indolent lymphomas in the next years.

Disclosure: Andreas Viardot: Advisory Role: Janssen, Gilead, Amgen, Roche; Financing of Scientific Research: Janssen, Roche, Pfizer; Other Financial Relationships: Reisekosten: Roche, Amgen.

V279 - B-cell receptor signaling in lymphoid malignancies - oncogenic driver and therapeutic target

Schmitt C.1

1Charité - Universitätsmedizin Berlin, Campus Virchow-Klinikum, Hämatologie, Onkologie und Tumorimmunologie, Berlin, Germany

B-cell receptor (BCR) signaling emerged as a key therapeutic target in lymphoma. In normal B-cells, the BCR determines antigen specificity and marks their unique identity. Composed of two identical heavy and light chains, the BCR is the product of a complex B-cell-specific genomic affinity maturation process that includes V(D)J recombination, somatic hypermutation and class switch recombination. Typically following ligand-induced receptor aggregation, the BCR mediates phosphorylation of the associated Ig-a/Ig-b (CD79A/CD79B) immunoreceptor tyrosine-based activation motifs (ITAMs) by SRC kinases, which initiates a cascade of downstream events including the Bruton's Tyrosine Kinase (BTK), phosphatidylinositol 3-kinase (PI3K) and NF-kB signaling, collectively amplifying the original BCR signal. Antigen-dependent and potentially ligand-independent “tonic” BCR signaling is essential for normal B-cell survival and function.

Importantly, most mature B-cell malignancies rely on functional BCR signaling as well, although the underlying mode of activation may be diverse (including tonic signaling, exposure to [auto]-antigens, or self-aggregation of the BCR). This is further underscored by their usage of a restricted, non-random set of stereotyped IgH V segments, aberrant somatic hypermutation, and activating mutations in signaling components (such as CD79/ITAM or CARD11) of the BCR cascade.

B-cell lymphomas may present with BCR signaling-related vulnerabilities that can be exploited by novel targeting approaches. Most prominently, the BTK inhibitor Ibrutinib or the d-isoform-specific PI3K inhibitor Idelalisib have been clinically approved, and numerous novel compounds are in pre-clinical or early clinical testing. However, functionally poorly understood BCR-governed signaling networks and their different wiring across B-cell lymphoma entities, as well as considerable genetic heterogeneity within histological or even molecularly defined subtypes seem to account for the remarkable diversity in clinical responsiveness to these novel agents. While BCR signaling-targeting agents undoubtedly opened a new era in lymphoma therapy, their specific mode(s) of action and the signaling conditions under which they exert their activity require further investigation regarding enhanced efficacy, less toxicity, personalized stratification and treatment optimization, at least in part, via rational co-targeting of critical collateral pathways.

Disclosure: No conflict of interest disclosed.

Freier Vortrag


V285 - European working group for adult ALL phase II trial of nilotinib in combination with chemotherapy for first-line treatment in elderly patients with de novo Philadelphia-chromosome positive acute lymphoblastic leukemia (EWALL-PH-02)

Pfeifer H.1, Cayuela J.-M.2, Spiekermann K.3, Beck J.4, Jung W.5, Viardot A.6, Schäfer-Eckhart K.7, Reichle A.8, Maury S.9, Schmitz N.10, Heidenreich D.11, Panse J.12, Junghanß C.13, Raffoux E.14, Suarez F.15, Guillerm G.16, Alexis M.17, Lissandre S.18, Huguet F.19, Isnard F.20, Lepetre S.21, Escofffre-Barbe M.22, Ribera J.-M.23, Goekbuget N.1, Dombret H.14, Hoelzer D.24, Rousselot P.25, Ottmann O.1

1Goethe University Hospital, Frankfurt/Main, Medizinische Klinik II, Frankfurt am Main, Germany, 2Hôpital Saint Louis, Molecular Biology Department, Paris, France, 3Ludwig-Maximilians-Universität, München, Germany, 4Universitätsklinik, Mainz, Germany, 5Universitätsklinik, Göttingen, Germany, 6Universitätsklinik, Ulm, Germany, 7Klinikum Nürnberg Nord, Nürnberg, Germany, 8Universitätsklinik, Regensburg, Germany, 9Hopital Henri Mondor, Creteil, France, 10Asklepios Klinik St. Georg, Hamburg, Germany, 11Universitätsmedizin, Mannheim, Germany, 12RWTH Aachen University Hospital, Aachen, Germany, 13Universitätsklinik, Rostock, Germany, 14Hopital Saint-Louis, APHP, Paris, France, 15Hopital Necker, Paris, France, 16Hopital Norvan, Brest, France, 17CHR Orleans, Orleans, France, 18CHU de Tours, Tours, France, 19Hopital Purpan, University, Toulouse, France, 20Hopital Saint Antoine, APHP, Paris, France, 21CLCC H Becquerel, Rouen, France, 22CHU Rennes, Rennes, France, 23Universitat Autonoma de Barcelona, Barcelona, Spain, 24Onkologikum, Frankfurt am Main, Germany, 25Universite Versailles Saint-Quentin-en-Yvelines, Versailles, France

Introduction: Imatinib in combination with chemotherapy is considered to be the gold standard for treatment in patients with Philadelphia positive (Ph+) acute lymphoblastic leukaemia (ALL). But the overall prognosis particularly in elderly patients (pts.) remains poor. Nilotinib is a more effective second generation ABL kinase inhibitor (TKI). Data on its efficacy in Ph+ ALL is limited. The EWALL (European Working Group for Adult ALL) initiated a prospective, investigator-initiated multicenter European clinical trial to examine the efficacy and safety of nilotinib in conjunction with chemotherapy in newly diagnosed elderly Ph+ ALL pts (registered under EudraCT no.:2010-022855-46).

Methods: Male or female pts. >55 years with untreated Ph+ and/or BCR-ABL1 positive ALL were treated after a pre-phase, with nilotinib 400 mg BID during the induction period in combination with weekly vincristine (VCR) and dexamethasone (DEX) for 4 weeks. Consolidation consisted in nilotinib with methotrexate and asparaginase for cycles 1, 3 and 5 and cytarabine for cycles 2, 4 and 6. Maintenance phase consisted of nilotinib sequentially with 6-MP and methotrexate orally one every other month and Dex/VCR every two months up to 24 months of treatment.

Results: Between 2012 and 2014, 56 pts. (25 male, 31 female) have been enrolled. Median age is 65 years (55-85 years), twelve pts. are older than 70 years of age. To date, all pts. are evaluable for safety and 47 pts. are evaluable for efficacy. The CHR rate is 87%, one pt. was refractory (2%), one pt. had a partial remission (2%). One pt. died during induction therapy (2%), three pts. discontinued therapy before CR evaluation (6%). With a median follow-up of 5.5 months, 34 of the 41 pts. who achieved CR are in CCR and 3 pts. relapsed, two of whom had discontinued study treatment to undergo allogeneic SCT. 9 pts. have discontinued study treatment prematurely because of transfer to allogeneic SCT, as explicitly permitted by the protocol. The rate of complete molecular remission (B/A < 10E-03) after induction (25 pts. evaluable) was 45.5%, 5 pts. had undetectable BCR-ABL1 transcripts. Tolerability has been acceptable. Infectious events and neutropenic fever predominated.

Conclusion: Nilotinib in conjunction with chemotherapy according to the EWALL-PH-02 protocol is highly effective and well tolerated in elderly pts. with newly diagnosed Ph+ ALL. Molecular response rates are high and MRD levels in responding pts. decrease with time.

Disclosure: Heike Pfeifer: No conflict of interest disclosed. Oliver Ottmann: Advisory Role: Novartis, BMS, Ariad, Pfizer; Financing of Scientific Research: Novartis, BMS, Ariad, Pfizer.

V286 - Silencing of GATA3 defines a new stem cell subgroup of T-ALL

Fransecky L.1, Neumann M.1, Heesch S.1, Schlee C.1, Ortiz Tanchez J.1, Heller S.1, Schwartz S.1, Mochmann L.H.1, Isaakidis K.1, Bastian L.1, Mossner M.2, Herold T.3, Goekbuget N.4, Baldus C.D.1

1Charité, University Hospital Berlin, Campus Benjamin Franklin, Hematology/Oncology, Berlin, Germany, 2University Hospital Mannheim, Medical Faculty Mannheim of the University of Heidelberg, Hematology/Oncology, Mannheim, Germany, 3University Hospital Grosshadern, Ludwig-Maximilians-University (LMU), Department of Internal Medicine III, Munich, Germany, 4Goethe University Hospital, Hematology/Oncology, Frankfurt/Main, Germany

Introduction: The critical role of GATA3 is well characterized in later stages of T-cell differentiation, but its role for the generation of early-T-cell precursors (ETP) has only recently been explored. As aberrant GATA3 mRNA expression has been linked to tumorigenesis, we here investigated the role of GATA3 in T-cell acute lymphoblastic leukemia (T-ALL).

Patients and methods: Seventy-one ETP-ALL samples, sent to the central diagnostic reference laboratory of the German Multicenter Study Group for Adult Acute Lymphoblastic Leukemia, were investigated by RT-PCR analysis for GATA3 mRNA expression. In addition, pyrosequencing was performed to determine GATA3 DNA methylation in 69 of 71 samples. Additionally, global DNA methylation was assessed in 12 ETP-ALL samples using the Illumina Infinium® HumanMethylation450 BeadChip platform. Microarray expression data (Affymetrix HG-U133 Plus 2.0) of an independent cohort of 83 T-ALL patients, which included both ETP-ALL and non-ETP-ALL patients, was used for read out of a GATA3-specific gene expression profile.

Results: One third of ETP-ALL samples lacked GATA3 expression (23/71, 32%, GATA3null ETP-ALL). Global DNA methylation analysis revealed a 6kb segment within the GATA3 locus with significant DNA hypermethylation (46% v. 19%, p < 0.001) containing 35 differentially methylated sites. We confirmed GATA3 hypermethylation in the ETP-ALL cohort (n = 69) and found that GATA3 mRNA expression and DNA methylation were inversely correlated (r = -0.73, p < 0.001). GATA3null ETP-ALL was characterized by a high frequency of FLT3 mutations (15/19, 79%) and a low frequency of NOTCH1 mutations (2/19, 11%). Gene set enrichment analysis of GATA3null in an independent cohort of T-ALL revealed enrichment of genes associated with hematopoietic stem cells and depletion of genes involved in T cell differentiation. Among upregulated genes in GATA3null samples, we identified molecules such as FLT3, C-KIT, BCL2, and BTK that could potentially be accessible for targeted therapies.

Conclusion:GATA3 silencing occurs in a third of adult ETP-ALL patients and is associated with GATA3 DNA hypermethylation. While GATA3positive ETP-ALL might be susceptible to conventional lymphoid directed therapies, GATA3null ETP-ALL exhibits stem cell leukemia properties, which might be associated with distinct sensitivities towards targeted therapies.

Disclosure: No conflict of interest disclosed.

V287 - Droplet digital PCR to quantify minimal residual disease in acute lymphoblastic leukemia

Koopmann J.1, Pott C.1, Füllgrabe M.1, Knecht H.1, Gökbuget N.2, Kneba M.1, Brüggemann M.1

1Universitätsklinikum Schleswig-Holstein, Campus Kiel, Medizinische Klinik II, Kiel, Germany, 2Goethe University Hospital, Medizinische Klinik II, Frankfurt, Germany

Introduction: Minimal residual disease (MRD) has been proven to be the most powerful prognostic factor in ALL. The current molecular gold standard for MRD quantification is the allele specific real-time quantitative PCR (ASO RQ-PCR) of clonal immunoglobuline (IG) and T-cell receptor (TR) gene rearrangements. However, new high throughput tools emerge to overcome some limitation of RQ-PCR. Therefore, we compared the performance of droplet digital (dd)PCR and RQ-PCR to quantify MRD in ALL.

Material and methods: We adapted ASO ddPCR for MRD monitoring in ALL and checked linearity, reproducibility, repeatability, lower limit of detection and lower limit of quantification. Then we analyzed a total of 154 samples (17 diagnostic, 93 follow-up, 44 serial dilutions of diagnostic samples) of adult patients with ALL using IG/TR ASO ddPCR and compared results with standardized RQ-PCR. RQ PCR was performed according to EuroMRD guidelines.

Results: Sensitivity and lower limit of quantification of ddPCR as defined by EuroMRD guidelines was comparable to RQ-PCR using the same amount of input DNA. ddPCR results proved to be highly reproducible, even at the lower limit of quantification. Comparison of MRD analysis using ddPCR and ASO RQ-PCR showed good concordance (r = 0.83, p < 0.0001) between both methods. In contrast to RQ-PCR MRD quantification of follow up samples using ddPCR was possible without the usage of serial dilutions of the diagnostic sample as external calibrator thereby significantly reducing labor intensiveness compared to RQ-PCR. In addition, it allowed the exact quantification of the clonal marker in the diagnostic sample. This represents a major advantage of ddPCR compared to RQ-PCR as it enables the exact quantification of tumor load and thereby the identification of oligoclonality at diagnosis even in patients with low infiltrated diagnostic samples. This was highlighted by comparison of the two molecular MRD approaches in one patient where ddPCR of the diagnostic sample identified subclonality of the marker IGH gene rearrangement. In contrast, RQ-PCR of serial dilutions of the calibrating diagnostic sample showed late amplification and poor sensitivity which precluded correct RQ-PCR based MRD quantification of follow-up samples in this case.

Conclusion: We showed that ddPCR is a promising tool for IG/TR based MRD analysis in ALL. Prospective analyses of unselected cases have to be performed to verify the clinical impact of ddPCR-based MRD detection.

Disclosure: No conflict of interest disclosed.

V288 - Treatment of B-lymphoblastic leukemia cells with demethylating agents enhances the sensitivity to cytostatic drugs

Konkolefski C.1, Roolf C.1, Sklarz L.-M.1, Sekora A.1, Murua Escobar H.1, Junghanss C.1

1Universitätsmedizin Rostock, Klinik III, Hämatologie, Onkologie, Palliativmedizin, Rostock, Germany

Introduction: Aberrant methylation of tumorsuppressor gene promoters is frequently observed in acute lymphoblastic leukemia (ALL). Several studies have shown that hypermethylation is an independent prognostic factor for disease-free survival and play a critical role in drug resistance.

Azacitidine (AZA) and decitabine (DAC) are methyltransferase inhibitors which partially reverse aberrant DNA methylation and sensitize malignant myeloid cells to cytostatics.

However, it is not yet clear whether these agents are equally effective in ALL cells. Therefore, we examined the impact of AZA and DAC on B-ALL cells in single application and in combination with other cytostatics to investigate the sensitizing effect.

Methods: B-ALL cell lines SEM and RS4;11 (both harbouring a t(4;11)) were treated with increasing concentrations of AZA or DAC (100-1.000 nM) for up to 72 h. Cell proliferation and metabolism were examined by trypan blue staining and WST-1 testing. Cell cycle and apoptosis changes were analyzed by flow cytometry. For drug combination experiments, cells were exposed to AZA or DAC (500 or 100 nM) in combination with IC80 values of ara-C, dexamethasone and doxorubicin. Thereby, the demethylating agents were applied in different preparations: simultaneously as well as 24 hours before or after the respective cytostatics. Changes in cell proliferation were determined by WST-1 and drug synergy was evaluated using BLISS statistic.

Results: In SEM cells, AZA and DAC induced dose-dependent antiproliferative effects starting at a concentration of 100 nM. Metabolic activity was significantly decreased by AZA: to 65.6 ± 15.3% and DAC: to 32.7 ± 9.3% compared to control cells (100%). Both drugs increased the number of cells in G0/G1 stage of the cell cycle and induced apoptosis. Treatment with AZA or DAC was less effective in RS4;11 cells compared to SEM.

Enhanced antiproliferative effects on cells were detected when DAC was combined with doxorubicin or ara-C. Here, metabolic activity decreased significantly when DAC and doxorubicin or ara-C were given simultaneously or when cells were pre-treated with doxorubicin, compared to single drug treatment.

Conclusion: Demethylating agents possess antiproliferative effects on B-ALL cells in vitro and may be beneficial for combined chemotherapy in ALL. Different response rates of the cell lines on AZA and DAC may be referred to altered methylation patterns, which is currently under investigation.

Disclosure: No conflict of interest disclosed.

V289 - Deletions of CDKN2A/B and PAX5 are frequently restricted to the CD19+ compartment and associated with concurrent deletions of ABL in adult BCR-ABL+ BCP-ALL

Bartels M.1, Nagel I.2, Ussat S.3, Ottmann O.4, Pfeifer H.4, Trautmann H.1, Böttcher S.1, Gökbuget N.4, Kneba M.1, Oberg H.-H.3, Siebert R.2, Brüggemann M.1

1Universitätsklinikum Schleswig-Holstein, Campus Kiel, 2. Medizinische Klinik, Kiel, Germany, 2Universitätsklinikum Schleswig-Holstein, Campus Kiel, Institut für Humangenetik, Kiel, Germany, 3Universitätsklinikum Schleswig-Holstein, Campus Kiel, Institut für Immunologie, Kiel, Germany, 4Universitätsklinikum Frankfurt, 2. Medizinische Klinik, Frankfurt, Germany

Introduction: BCR-ABL fusion positive (BCR-ABL+) B-cell precursor acute lymphoblastic leukemia (BCP-ALL) may originate at distinct stages of hematopoiesis. It is unclear, weather concurrent CDKN2A/B- and PAX5-Deletions can also be assigned to certain hematopoietic compartments. Therefore we traced the distribution of driver mutations within relevant hematopoietic compartments of BCR-ABL+ adult BCP-ALL cases with concurrent CDKN2A/B- and PAX5-Deletions in order to detect targetable immunophenotypic patterns of involvement.

Methods: Cryo-conserved samples from initial diagnosis of 12 adult BCR-ABL+ BCP-ALL patients with known CDKN2A/B deletions treated within the GMALL 07/2003 trial were flow sorted and screened for BCR-ABL, CDKN2A/B and in 5 cases also for PAX5 deletions by fluorescence in situ hybridization (FISH) using one customized (PAX5) and two commercial probes (Vysis LSI BCR/ABL Dual Color, Dual Fusion Translocation probe, Vysis LSI CDKN2A/CEP 9; both Abbott Molecular, Illinois, USA). The following compartments were analyzed: CD34+38-19-10-3-, CD34+38+19-3-, CD34+19+20-3-, CD34+19+20+3-, CD34-19+20+3-, CD34-19-20-3+, CD34+19-13/33+10-3/16-, CD34-19-13/33+10-3/16-, CD34-19-13/33+10-3/16+.

Results: Diverse individual subclones with CDKN2A/B deletions were exclusively detectable within the B-committed progenitor compartment (CD34+19+) in 10 out of 12 cases although concurrent BCR-ABL+ multipotent progenitors were evident in 4 cases. Cells with a PAX5 deletion had a CD34+19+ phenotype in all 5 investigated cases, including 3 cases with evident BCR-ABL+ multipotent progenitors. Finally, 10 patients of the cohort showed major leukemic (sub)clones with atypical signal patterns after FISH with the BCR/ABL probe: in 7 cases the major subclone was characterized by a missing ABL signal.

Conclusions: While the t(9;22) frequently represents an antecedent event prior to B-lineage determination in the genesis of BCR-ABL+ BCP-ALL this does not hold true for CDKN2A/B and PAX5. In our cohort the different subclones with deletions of CDKN2A/B and PAX5 were commonly restricted to the CD34+19+ B-committed progenitor compartment regardless of BCR/ABL+ multipotent progenitors. Our observation may help to identify patients that particularly profit from B-cell specific immunotherapy. Monosomy 9 probably accounts for the association of ABL-deletions with CDKN2A/B cna in some but not all cases.

Disclosure: No conflict of interest disclosed.

Freier Vortrag

CLL klinisch

V290 - Impact and predictors of reducing prescribed doses of fludarabine, cyclophosphamide and rituximab (FCR) in frontline treatment of chronic lymphocytic leukemia (CLL)

Kovacs G.1, Bahlo J.1, Kluth S.1, Fink A.M.1, Cramer P.1, von Tresckow J.1, Maurer C.1, Langerbeins P.1, Groß-Opphoff-Müller C.1, Fischer K.1, Wendtner C.-M.2, Kreuzer K.-A.1, Stilgenbauer S.3, Hallek M.1, Eichhorst B.1, Goede V.1

1Uniklinik Köln, Klinik I für Innere Medizin (DCLLSG), Köln, Germany, 2Klinikum Schwabing, Klinik für Hämatologie, Onkologie, Immunologie, Palliativmedizin, Infektiologie und Tropenmedizin, München, Germany, 3Uniklinik Ulm, Klinik III für Innere Medizin, Ulm, Germany

Introduction: Frontline treatment with full-dose FCR is considered standard of care for physically fit patients with CLL. In routine practice, however, adverse events often lead to reduction in dose of FCR which might result in a decrease of efficacy. The aim of our study was to systematically evaluate the impact of reducing the prescribed dose of FCR on its efficacy and to identify clinical and biological predictors for such dose modification.

Methods: Patients treated with FCR within two randomized phase III trials of the German CLL Study Group (GCLLSG) (CLL8: FC (n = 409) vs. FCR (n = 408); CLL10: FCR (n = 282) vs. bendamustine plus R (n = 279)) were pooled (n = 690). The planned FCR dose (according to protocol) was compared with the actually applied dose in each patient and differences were provided in%. Patients with ≤20% and with >20% reduction in planned dose were compared with regard to progression free (PFS) and overall survival (OS). Logistic regression was used to identify predictors of such modification in dosage.

Results: 681 of 690 patients were treated with at least one dose of FCR. Of these, 253 (37.2%) received FCR with either FC, R, or both reduced by 20% or more. These patients had significantly shorter PFS (41.6 vs. 64.0 months, HR = 1.78, 95%CI = 1.45-2.23) and OS (90.2 months vs. not reached, HR = 2.15, 95%CI = 1.57-2.96) compared to patients with a ≤20% reduction in dose (Figure 1-2). In univariate and multivariate analyses, age >60 years, Binet stage C and serum β2-microglobuline >3.5 mg/l were identified as independent predictors for reducing the prescribed dose of FCR.

Conclusions: This pooled analysis of two GCLLSG trials showed that reduction of full-dose FCR treatment significantly mitigates its efficacy and results not just in impaired PFS, but also OS. Predictors of dose reductions during therapy with FCR may help to refine treatment decision-making in CLL.


Disclosure: Gabor Kovacs: Other Financial Relationships: travel grant by Mundipharma. Valentin Goede: Advisory Role: F. Hoffmann-La Roche; Financing of Scientific Research: F. Hoffmann-La Roche, Mundipharma, Glaxo Smith Kline, Bristol Myers Squibb.

V291 - Results from the phase 2 RESONATE™-17 trial: Efficacy and safety of Ibrutinib in patients with relapsed or refractory chronic lymphocytic leukemia or small lymphocytic lymphoma with Del17p

Stilgenbauer S.1, Jones J.A.2, Coutre S.E.3, Mato A.R.4, Hillmen P.5, Tam C.6, Osterborg A.7, Siddiqi T.8, Thirman M.J.9, Furman R.R.10, Ilhan O.11, Keating M.12, Call T.G.13, Brown J.R.14, Stevens-Brogan M.15, Li Y.15, Clow F.15, James D.F.15, Chu A.D.15, Hallek M.16, O'Brien S.12

1University of Ulm, Ulm, Germany, 2The Ohio State University, Division of Hematology, Columbus, United States, 3Stanford University School of Medicine, Division of Hematology, Stanford Cancer Center, Stanford, United States, 4Hackensack University Medical Center, Hackensack, United States, 5The Leeds Teaching Hospitals, St. James Institute of Oncoogy, Leeds, United Kingdom, 6Peter MacCallum Cancer Centre, East Melbourne, Australia, 7Karolinska University Hospital Solna, Department of Hematology, Solna, Sweden, 8City of Hope National Medical Center, Department of Hematology/Hematopoietic Cell Transplantation, Duarte, United States, 9The University of Chicago Medicine, Section of Hematology/Oncology, Chicago, United States, 10Weill Cornell Medical College, Division of Hematology-Oncology, New York, United States, 11Ankara University Medical Faculty, Ankara, Turkey, 12The University of Texas MD Anderson Cancer Center, Department of Leukemia, Houston, United States, 13Mayo Clinic, Division of Hematology, Rochester, United States, 14Dana Farber Cancer Institute, Division of Hematologic Malignancies, Boston, United States, 15Pharmacyclics, Inc., Sunnyvale, United States, 16University Hospital Cologne, Department I of Internal Medicine and Center of Integrated Oncology Cologne Bonn, Cologne, Germany

Introduction: Patients with relapsed/refractory (R/R) chronic lymphocytic leukemia (CLL) with del17p have poor treatment outcomes. Ibrutinib (Imbruvica®) is a first-in-class BTK inhibitor approved by EMA for adult CLL patients with ≥1 prior therapy and first-line CLL patients with del17p or TP53 mutation who are unsuitable for chemotherapy. We report a primary analysis of the Phase 2 RESONATE™-17 trial evaluating efficacy and safety of single-agent ibrutinib in R/R del17p CLL or small lymphocytic lymphoma (SLL).

Methods: Patients with del17p CLL/SLL who failed ≥1 prior therapy received oral 420 mg ibrutinib once daily until disease progression. Primary endpoint was overall response rate (ORR) per independent review committee (IRC). Other endpoints were duration of response (DOR), progression-free survival (PFS), overall survival (OS) and safety. The analysis was conducted 12 months after final enrollment and included all patients receiving ≥1 dose of ibrutinib.

Results: Of 144 patients, 137 had CLL and 7 had SLL. Median age was 64. At baseline, 63% were Rai Stage III/IV; 49% had bulky lymphadenopathy ≥5 cm. Patients had a median of 2 prior therapies. Investigator-assessed ORR was 83%, with complete response (CR) or CR with incomplete bone marrow recovery (CRi) in 2% of patients and partial response with lymphocytosis (PR-L) in 17%. ORR by IRC was 65%, including 4% PR-L. At 11.5 months of median follow-up, median PFS, OS and DOR had not been reached. The 12-month PFS rate was 79.3% and 12-month OS was 83.5%. Of 20 patients (13.9%) who progressed, 11 (7.6%) had Richter's transformation. Most common adverse events (AE) were diarrhea (36%), fatigue (31%), cough (24%) and arthralgia (22%); most were Grade 1-2. Most common Grade 3-4 AEs were neutropenia (14%), anemia (8%), pneumonia (8%) and hypertension (8%). Atrial fibrillation occurred in 11 patients (7.6%; 3.5% Grade 3/4) and major hemorrhage in 7 patients (4.9%, all Grade 2/3). Sixteen patients (11.1%) discontinued treatment due to AEs. At data cut, 70% of patients continued on ibrutinib.

Conclusions: In the largest prospective study to date of patients with del17p CLL/SLL, ibrutinib demonstrated marked efficacy in measures of ORR, DOR, PFS and OS, with an AE profile similar to that of earlier studies. These results support ibrutinib as an effective treatment for patients with del17p CLL/SLL.

Disclosure: Stephan Stilgenbauer: Advisory Role: Pharmacyclics; Financing of Scientific Research: Pharmacyclics. Susan O´Brien: Financing of Scientific Research: Janssen; Expert Testimony: Pharmacyclics.

V292 - Bendamustine and Rituximab in combination with Lenalidomide in patients with relapsed or refractory and in patients with previously untreated chronic lymphocytic leukemia: A multicenter phase I/II safety and efficacy trial of the German Chronic Lymphocytic Leukemia Study Group (GCLLSG)

Pflug N.1, Maurer C.1, Bahlo J.1, Kluth S.1, Rhein C.1, Cramer P.1, Gross-Ophoff C.1, Langerbeins P.1, Fink A.-M.1, Eichhorst B.1, Kreuzer K.-A.1, Tausch E.2, Stilgenbauer S.2, Böttcher S.3, Döhner H.2, Kneba M.3, Hallek M.1, Wendtner C.-M.4, Bergmann M.4, Fischer K.1, Deutsche CLL Studiengruppe

1Klinik I für Innere Medizin, Uniklinik Köln, Köln, Germany, 2Klinik III für Innere Medizin, Uniklinik Ulm, Ulm, Germany, 3Klinik II für Innere Medizin, Uniklinik Schleswig-Holstein, Kampus Kiel, Kiel, Germany, 4Klinik für Hämatologie, Onkologie, Immunologie, Palliativmedizin, Infektiologie und Tropenmedizin, München, Germany

Introduction: The combination of Bendamustine plus Rituximab (BR) is indicated in treatment-naïve as well as relapsed/refractory (R/R) patients with CLL. The immunmodulatory drug lenalidomide (L) has proofed efficacy in heavily pre-treated CLL. This phase I/II trial was initiated to assess safety/efficacy of the combination BRL in previously untreated (1st-line) and R/R patients with CLL.

Patients and methods: Seventeen R/R and five previously untreated medically fit patients with CLL (CIRS score < = 6) were enrolled to receive the combination of BRL. In the R/R phase I- cohort, four different dose levels of L (maximum 15 mg per day [pd]) were tested. In the 1st-line cohort, L was dose escalated from 5 to 15 mg pd. B was administered intravenously (i.v.) with a dose of 50/90 mg/m2 (R/R/1st-line) on day 1 & 2 of each 28 day cycle while R i.v. was dosed with 375 mg/m(2) on day 0 of the first cycle and 500 mg/m(2) on day 1 during subsequent cycles for up to six cycles.

Results: The maximal tolerable dose of L determined in the phase I-part of the R/R cohort was 5 mg pd. Overall response rate was 47.1% in the R/R and 60% in the 1st-line cohort, respectively. Median progression-free survival (PFS) was 8.0 months for both cohorts. Median overall survival was 22.9/12.3 months (R/R/1st-line). Grade 3/4 hematologic toxicity was observed in 75% of the R/R- and in 60% of patients receiving 1st-line treatment. Severe infections CTC grade 3-4 occurred in 60% and 43.8% of the patients respectively. Due to high toxicity and low remission rate of BRL the trial was closed prematurely.

Conclusion: BRL was associated with a high rate of severe infections, a low response rate and short PFS compared to other 1st- and 2nd-line regimens for CLL and cannot be considered an appropriate treatment option for those patients.

Disclosure: No conflict of interest disclosed.

V293 - Outcomes of anticoagulant (AC) or antiplatelet (AP) use in patients (pts) with chronic lymphocytic leukemia (CLL) or indolent non-Hodgkin's lymphoma (iNHL) in idelalisib (IDELA) trials

Stilgenbauer S.1, Barrientos J.2, Ghia P.3, Pagel J.4, Salles G.A.5, Sharman J.P.6, Gurtovaya O.7, Kim Y.7, Philip B.7, Zelenetz A.D.8

1Universität, Ulm, Germany, 2Hofstra North Shore-LIJ School of Medicine, Hempstead, United States, 3Università Vita-Salute San Raffaele and Instituto Scientifico San Raffaele, Milano, Italy, 4University of Washington Fred Hutchinson Cancer Research Center Medical Oncology, Seattle, United States, 5Université Claude Bernard, Pierre Bénite, France, 6Willamette Valley Cancer Institute and Research Center, Springfield, United States, 7Gilead Sciences, Inc., Foster City, United States, 8Memorial Sloan Kettering Cancer Center, New York, United States

Introduction: IDELA, a selective oral PI3Kδ inhibitor, is approved for use in relapsed CLL (in combination with rituximab [R]) and iNHL (as monotherapy). Both diseases occur mainly in the elderly, who have comorbidities that increase thrombotic risk. This post hoc analysis characterized the use and outcomes of AC/AP therapy, which was allowed in IDELA registrational clinical trials.

Methods: In the phase 3 Study 312-116 (NCT01539512), frail pts with relapsed CLL (including those with any degree of thrombocytopenia) were randomized to receive a combination of continuous IDELA 150 mg BID or placebo (PBO) with 8 R doses. In the phase 2 Study 101-09 (NCT01282424), pts with refractory iNHL received IDELA 150 mg BID until disease progression or unacceptable toxicity. Grade 1, 2, and ≥3 bleeding events were analyzed using MedDRA preferred terms and CTCAE.

Results: The 2 trials included 343 pts. In the CLL study, 18 pts (16%) on IDELA + R and 31 (29%) on PBO + R had grade ≥3 thrombocytopenia at baseline. Concomitant AC/AP use was frequent (45% in each study); the most common were aspirin, enoxaparin, and warfarin. AC/AP use was more frequent in pts treated with IDELA + R vs PBO + R. The incidence of bleeding events was similar with IDELA, IDELA + R, and PBO + R. Grade ≥3 bleeding events occurred in 1 IDELA + R, 1 PBO + R, and 3 IDELA pts.

Conclusions: AC/AP use involved 45% of the IDELA registrational trial population. Overall, rates of bleeding events were moderate and similar with IDELA + R vs IDELA + PBO; grade ≥3 events were uncommon. There was no specific trend with regard to AC/AP and bleeding events in the 2 arms of the CLL study.


Disclosure: Stephan Stilgenbauer: Advisory Role: Gilead Sciences; Financing of Scientific Research: Gilead Sciences; Expert Testimony: Gilead Sciences. Andrew Zelenetz: Advisory Role: Gilead Sciences; Expert Testimony: Gilead Sciences.

V294 - Ibrutinib as salvage therapy for relapsed high-risk chronic lymphocytic leukemia (CLL) in patients after allogeneic hematopoietic stem cell transplantation (HSCT)

Hahn M.1, Dietrich S.1, Görner M.2, Welslau M.3, Zenz T.1, Rummel M.4, Schmitt M.1, Köhler A.5, Ho A.D.1, Dreger P.1

1Universität Heidelberg, Medizinische Klinik V, Heidelberg, Germany, 2Klinikum Bielefeld, Klinik für Hämatologie, Onkologie und Palliativmedizin, Bielefeld, Germany, 3Klinikum Aschaffenburg, Praxis für Onkologie, Aschaffenburg, Germany, 4Universitätsklinikum Gießen und Marburg, Medizinische Klinik IV, Hämatologie, Gießen, Germany, 5Asklepiosklinik Langen, Praxis für Hämatologie und Onkologie, Langen, Germany

Introduction: Patients with high-risk CLL relapsed after HSCT have an overall poor prognosis. The upcoming era of small molecule inhibitors may offer new therapeutic options in that situation.

Study design and patients: In a single centre retrospective analysis, the outcome of ibrutinib salvage therapy was assessed in patients with high-risk CLL relapsed after HSCT.

Results: Ibrutinib was administered to 8 patients (m/f= 1/1) with a median age of 53y at ibrutinib start. 7/8 had been transplanted from an unrelated donor (6 MUD, 1 MMUD), 4/8 were 17p- at relapse.

The median time from HSCT to ibrutinib start was 33 (5-88) months. Ibrutinib was the primary relapse therapy in 4 patients. The remainder received ibrutinib after failure of pretreatment with rituximab + DLI (2 patients) or with rituximab + DLI followed by rituximab + lenalidomide (2 patients).

With a median therapy duration of 7 (1-14) months, no grade 3/4 toxicity has been observed. All 8 patients are in ongoing response: Six patients have accomplished a partial remission (PR), one patient being under therapy only since one month reported symptom relief, and one patient has achieved an MRD-negative complete remission (CR). This patient had primary DLI failure and subsequently received ibrutinib for lowering of tumor burden, while DLI applications were continued. After having achieved CR upon simultaneous treatment with ibrutinib and DLI, the patient developed cGvHD and for the first time became MRD-negative.

Another patient had been transplanted in an ibrutinib-induced CR. After ibrutinib had been discontinued in the context of HSCT, an early relapse with rapidly rising leucocyte count ocurred 72d post HSCT. As a consequence, immunosuppression was tapered and ibrutinib treatment was resumed immediately after immunosuppression withdrawal, which resulted in a PR with normalized leucocyte count after 4 months.

Conclusion: This preliminary data suggest that ibrutinib is a safe and effective therapy for relapse of high-risk CLL after HSCT, even in patients pre-exposed before HSCT. Ibrutinib seems to allow for GvL activity and might serve as a priming therapy for DLI. Further studies are needed to decipher the immunologic interactions between ibrutinib and the allograft, and to analyze long-term outcome of patients treated with ibrutinib for relapsed high risk CLL.

Disclosure: No conflict of interest disclosed.

V295 - CLL - patient registries confirm bendamustine-containing regimen (BR) as an effective first-line therapy

Linde H.1, Göttel R.2, Tessen H.-W.3,4

1Medizinisches Versorgungszentrum für Blut- und Krebserkrankungen, Potsdam, Germany, 2rgb Onkologisches Management GmbH, Sarstedt, Germany, 3Onkologische Kooperation Harz, Goslar, Germany, 4Projektgruppe Internistische Onkologie (PIO), Goslar, Germany

Introduction: The chronic lymphocytic leukaemia (CLL) is the most common leukemic disease in Central Europe. The median age of onset is between 70 and 75 years. The combination of bendamustine and rituximab has proven effective for the treatment of this disease in clinical trials and everyday use.

Methods: Since 2008, 61 hemato-oncological practices from 16 federal states within the project team of internal oncology (PIO) have been documenting disease histories of patients with chronic lymphocytic leukaemia in the registry ONCOReg. 769 patients received a bendamustine-containing therapy, 525 (68.3%) as first-line therapy, 124 (23.6%) of which bendamustine mono and 401 (76.4%) bendamustine/rituximab (BR).

Results: This analysis presents the results of the use of bendamustine/rituximab in the first-line treatment of CLL patients in clinical practice.

Patient characteristics:

Gender: 66% m / 34% f

Median age: 71 years

Without B-symptoms: 61%

ECOG 0/1/2: 26% / 59% / 15%

BINET A/B/C: 11% / 55% / 34%

Period from initial diagnosis until first therapy: 22.3 months

Comorbidities: 37% hypertension, 19% diabetes; 10% CHD, 8% AIHA/ITP

Therapy: A median of 6 (1-8) cycles were administered. The median total dose of bendamustine was 880 mg/m².

Response: The objective remission rate is at 88.8%, 48% of which CR (haematologically tested) and 41% PR.


The median follow up is 24.3 months.

The median progression-free survival is at 44.5 months.

The median overall survival has not been reached yet. The 3-year survival rate is at 84%.

Conclusions: The analysis of the registry reflects the treatment routine and reveals the treatment of mostly older comorbid patients.

The therapy with bendamustine in combination with rituximab proves to be highly effective and safe. The remission rates and PFS of ONCOReg are comparable to those of other patients registries (TLN) or to clinical studies such as CLL2M or CLL10. Recent studies in which older and comorbid (“not fit”) patients were treated with newer substances do not show better results with regard to CR and PFS than the combination BR.

The results emphasize the high quality of the treatment of patients in the everyday routine and in specialized oncology practices.

Disclosure: Hartmut Linde: Expert Testimony: Honorare für Teilnahme an Studien (Firma Mundipharma, Roche, Universität Köln); Other Financial Relationships: Übernahme von Fortbildungs-, Reise- und Beherbergungskosten (Firma Mundipharma, Roche) Hans-Werner Tessen: No conflict of interest disclosed.


Ethik Handeln am Lebensende

V296 - Physicians´ practice at the end of life. Empirical data, ethical challenges

Schildmann J.1

1Ruhr-Universität Bochum, Institut für Medizinische Ethik und Geschichte der Medizin, Bochum, Germany

Introduction: Ethical questions at the end of life are at the centre of scientific and public debates. Up to the present there is scarcity of empirical data regarding physicians' end-of-life practices in Germany.

Methods: Cross-sectional study among a random sample of German physicians by means of the questionnaire of the EURELD Consortium with additional questions on experiences and attitudes regarding physician assisted suicide.

Results: 734 physicians from five state chambers of physicians responded (response rate 36.9%). 403 physicians reported about end-of-life practices regarding adult patients. Alleviation of symptoms took place in 86.7% of cases and in 50.7% medical treatment had been withheld. 30.8% of patients received continuous sedation till death. In three cases death was the consequence of a drug which was provided or administered by respondents. 20.7% of respondents had been requested to perform physician assisted suicide (PAS). 41.7% of participants could not imagine participating in PAS, whereas 40.2% could imagine this under certain circumstances. A prohibition of PAS by professional law war rejected by 33.7%, 25.0% support such a ban and 41.4% were undecided.

Conclusion: The published data will be presented together with data of an online survey among members of the German Society for Hematology/Oncology on their experiences and views regarding physician assisted suicide. The relevance of such data for the current societal and political debate and as starting point for reflections on ethical issues at the end of life in cancer care will be discussed.

Disclosure: No conflict of interest disclosed.

V298 - The role of physicians in the area of assisted suicide. Legal and medical ethical standards in Switzerland

Güth U.1

1Kantonsspital Winterthur, Department Geburtshilfe & Gynäkologie, Brustzentrum Senosuisse, Winterthur, Switzerland

Unlike in most European countries, physician-assisted suicide is not illegal in Switzerland, insofar as the actions of the person are not in self-interest (Art. 115 of the Swiss Penal Code). The number of assisted suicides aided by right-to-die organizations such as Exit or Dignitas has sharply increased in the last 20 years. Approximately half of the cases of assisted suicides are comprised of people with late stage cancer; the second most common reason for choosing assisted suicide is affliction with neuro-degenerative diseases (above all multiple sclerosis, amyotrophic lateral sclerosis and Parkinson's disease). According to current data, approximately 25% of the officially reported suicides in Switzerland are physician-assisted. Physician-assisted suicide is widely accepted in the Swiss society. In the canton of Zurich in 2011, a petition to ban assisted suicide was rejected by 85% of voters in a referendum.

The Schweizerische Akademie der Medizinischen Wissenschaft (SAMW) rejects assisted suicide as a physician`s duty, because it contradicts the basic principals of medicine. Nevertheless, it concedes that a doctor's conscientious and personal decision to perform assisted suicide for individual terminally-ill patients should be respected.

The central part of the doctor's involvement in assisted suicide is the prescription of a lethal dose of sodium pentobarbital. In doing so, the doctor has to apply to the rules of due medical care. A prescription can only be issued if the patient has been examined by the physician concerned. The patient must be informed about his diagnosis, about the expected prognosis, and about different alternative treatment options, in particular about the possibilities offered by palliative care. According to the SAMW, assisted suicide is only possible when the end of life is in the foreseeable future and the person is capable of making the decision willingly on his own without outside influences or pressure. A third party with the capability to assess whether these criteria have been met must confirm the decision before assisted suicide can legally be performed.

The practice of assisted suicide in Switzerland is characterized by the close cooperation between medical and non-medical personnel. In general, a staff member of the right-to-die organization, but not the doctor, is present during the suicide. Following death, the assisted suicide has to be reported to the authorities as a death under special circumstances.

Disclosure: No conflict of interest disclosed.

Freier Vortrag

Lymphome aggressiv experimentell

V299 - USP9X promotes resistance to spindle poisons in aggressive B-cell lymphoma by regulating the mitotic cell fate decision

Engel K.1, Rudelius M.2, Altmann B.3, Abhari B.A.4, Brunner A.1, Kurutz J.1, Targosz B.-S.1, Loewecke F.1, Knorn A.-M.1, Fernandez-Sáiz V.1, Baumann U.1, Glöckner J.5, Pfreundschuh M.6, Trümper L.7, Jost P.1, Klapper W.8, Fulda S.4, Peschel C.1, Bassermann F.1

1III. Medizinische Klinik, Klinikum rechts der Isar, Technische Universität, München, Germany, 2Pathologisches Institut/Universität, Würzburg, Germany, 3IMISE, Leipzig, Germany, 4Institut für Experimentelle Tumorforschung in der Pädiatrie, Frankfurt, Germany, 5DZNE, Tübingen, Germany, 6Klinik für Innere Medizin I, Universitätsklinikum des Saarlandes, Homburg, Germany, 7Klinik für Hämatologie und Medizinische Onkologie, Göttingen, Germany, 8Institut für Pathologie, Kiel, Germany

Introduction: Despite advances in the first line therapy, a significant number of patients with aggressive B-cell lymphoma succumb to relapsing or refractory disease. The mitotic spindle assembly checkpoint (SAC) is an important target of lymphoma therapy as it regulates resistance to spindle poisons. To date it has remained unclear how cells execute cell fate decisions under conditions of SAC induced mitotic arrest.

Methods: Binding and stability assays were performed using immune precipitation, induced expression or siRNA-mediated knockdown. Ubiquitylation was assessed under denaturing conditions. Analysis of cells synchronized with thymidine and nocodazol or paclitaxel was supported by two dimensional cell cycle analysis using FACS (BrdU, PI). BL57 mice were injected with lentivirally transduced Eµ-myc lymphoma cells. Patient samples were obtained from the German High Grade Non Hodgkin's Lymphoma Study Group (DSHNHL, Ricover-60 Trial) and analysed by immunohistochemistry.

Results: We identify USP9X as the mitotic deubiquitinase of the X-linked inhibitor of apoptosis protein (XIAP), and demonstrate that deubiquitylation and stabilization of XIAP by USP9X induces resistance towards mitotic spindle poisons[FB1] . Mitotic cell death in response to prolonged taxol exposure is increased by USP9X knockdown in WT, cIAP2 k.o. and MCL-1 k.o. but not in XIAP k.o. MEFs, indicating that XIAP marks the biologically relevant target of USP9X. Mapping experiments delineate the USP9X binding site in XIAP to the Gly188 residue. Accordingly, overexpression of USP9X and XIAP, but not of instable XIAP(G188) mutants, promote cell survival despite taxol exposure. USP9X-XIAP overexpressing DLBCL lines exhibit increased resistance to spindle poisons and undergo substantial apoptosis in response to USP9X knockdown or pharmacological inhibition and treatment with the XIAP inhibiting SMAC mimetic BV6. Likewise, knockdown of USP9X or XIAP delays lymphoma development in a murine Eµ-myc-lymphoma model. In 121 human DLBCL samples high expression of USP9X significantly correlated with XIAP overexpression (p = 0.004) and overexpression of USP9X and XIAP was associated with significantly reduced event free survival in CHOP treated DLBCL patients (p = 0.05).

Conclusion: Antiapoptotic signaling via the USP9X-XIAP axis mediates resistance to mitotic cell death. USP9X-XIAP expression may serve as an attractive novel prognostic and therapeutic target in aggressive B-cell lymphoma.

Disclosure: Katharina Engel: No conflict of interest disclosed. Florian Bassermann: Expert Testimony: Celgene.

V300 - Vitamin D promotes tumoricidal activity of macrophages and improves the efficacy of antibody-dependent cellular cytotoxicity

Bruns H.1, Büttner M.2, Mougiakakos D.2, Bittenbring J.3, Beier F.4, Pasemann S.2, Fabri M.5, Neumann F.3, Mackensen A.2, Gerbitz A.2

1Uniklinikum Erlangen, Dept. of Internal Medicine 5 - Hematology/Oncology, University of Erlangen, Erlangen, Germany, 2Uniklinikum, Erlangen, Germany, 3Universitätsklinikum des Saarlandes, Homburg, Germany, 4Uniklinik, Aachen, Germany, 5Uniklinik, Köln, Germany

Infiltration by macrophages represents a characteristic morphological hallmark in high-grade lymphatic malignancies such as Burkitt's lymphoma (BL). Although macrophages can, in principle, target neoplastic cells and mediate antibody-dependent cytotoxicity (ADCC), tumor-associated macrophages (TAMs) regularly fail to exert direct cytotoxic functions. The underlying mechanisms responsible for this observation remain unclear. Here, we demonstrate that inflammatory M1 macrophages kill proliferating high-grade B cell lymphoma cells by releasing the antimicrobial peptide cathelicidin in a vitamin D-dependent fashion. We show that cathelicidin directly induces cell death by targeting mitochondria of BL cells. In contrast, anti-inflammatory M2 macrophages and M2-like TAMs in BL exhibit an altered vitamin D metabolism, resulting in a reduced production of cathelicidin and consequently in inability to lyse BL cells. However, treatment of M2 macrophages with the bioactive form of vitamin D, 1,25D3, or a vitamin D receptor agonist effectively induces cathelicidin production and triggers tumoricidal activity against BL cells. Furthermore, rituximab-mediated cytotoxicity of vitamin D-treated M2 macrophages is cathelicidin-dependent. Finally, vitamin D treatment of 25D-deficient volunteers in vivo or primary TAMs in vitro improves rituximab-mediated ADCC against B cell lymphoma cells. These data indicate that activation of the vitamin D signaling pathway activates antitumor activity of TAMs and improves the efficacy of ADCC.

Disclosure: No conflict of interest disclosed.

V301 - Arylindolylmalemide PDA-66 and Its derivate PDA-377 show antiproliferative effects on B-cell non-Hodgkin lymphoma cell lines with MYC and BCL rearrangements

van der Wall K.1, Roolf C.1, Sklarz L.-M.1, Eschenburg A.1, Sekora A.1, Murua Escobar H.1, Rolfs A.2,3, Pews-Davtyan A.4, Beller M.4, Junghanss C.1

1Universitätsmedizin Rostock, Klinik III, Hämatologie, Onkologie, Palliativmedizin, Rostock, Germany, 2Universitätsmedizin Rostock, Albrecht-Kossel-Institut für Neuroregeneration, Rostock, Germany, 3Centogene AG, Rostock, Germany, 4Universität Rostock, Leibniz-Institut für Katalyse, Rostock, Germany

Introduction: Diffuse large B-cell lymphoma (DLBCL) as well as Burkitt`s lymphoma are a frequent subtypes of non-Hodgkin´s lymphoma (NHL). Both subtypes usually are characterized by short cell doubling times making them susceptible to cell cycle modifying agents. Previous own results showed that the novel arylindolmaleimide PDA-66 inhibits the polymerisation of microtubules inducing antiproliferative effects in acute lymphoblastic leukemia cells. Here, we evaluated the potency of PDA-66 and its derivate PDA-377 on B-cell NHL cell lines.

Methods: Burkitt lymphoma and DLBCL cell lines with BCL2 and MYC rearrangements (DOGKIT; SU-DHL-4 and WILL-2) were incubated for 72 h with increasing concentrations ranging from 1 to 5 µM of PDA-66 and PDA-377, respectively. Metabolic activity was determined by WST-1 assay. Furthermore, PDA-66 was combined with low dose of AraC, Doxorubicin (Doxo) or Dexamethasone (Dexa) in different sequences (PDA before, concomitant, after 2nd drug). Changes in cell cycle and apoptosis were analyzed flow cytometricaly by propidium iodide and annexin staining.

Results: In all cell lines PDA-66 and PDA-377 inhibited proliferation significantly in a dose dependent manner. Strongest antiproliferative effects were detected with PDA-66. Incubation with 5 µM PDA-66 decreased metabolic activity to 22.4% in WILL-2, 47.1% in SU-DHL-4 and 43.3% in DOGKIT cells. Further, treatment with PDA-66 resulted in an increase of cells arresting in G2/M phase of the cell cycle (41.4% vs. control: 14.1%). This effect was enhanced when PDA-66 and Dexa were combined together (68.9% in G2/M) even when cells were less sensitive to single Dexa. Interestingly, the sequence of PDA-66 and Dexa application to DBCL cells had an impact on the antiproliferative effects. A 24 h pre-treatment with PDA-66 (2.5 µM) before adding Dexa (50 µM) increased antiproliferative effects compared to other sequences.

Conclusion: Treatment with PDA-66 and PDA-377 induced antiproliferative effects on B-NHL harbouring BCL2 or MYC gene rearrangements. Moreover, PDA-66 seems to sensitize DLBCL cells to Dexa.

Disclosure: No conflict of interest disclosed.

V302 - Identification of a N-hyperglycosylated domain of CNS proteins as B cell receptor antigen in primary CNS lymphoma

Thurner L.1, Kemele M.1, Fadle N.1, Regitz E.1, Roth P.2, Weller M.2, Schäfer H.3, Schorb E.3, Illerhaus G.3, Szczepanowski M.4, Klapper W.4, Monoranu C.-M.5, Rosenwald A.5, Buslei R.6, Bohle R.M.7, Preuss K.-D.1, Pfreundschuh M.1

1Saarland University Medical School, José Carreras Center for Immuno- and Gene Therapy and Internal Medicine, Homburg/Saar, Germany, 2Department of Neurology, University Hospital, Zürich, Switzerland, 3Department of Hematology and Oncology, University Medical Center, Freiburg, Germany, 4Department of Pathology, Hematopathology Section and Lymph Node Registry, University of Kiel, Kiel, Germany, 5Institute of Pathology, University of Würzburg, Würzburg, Germany, 6Institute of Neuropathology, University Hospital Erlangen-Nürnberg, Erlangen, Germany, 7Saarland University Medical School, Institute of Pathology and Neuropathology, Homburg/Saar, Germany

Background: A reported VH4-34 bias in PCNSL has raised speculations of a possible chronic stimulation of BCR by a CNS-autoantigen. The present study focused on the search for these hypothetic autoantigens in immunocompetent patients.

Methods: Recombinant Fabs were expressed based on corresponding pairs of functional variable region heavy and light chain genes, which had been amplified from snap-frozen lymphoma specimens by semi-nested PCRs. Subsequently these rec. Fabs were screened on protein arrays.

Results: Rec. Fab expression was attempted in 21 and successful in twelve PCNSL cases. Protein array screening of the rec. Fabs revealed that 8 of 12 rec. PCNSL-BCRs reacted with SAMD14 and the SAM domain of neurabin-1, two proteins with high homology and preferential expression in the CNS. Proteomic analysis of lymphoma specimens showed that SAMD14 and the SAM domain of neurabin-1 were alternatively N-glycosylated in patients with a Fab-specificity against SAMD14/neurabin-1, but not in the remaining cases with other BCR specificities. Atypical (N-L-E-Q) instead of (N-X-S/T) N-hyperglycosylations of SAMD14/neurabin-1 were shown for every case with sufficient biopsy material for this proteomic analysis and a PCNSL-BCR specific for SAMD14/ neurabin-1. Of the rec. BCRs of all cases with sufficient material to test for hyperglycosylation, only the rec. Fabs derived of the cases with hyperglycosylated SAMD14/neurabin-1 reacted against SAMD14/neurabin-1. No N-hyperglycosylations of SAMD14/ neurabin-1 were neither detected in the peripheral blood of 400 healthy controls, 100 newborns, 50 nursery home residents nor in 86 established cell lines of various cellular origin. Moreover, antibodies against SAMD14/neurabin-1 were detected in the sera and cerebrospinal fluids of an independent second cohort of patients with PCNSL (8/22), but not in sera of patients with secondary CNS manifestations of systemic DLBCL (0/17) or of healthy controls (0/92).

Conclusion: Our results suggest that atypical (NLEQ) glycosylation of the highly homologous SAM domain of SAMD14 and neurabin-1 maintains a chronic autoantigenic stimulation in the CNS, ultimately leading to the malignant transformation of B-cells with a BCR specific for these atypically N-hyperglycosylated proteins into an aggressive B-cell lymphoma in the CNS, providing strong evidence for the role of chronic autoantigenic stimulation as a first step in the pathogenesis of aggressive B-cell lymphomas.

Disclosure: No conflict of interest disclosed.

V303 - Functional investigation of putative genetic lesions on the CNS tropism of diffuse large B-cell lymphomas in vivo

Reimann M.1, Maßwig S.1, Schleich K.1, Herrmann A.1, Lohneis P.2, Schrezenmeier J.F.1, Dörken B.1,3, Schmitt C.1,3

1Charité - Universitätsmedizin Berlin, Medical Department of Hematology, Oncology and Tumor Immunology and Molekulares Krebsforschungszentrum, Berlin, Germany, 2Charité - Universitätsmedizin Berlin, Institute of Pathology, Berlin, Germany, 3Max-Delbrück-Center for Molecular Medicine, Berlin, Germany

Introduction: Secondary central nervous system (CNS) lymphomas impose a dismal prognosis in patients suffering from systemic diffuse large B-cell lymphomas (DLBCL). Myc rearrangement, chromosomal deletions at the CDKN2A (a.k.a. INK4a/ARF) or ATM gene loci as well as recurrent NF-kB-hyperactivating mutations are frequent in CNS-tropic lymphoma, however, investigations that functionally validate these lesions in adequate in vivo model systems are missing.

Methods: We generated primary Eµ-myc transgenic lymphomas with distinct naturally occurring NF-kB mutations (within genes encoding for MyD88, CD79B, A20, IkBζ, IkBε or BIRC3) or deletions at the INK4a/ARF and ATM loci. Wild-type recipient mice were i.v. transplanted with these lymphoma cells, and subsequently monitored for systemic lymphoma development, at which the brain was isolated and examined regarding lymphoma infiltration.

Results: Underscoring Myc's role as a putative co-driver of CNS involvement, we found in about 40% of primary Eµ-myc lymphomas (with no additional exogenous lesions) meningeal CNS tropism. Genome-wide transcriptome analysis unveiled NF-kB hyperactivation in the CNS-tropic lymphoma group, suggesting that NF-kB-activating mutations promote CNS-prone pathogenesis in vivo. Transplantation of Eµ-myc hematopoietic stem cells expressing a variety of NF-kB-activating mutants resulted in a significant acceleration of Eµ-myc-driven lymphomagenesis, with some, but not all of these mutants conferring a CNS-tropic lymphoma phenotype. Global NF-kB suprression in CNS-tropic Eµ-myc lymphomas via the NF-kB-antagonizing IkBΔN super-repressor did not fully abrogate lymphoma infiltration of the brain, suggesting that additional factor(s) must contribute. Importantly, targeted ablation of the INK4a/ARF and ATM loci robustly enhanced CNS tropism of Eµ-myc lymphomas.

Conclusions: The Eµ-myc mouse lymphoma model is well-suited to genetically dissect and rebuild components of CNS tropism. We identified CDKN2A or ATM deletions as critical determinants of CNS tropism in vivo. Our systematic analyses of different NF-kB mutants - so far rather recognized as functionally interchangeable - indicated that only distinct NF-kB mutants contribute to CNS tropism in B-cell lymphomas. Our findings underscore the need for functional analyses of oncogenic network contexts, and provide important insights into candidate target lesions for personalized CNS-directed therapies in DLBCL patients in the future.

Disclosure: No conflict of interest disclosed.

V304 - A modified autoantigen is the first molecularly defined risk factor and a dominant antigenic target / stimulus of the B-cell receptor from ABC-type DLBCL

Pfreundschuh M.1, Preuss K.-D.1, Fadle N.1, Regitz E.1, Kemele M.1, Bohle R.-M.2, Hansmann M.-L.3, Thurner L.1

1Universität des Saarlandes, Innere Medizin I, Homburg/Saar, Germany, 2Universität des Saarlandes, Institut für Allgemeine und Spezielle Pathologie, Homburg/Saar, Germany, 3Universitätsklinikum Frankfurt, Dr. Senckenbergisches Institut für Pathologie, Frankfurt am Main, Germany

Introduction: Chronic antigenic stimulation may play an important role in the pathogenesis of malignant lymphomas. We have previously shown that hyperglycosylated neurabin/SAMD14 is the antigenic target of the B-cell receptor (BCR) of 2/3 of all primary CNS lymphomas, but BCR for peripheral DLBCL have not been defined to date.

Methods: BCRs were expressed as recombinant Fabs based on corresponding pairs of functional variable region heavy and light chain genes, which had been amplified from isolated genomic DNA of snap-frozen lymphoma specimens and DLBCL-derived cell lines. The purified BCR-Fabs were checked for binding to proteins expressed on macroarrays of human cDNA expression libraries.

Results: The BCR from 10 DLBCL cell lines (5 of the germinal center type and 5 of the activated B-cell type) were tested on the protein macroarray. None of the GC-type BCR reacted with any of the proteins expressed on the protein macroarray, but the BCR from 3/5 (60%) of the ABC-derived cell lines reacted with ARS2 (arsenite resistance protein 2), a conserved mammalian protein which is important for microRNA biogenesis. Isoelectric focusing and phosphatase treatment of ARS2 derived from ABC cell lines with a BCR specific for ARS2 revealed that ARS2 was hypophosphorylated (hypo-ARS2) in the respective cell lines. Analysis of peripheral blood lymphocytes from patients with DLBCL of unknown cell of origin and healthy controls revealed that 5/100 (5%) of patients, but only 1/400 (0.25%) of controls were carriers of hypo-ARS2, resulting in a 20x increased risk for healthy carriers of hypo-ARS2 to develop DLBCL. All patients with BCRs targeting ARS2 had polyclonal antibodies against ARS2 in their serum.

Conclusions: Hypo-ARS2 is the first molecularly defined risk factor for DLBCL identified to date. The increased risk for healthy carriers of this posttranslational modification to develop DLBCL supports the hypothesis of chronic antigenic stimulation as an important factor in the pathogenesis of DLBCL and indicates that posttranslationally modified autoantigens are a frequent target and stimulus for DLBCL-BCR. That antibodies against ARS2 are found in the respective patients suggests that the DLBCL evolves from a polyclonal B-cell response against this autoantigen. Investigations into the mechanism underlying the hypophosphorylation of ARS2 are underway and therapeutic consequences will be discussed.

Supported by Wilhelm-Sander-Stiftung.

Disclosure: No conflict of interest disclosed.

Freier Vortrag

Tumor-/Zellbiologie I

V305 - The tumor-supporting role of oxidative and ER stress tolerance: New lessons from fibroblasts and cancer cells to understand tumor biology and chemoresistance

Venkataramani V.1, Rumkamp T.1, Pape V.1, Kiecke C.1, Küffer S.2, Ströbel P.2, Trümper L.1, Wulf G.G.1

1University Medicine Goettingen, Department of Hematology and Oncology, Göttingen, Germany, 2University Medicine Goettingen, Department of Pathology, Göttingen, Germany

Introduction: Conversely to untransformed cells, where deregulated cellular stress conditions drive the activation of death pathways, malignant cells exploit oxidative milieu for its advantage. Therefore, resistance to oxidative stress appears to be a major mechanism of cancer initiation/propagation and chemoresistance. In this regard, we have identified a novel role for the redox modulator amyloid precursor protein (APP) in effectively modulating the stress tolerance of cancer cells. We previously established that APP is overexpressed in several solid and haematological cancer types and underlined the function in the cell proliferation, colony formation and propagation of cancer stem cells. Now, we show that downregulation of APP in cancer cells results in a feed-forward loop with oxidative stress and its induction of endoplasmic reticulum stress (ER stress). The resulting selective DNA damage response (DDR) impacts growth of cancer cells/transformed fibroblasts and confers sensitization especially to genotoxic chemotherapeutics agents.

Methods: Murine embryonal fibroblasts (MEFs) from wild-type (APP+/+) and APP-/- (APP-knockout) mice were isolated and transformed with Ha-RasV12 via adenoviral transfer. Functional assays such as cell proliferation, colony forming assays, RNA seq, and western blot as well as in ovo analysis via CAM-Assay were performed. Intracellular ROS and lipidperoxidation levels were determined via DCFH-DA dye and BODIPY 581⁄591 C11 dye, respectively.

Results: Consistent with our hypothesis that APP modulates intracellular iron homeostasis, we show that loss-of-APP either in cancer cells or APP-/- MEFs contain higher ROS and lipidperoxidation levels. Beside oxidative stress, we provide direct genetic evidence that APP causes cell death in a pathologically relevant form leading to ER stress inducing DNA damage. Novel selective APP-Inhibitors also induced ROS and ER-stress causing activation of DDR without inducing apoptosis or autophagy. By dissecting the molecular hallmark events, we present evidence that loss-of-APP suppresses tumorigenicity of transformed fibroblasts and reveal that APP-induced alteration of the DNA damage-signaling effectively re-sensitize several cancer types to various cancer therapies, including DNA damaging agents.

Conclusion: Here, we identify APP as regulator of oxidative and ER stress selectively modulating DNA damage signaling representing an effective drug target for enhanced cancer therapy.

Disclosure: No conflict of interest disclosed.

V306 - Inhibition of PARP selectively sensitizes KRAS-mutant cancer cells to chemotherapy in vivo - an update

Hähnel P.S.1, Sasca D.1, Enders B.1, Lehmann N.1, Roos W.P.2, Kaina B.2, Theobald M.1, Kindler T.1

1University Medical Center of the Johannes Gutenberg-University, Third Department of Medicine, Division of Hematology, Medical Oncology & Pneumology, Mainz, Germany, 2University Medical Center of the Johannes Gutenberg-University, Institute of Toxicology, Mainz, Germany

Introduction: Activating KRAS mutations are detected in a substantial number of different malignancies and often appear during early stages of tumorigenesis. KRAS-mutations are already found in patients suffering from colorectal adenomas or pancreatic ductal hyperplasias. Full malignant transformation is thought to be triggered by additional mutations. In preliminary work we demonstrated that KRAS-mutated cells rely on the alternative non homologeous end-joining (alt-NHEJ) repair pathway upon genotoxic stress, while KRAS wild-type cells do not. RNAi-mediated down regulation of the alt-NHEJ component DNA ligase 3α abolished drug-resistance to apoptosis in KRAS-mutant cells in vitro. In vivo, treatment with the PARP-inhibitor Olaparib sensitized specifically KRAS-mutated cells to chemotherapeutic agents. Combination therapy with Irinotecan or VP-16 was able to control tumor growth in NSG mice transplanted with colon cancer and NSCLC cells, respectively.

Methods: The sensitivity to drug-induced apoptosis of different colon cancer and NSCLC cell lines with either mutated or wild-type KRAS was analyzed by molecular assays and FACS analysis. Cell viability and cytotoxicity were determined via cell proliferation assays and cell cycle analysis. In vivo, the antitumor activity of Olaparib in combination with different genotoxic agents was assessed in mice bearing tumor xenografts.

Results: Pharmacologic inhibition of PARP, which has been implicated in promoting end joining by alt-NHEJ, upon treatment with Olaparib caused increased apoptosis induced by standard genotoxic agents compared to controls. Mice bearing tumor xenografts either from colon cancer or NSCLC cell lines showed a diminished tumor growth compared to animals treated with monotherapy, finally resulting in longer overall survival. Interestingly, the reversal of genotoxic drug resistance upon inhibition of the alt-NHEJ pathway seems to be specific for KRAS-mutant cells, as no effect was observed in tumors derived from KRAS-wild-type cell lines.

Conclusion: Our data provide evidence for a synthetic lethal interaction between KRAS-mutations and DNA damage repair. Furthermore, they indicate that targeting components of the alt-NHEJ pathway, e.g., PARP, sensitizes KRAS-mutant cancer cells to standard chemotherapeutics and represents a promising approach for inducing synthetic lethal vulnerability in cells harboring otherwise non-druggable KRAS mutations.

Disclosure: No conflict of interest disclosed.

V307 - Loss of the tumor- and metastasis suppressor RAF kinase inhibitor protein is caused by the overexpression of miRNA-23a

Hatzl S.1, Geiger O.1, Kuepper M.K.1, Seime T.2, Nußbaumer E.1, Wieser R.3, Pichler M.4, Scheideler M.5,6,7, Nowek K.8, Jongen-Lavrencic M.8, Wölfler A.1, Troppmair J.2, Sill H.1, Zebisch A.1

1Division of Hematology, Medical University of Graz, Graz, Austria, 2Daniel Swarovski Research Laboratory, Innsbruck Medical University, Innsbruck, Austria, 3Department of Medicine I, Medical University of Vienna, and Comprehensive Cancer Center of the Medical University of Vienna, Vienna, Austria, 4Division of Oncology, Medical University of Graz, Graz, Austria, 5Institute for Diabetes and Cancer, Helmholtz Center Munich, Neuherberg, Germany, 6German Center for Diabetes Research (DZD), Neuherberg, Germany, 7University Hospital, Heidelberg, Germany, 8Department of Hematology, Erasmus University Medical Center, Rotterdam, Netherlands

Introduction: RAF kinase inhibitor protein (RKIP) is a negative regulator of MAPK/ERK signaling and acts as a tumor- and metastasis suppressor. A complete or partial loss of RKIP expression has been observed in several human malignancies, including acute myeloid leukemia (AML). As the mechanisms leading to RKIP loss are still unclear, we analyzed the potential involvement of miRNAs within this process.

Methods: RKIP protein expression was assessed in 33 AML specimens, which had been characterized by miRNA microarrays previously (Rommer et al., BMC Cancer 2013). Chip data of miRNAs related to RKIP expression were further validated by qPCR. U937 cells were transduced with shRNA or overexpression constructs for RKIP using lentiviral transduction. miRNA mimics, as well as an RKIP 3'UTR Luciferase reporter plasmid were transfected by lipofection (in HEK-293) and nucleofection (in NB-4), respectively. qPCR and Western blot were used to monitor the expression of RKIP and miRNAs. Correlation between miRNA-23a and RKIP expression was further corroborated in mRNA and miRNA microarray data from an independent AML cohort (n = 214), as well as in a database retrieval using The Cancer and Genome Atlas (TCGA; AML, n = 173; invasive breast carcinoma, n = 283; renal clear cell carcinoma, n = 171; ovarian serous cystadenocarcinoma, n = 516).

Results: By combining RKIP expression with both, miRNA microarray and qPCR data, we identified a set of five miRNAs showing increased expression levels in AML specimens with RKIP loss. Increased expression of miRNA-23a could be corroborated in two independent AML cohorts, comprising almost 400 patients. In functional experiments, modulation of RKIP expression failed to influence miRNA-23a, thereby excluding that miRNA-23a overexpression is an effect of RKIP loss. On the contrary, overexpression of miRNA-23a by miRNA-mimics markedly decreased RKIP mRNA and protein expression, indicating that it is functionally involved in the downregulation of this tumor-suppressor. Preliminary results with an RKIP 3'UTR Luciferase reporter clone suggest, that this effect is mediated by binding of miRNA-23a to this region. Finally, a database retrieval comprising almost 1000 patients with other cancer entities also showing RKIP loss, further corroborated association of miRNA-23a overexpression with decreased RKIP expression, thereby supporting the relevance of this finding.

Conclusions: Our data have identified miRNA-23a as a negative regulator of RKIP expression.

Disclosure: No conflict of interest disclosed.

V308 - Very early functional in-vivo imaging to predict response to B cell receptor pathway targeting in lymphoma

Habringer S.1,2, Li Z.2, Pietschmann E.2, Slawska J.2, Walch A.3, Peschel C.2,4, Keller U.1,2

1DKFZ/DKTK, Partnerstandort München, München, Germany, 2Technische Universität München, III. Medizinische Klinik, München, Germany, 3Helmholtz Zentrum, München, Germany, 4DKFZ/DKTK, Partnerstandort München, Heidelberg, Germany

B cell receptor (BCR) signaling plays a pivotal role for disease development and progression in both activated B cell (ABC) and germinal center B cell (GCB) diffuse large B cell lymphoma (DLBCL). However, there is evidence that ABC DLBCL primarily depends on chronic active BCR signaling with strong NFκB activation, whereas GCB-DLBCL is characterized by tonic BCR signaling, leading to PI3K activation. These differences in BCR signaling impact the response of ABC and GCB DLBCL to kinase inhibitors affecting BCR pathway components.

We assessed response to ibrutinib and idelalisib in ABC and GCB cell lines harboring different mutations in the BCR pathway in-vitro and in a xenograft mouse model in-vivo and evaluated the ability of functional imaging technologies to predict the response to these inhibitors.

Both idelalisib and ibrutinib inhibited proliferation in the CD79A mutated ABC cell line OCI-Ly10 in-vitro, but had no effect on the TAK1 mutated ABC cell line U2932 and the GCB cell line SUDHL-6, as was predicted by the mutational status of BCR components in these cells. To further analyze if response to BCR signaling inhibitors could be imaged in vivo, we xenotransplanted OCI-Ly10, SUDHL-6 and U2932 cells subcutaneously into SCID mice and administered both inhibitors orally after successful engraftment of lymphoma cells had occurred. To monitor response to therapy, mice were imaged with FDG- and FLT-PET immediately before and 48 hours after initiation of therapy. Both inhibitors induced rapid reduction in tumor volumes of OCI-Ly10 and, surprisingly, U2932 lymphomas, but had no effect on SUDHL-6 tumors, which indicated that mutational status of BCR signaling components alone is not sufficient to predict response to BTK and PI3K inhibition in this in-vivo model. Finally, MALDI imaging was performed on xenograft lymphomas to assess patterns of proteomic changes associated with response or resistance to BCR pathway inhibition. In OCI-Ly10 lymphomas, we found a signature of proteomic changes induced by ibrutinib that separated treated and untreated lymphomas in PCA-clustering.

We found that mutational status of BCR components was sufficient to predict response of OCI-Ly10, U2937 and SUDHL-6 to ibrutinib and idelalisib in-vitro, but not in-vivo. MALDI-imaging to detect changes in the proteome could serve as a potential method to predict response or resistance to BCR pathway inhibition early after initiation of therapy.

Disclosure: No conflict of interest disclosed.

V309 - Multi-molecular complexes of Bcl-2 proteins regulate membrane permeabilization by Bax

Bogner C.1,2, Kale J.2, Chi X.2, Leber B.2, Andrews D.2

1III. Medizinische Klinik Klinikum rechts der Isar, Hämatologie/Onkologie, München, Germany, 2Sunnybrook Research Institute, Biological Sciences, Toronto, Canada

The core event in apoptosis is the mitochondrial outer membrane permeabilization (MOMP), the “point of no return” in the commitment to cell death. MOMP is regulated by the tight interplay of members of the Bcl-2 family of proteins, which play the central role in this crucial step. The interactions and balance between pro- and anti-apoptotic members of this family ensures the fine-tuning of life or death decisions at the outer mitochondrial membrane (OMM). Pro-survival members sequester activator BH3 proteins or executioners to prevent pore formation. Sensitizer proteins sequester and inhibit the pro-survival members and promote MOMP, thereby getting sequestered themselves to limit their impact. Changes for the equilibria of these mutual sequestrations and inhibitions within dynamic complexes of Bcl-2 proteins define the decision for survival or death at the OMM. In our study we use fluorescence spectroscopy techniques to analyze these complex interactions of anti-apoptotic Bcl-XL with pro-apoptotic Bax and Bid and Bad in solution, in an in vitro membrane system modeling the OMM and in isolated mitochondria. Our findings show that the interplay of pro-survival Bcl-XL with the pro-apoptotic counterparts is not limited to heterodimeric interactions but involves formation of higher order complexes that are crucial in the regulation of membrane permeabilization. As this key step of apoptosis is embedded in the mitochondrial membrane, the finding of new membrane associated complexes of Bcl-2 family proteins will extent our tools for the development of powerful therapeutics, which will target were it matters, in the membrane.

Disclosure: No conflict of interest disclosed.

V310 - Endothelial cell derived micro-vesicular proteins induce breast cancer cell migration

Ferraro D.1, Goosen R.1, Zanivan S.2, Patella F.2, Christofori G.1, Buess M.3

1Department Biomedizin Universität, Basel, Switzerland, 2Beatson Institute for Cancer Research, Glasgow, United Kingdom, 3St. Claraspital, Basel, Switzerland

Introduction: The microenvironment is a central regulator of tumor biology. While the contribution of fibroblasts has been largely studied, the role of endothelial cells as regulators of cancer cell behavior is still poorly understood. As in a diverse spectrum of physiological processes in normal tissue, endothelial cells may exert a similar regulatory control in malignant cancer progression and metastasis, not only contributing to vessel formation, but also through endothelial cell specific signaling.

Methods: To characterize the functional effects of endothelial-cancer interaction we focused on an in vitro model of SKBR3 breast cancer cells and human endothelial cells.

Results: SKBR3 cells treated with HUVEC derived supernatant show significantly increased migratory potential, without a parallel increase in proliferation, an elongated phenotype and expression of mesenchymal markers (up-regulation of FN1, Stress Fibers and Focal Adhesion formation). The pro-migratory effect is significantly more pronounced when the supernatant is obtained from a sparse and highly proliferative endothelial cell culture than from confluent and resting endothelial cells. To better understand the differential regulation on cancer cell migration, we analyzed the supernatant of sparse or dense endothelial cells by quantitative MS proteomics (SILAC analysis). Interestingly, extracellular matrix proteins were enriched in dense endothelium supernatant. Amongst them, Biglycan reduced the pro-migratory effect of treatment with sparse endothelium supernatant, suggesting a potential role of resting endothelium as an inhibitor of cancer cell migration. The proteomic analysis of sparse endothelial cell supernatant revealed an enrichment in proteins (e.g. rap-1, mmp1, annexinA2) belonging to the micro-vesicular compartment. Knocking down these proteins significantly reduced the pro-migratory effect of the endothelial supernatant on cancer cells, demonstrating that the microvesicular compartment can play a role in the modulation of tumor aggressiveness.

Conclusions: We suggest that identification of the endothelial cell´s role in cancer progression independent from vessel formation could reveal targets for novel therapeutic strategies.

Disclosure: No conflict of interest disclosed.


Hepatische Tumore

V313 - Biliray tract cancer: Adjuvant treatment - pros and cons

Stein A.1

1Universitätsklinikum Hamburg-Eppendorf, Hubertus Wald Tumorzentrum II. Medizinische Klinik und Poliklinik, Hamburg, Germany

The treatment of biliary tract cancer has made relevant progress during the last years with improved understanding of disease biology (e.g. detection of proliferation and inflammation subtypes in intrahepatic cholangiocarcinoma) or development of highly active and well tolerated systemic treatment for advanced disease stages (gemcitabine and cisplatin).

Despite decades of intense discussion wether and how to administer adjuvant treatment in biliary tract cancer available data are highly limited. Only two underpowered randomized trials about adjuvant chemotherapy are currently available showing either no benefit or a benefit limited to gallbladder carcinoma with a non contemporary chemotherapy-regimen containing mitomycin and 5FU. Pooled observational data point towards a benefit particularly in lymph node positive disease for chemotherapy or chemoradiation. Currently, several randomized trials have completed accrual (BILCAP: observation ± capecitabine; ACCORD18 - PRODIGE 12: observation ± gemcitabine and oxaliplatin) or are ongoing (ACTICCA 1: observation ± gemcitabine and cisplatin), which will likely solve the current dilemma about administration of adjuvant chemotherapy.

Disclosure: No conflict of interest disclosed.

V314 - Metastatic liver disease: Review on different entities and different loco-regional approaches

Wöll E.1

1St. Vinzenz Krankenhaus Zams, Innere Medizin, Zams, Austria

Surgery of hepatic metastases is well established in colorectal cancer if complete resection can be achieved and liver function can be obtained. Long term results show a curative potential for certain patients after complete resection with clear margins. Ablative techniques and stereotactic radiotherapy can be combined with resection in some cases or can substitute for resection. Optimal indication and patient selection for the different approaches however are still unclear. Surgery has the advantage of complete histological workup including information on resection margins. Different ablation techniques and radiotherapy however are less invasive and post intervention morbidity might be lower. Even if combined with modern MRI imaging techniques however exact information on circumferential margins after ablation or radiotherapy can not be given.

During the last decades surgery of liver metastases was performed not only in colorectal cancer and neuroendocrine cancer but also in melanoma, sarcoma, breast cancer, gastric cancer, and several other tumor entities. The role of local treatment of liver metastases in non colorectal, non neuroendocrine metastatic liver disease and the optimal approach, although feasible and safe, is not well defined yet.

Disclosure: No conflict of interest disclosed.


Stem Cell Biology

V316 - Quantitative approaches to single cell analysis: An update

Etzrodt M.1

1ETH Zurich, D-BSSE, Basel, Switzerland

Understanding the molecular processes that govern cell fates is essential to study the biology of normal and malignant hematopoiesis. This requires quantification of molecular and cellular behavior at the single-cell level, because bulk readouts can mask the inherent heterogeneity of the populations studied. Recent advances now permit high- throughput molecular readouts from single cells as well as continuous, noninvasive observation of cell behavior over time. Likewise new tools for single cell handling, sample preparation and culture, based on microfluidic technologies facilitate the execution of complex experiments with single cell precision, in an automated fashion and at reduced cost.

Each of the available approaches has specific advantages and limitations. Most molecular profiling approaches for instance provide only snapshots of a given cellular state and represent end-points, which prevents to link a molecular readout to future fates of the cells investigated. The knowledge of future fates is a prerequisite though when studying how a specific molecular signature influences the differentiation of a single normal or leukemic transformed hematopoietic stem or progenitor cell (HSPC). Consequently for many questions in the field the continuous, noninvasive observation of single cell behavior over time is an absolute requirement.

This seminar will address how single cell technologies, including advances in time-lapse imaging and molecular profiling, can be applied in hematology to study normal and leukemic HSPC fate. Further it will provide an interactive forum to discuss how both approaches (destructive and non-destructive) can be combined in a meaningful way to the study of normal and malignant hematopoiesis.

Disclosure: No conflict of interest disclosed.


Allogene Stammzelltransplantation

V317 - Allogeneic stem cell transplantation - current concepts and future directions

Hemmati P.1, Arnold R.1

1Charité - Universitätsmedizin Berlin, Medizinische Klinik m.S. Hämatologie, Onkologie und Tumorimmunologie, Berlin, Germany

Within the last two decades allogeneic hematopoietic stem cell transplantation (alloHSCT) has evolved into an important treatment option for a broad spectrum of malignant and non-malignant diseases of the hematopoietic system. As compared to conventional therapeutic approaches, e.g. chemotherapy, alloHSCT offers the highest potential for achieving remission and, ultimately, cure from the underlying disease. This strongly underscores the importance of alloreactivity in mediating the graft-versus-tumor effect as a prerequisite for long-term disease control. With 15.000 allogeneic transplants performed annually in Europe the use of alloHSCT has grown exponentially with respect to indications. Furthermore, the development of reduced-intensity/toxicity conditioning regimens has allowed expanding alloHSCT to elderly and/or frail patients ineligible to standard conditioning. Likewise, advances in supportive care have led to a substantial reduction in short and long-term morbidity and mortality, one of the major obstacles for achieving successful treatment outcome. Finally, improvements in HLA-typing have allowed for better donor selection.

Despite tremendous progress has been made in recent years, many aspects of alloSCT remain to be settled. Burning questions include: how to choose the most appropriate donor and what is the role of alternate stem cell sources? How and when to modify conditioning and post-transplant immunosuppression to optimize the balance between disease control and transplant-related toxicity? What is the optimal tool for post-transplant monitoring of minimal-residual disease? In addition to specific conditions during the intermediate post-transplant interval, e.g. acute graft-versus-host disease or infections, a substantial proportion of patients survive long-term and, therefore, are prone to a unique set of complications and late effects. This, in turn, prompts the need to develop novel follow-up strategies. These and other issues will be interactively presented and discussed during the session.

Disclosure: No conflict of interest disclosed.



V319 - Molecular diagnostic in ALL: Standards and novel approaches

Baldus C.D.1

1Charité Campus Benjamin Franklin, Hämatologie/Onkologie/Tumorimmunologie, Berlin, Germany

Acute lymphoblastic leukemia (ALL) represents a clinical and molecular highly heterogeneous disorder. Thus, a comprehensive diagnostic work up is necessary to allow risk stratification and the detection of therapeutic targets. Whereas the compulsory diagnostic program comprises standard morphology, FACS, cytogenetics, and molecular genetics, novel insights are gained by genome wide studies.

The morphological assessment of lymphoblasts by microscopy is essential to confirm or rule out L3 morphology as hint for a Burkitt leukemia. The immunophenotypic determination of lineage commitment is critical for the correct diagnosis of ALL; phenotypes with therapeutic implications are T-cell, mature B-cell, B-cell precursor phenotypes. The chromosomal analysis is an integral component of the initial work-up and identifies aberrations in 60-80% of patients. In addition, highly specific and sensitive molecular techniques, such as RT-PCR, fluorescence in-situ hybridization, allow detection of fusion transcripts and chromosomal translocations with prognostic or therapeutic relevance. Importantly, t(9;22)/BCR-ABL directs the use of tyrosine kinase inhibitors (TKI). As treatment response is highly predictive of the risk of relapse, the identification of leukemia-specific markers (immunoglobulin, T-cell receptor genes, fusion transcripts) at diagnosis is necessary to enable monitoring of residual clonal ALL cells, e.g. minimal residual disease (MRD), during the course of treatment.

In addition to this required diagnostic work-up, insights form exploratory studies are emerging and will shape a novel work-up in the near future. Gene-expression profiling has shown to accurately identify major ALL subtypes, and allowed the recognition of a new subtype, the Philadelphia-like ALL. By genome wide sequencing studies this novel high-risk subtype was characterized by alterations that activate cytokine receptors or tyrosine kinases amenable to inhibition with TKI. Genomic profiling has also enabled the identification of clonal heterogeneity and genetic alterations enriched at relapse. Many of these findings are of clinical relevance, thus efforts are made to capture molecular alterations in novel diagnostic tests including NGS based gene panels.

A detailed molecular classification at diagnosis and during the course of ALL is highly relevant to guide therapeutic decisions for the individual patient and to allow the implementation of molecular directed therapy approaches.

Disclosure: No conflict of interest disclosed.


Melanom neue Substanzen und Strategien

V324 - Radiation therapy for melanoma: Adjuvant, palliative and abscopal effects

Rogers S.1, Datta N.1, Bodis S.1

1Kantonsspital Aarau, Radio-Onkologie-Zentrum KSA-KSB, Aarau, Switzerland

Radiation therapy (RT) has long played a role in the management of melanoma, however historically its efficacy has been questioned due to the perceived radioresistance of melanoma cells in vitro. Apart from proton therapy for choroidal melanoma, RT has limited accepted indications as a definitive treatment. Surgery remains the first line therapy of choice for operable patients and adjuvant RT can be considered for high-risk primary tumours. Prospective randomised data are awaited following a Phase II TROG study reporting high local control rates (93%) following postoperative nodal irradiation in selected patients with node-positive melanoma. High response rates of melanoma (e.g. mucosal or inoperable) to primary radiotherapy have also been reported. Local radiation can achieve tumour-antigen release and clinical results suggest biological interaction with immunomodulating agents (a checkpoint inhibitor and anti-CTLA4 antibody), corroborated by preclinical data. Much interest also lies in enhancing melanoma radiation response rates through combination with clinical hyperthermia, which can achieve radiosensitisation, at least in part, by immune modulation. Novel developments in radiotherapy include the rise in radiosurgical treatment (SRS), involving high dose, small volume irradiation. SRS is increasingly the palliation of choice for patients with two or more brain metastases, following resection of a single brain metastasis and for extra-cranial oligometastases. The results of a Phase III trial evaluating whole brain radiotherapy are awaited. The radiobiological consequences of SRS may be even more immunomodulatory, congruent with retrospective data that fraction sizes greater than 4 Gray are more effective and SRS alone has been suggested to induce an abscopal effect. To date there are limited data reporting SRS with concomitant immunomodulators and the optimal combinations remain to be defined. Serious toxicities have been reported following administration of B-RAF inhibitors with and subsequent to both SRS and fractionated RT. Combination strategies are being actively researched as these should increase the effectiveness of radiation in the management of patients with melanoma by widening the therapeutic window between normal tissue toxicity and melanoma cell kill, and by potentially inducing an out-of-field abscopal effect.

Disclosure: No conflict of interest disclosed.


Mammakarzinom Standards in der adjuvanten Behandlung

V326 - Hereditary breast cancer - diagnosis, prevention, treatment

Morlot S.1, Schlegelberger B.1

1Hannover Medical School, Institute of Human Genetics, Hannover, Germany

Approximately 5% of all breast cancers and more than 10% of all ovarian cancers occur as a result of a hereditary predisposition. Possible indications of a hereditary cause can be found in the family and patient's own medical history (frequent breast, ovarian cancer, pancreatic and/or prostate cancer in the family, young age, triple-negative breast cancer, bilateral breast cancer, male breast cancer, serous ovarian cancer). In about 20-25% of these “high-risk” families, a mutation can be found in one of the known tumor predisposition genes (BRCA1, BRCA2, CHEK2 and other DNA repair genes). It is of the utmost importance to identify these “high-risk” families in order to offer 1) intensified screening measures, e.g. breast screening including breast MRI or early detection of pancreatic cancer, 2) prophylactic surgery, particularly prophylactic salpingo-oophorectomy and 3) predictive testing of healthy family members. The knowledge of a BRCA1 or BRCA2 mutation has implications for treatment: due to the significantly increased risk of secondary breast cancer (ipsilateral or contralateral) after the first diagnosis of breast cancer, therapeutic (and prophylactic contralateral) mastectomy versus breast-conserving therapy and radiotherapy have to be discussed. Chemotherapy may be adapted because of the better response to platinum derivatives and the promising new PARP inhibitors. Fast-track genetic diagnostics can be carried out within a few days to aid decision-making.

Disclosure: No conflict of interest disclosed.

Wissenschaftliches Symposium


V331 - Advances in the field of pancreatic cancer: What is the role of whole genome sequencing?

Wicki A.1

1Universitätsspital Basel, Onkologie, Basel, Switzerland

Next generation sequencing (NGS) is supposed to change the face of cancer care. With the help of a single sequencer, 8 complete cancer genomes can now be analysed in a single day. Thus, from a technical point of view, there are no obstacles preventing patients and physicians to look at individual cancer genomes in depth. However, the new challenge consists in interpreting NGS data and turning them into something that benefits cancer patients.

With regard to pancreatic cancer, whole genome sequencing plays a double role: on one hand, whole genome sequencing can identify mutations or copy number variations of druggable oncogenes that occur in a significant proportion of pancreatic cancers, but at a low individual patient prevalence. In addition, NGS has already identified new driver genes of pancreatic carcinogenesis, and those may become actionable in the future. On the other hand, whole genome sequencing can help to assess the occurrence and structure of neo-antigens in pancreatic cancer. Together with new approaches of modulating the immune environment of a tumor, this may help to develop tailored immunotherapies for this disease.

Disclosure: No conflict of interest disclosed.


Urothelkarzinom - Management 2015

V334 - Tumors of the upper urinary tract and urethra - multidisciplinary management

Gakis G.1

1Klinik für Urologie, Tübingen, Germany

Introduction: The treatment of rare urogenital tumors represents a major clinical challenge. Due to their low incidence, most tumors of the upper urinary tract and urethra are diagnosed in locally advanced stages. Moreover, by contrast to upper tract urothelial carcinoma (UTUC), urethral cancers can arise from different histological entities which inherits therapeutic implications.

Material and methods: Based on current data from European and North American cancer registries, improvements in the understading of the biology of tumors of the upper urinary tract and urethra have been recently made. In addition, the European Association of Urology has published this year the first update of the 2013 Guidelines on Primary Urethral Carcinoma.

Results: Radical nephroureterectomy (RNU) represents the mainstay of treatment for UTUC. Yet, solitary, low-grade and low-stage upper tract tumors can be treated safely with a kidney-sparing approach, especially when located in the ureter. Some retrospective studies suggest that neoadjuvant cisplatin-based, multi-agent chemotherapy can be offered to patients with locally advanced UTUC to reduce the risk of micrometastatic disease prior to RNU. In localized urethral carcinoma, a urethral-sparing approach should be considered, if negative surgical margins can be achieved intraoperatively. In locally advanced urethral cancer, cisplatin-based multiagent chemotherapy (or a radiosensitizing chemotherapy with concurrent radiotherapy for patients with squamous cell carcinoma) may exert a beneficial impact on survival and even enable genital preservation. In case of urethral recurrence, recent data suggest that salvage surgery is associated with an improved overall survival.

Conclusions: Low-stage and low-grade tumors of the upper urinar tract and urethra can be safely treated with organ-sparing surgery, whereas a multidisciplinary approach should be especially considered for patients with locally advanced tumors.

Disclosure: No conflict of interest disclosed.



V339 - “Cancer-related fatigue: What is to do?”

Fischer I.1,2

1Institut für Tumor-Fatigue-Forschung, Emskirchen, Germany, 2Deutsche Fatigue Gesellschaft, Köln, Germany

Cancer-related fatigue (CrF) is defined as a persistent, unusual , subjective sensation of physical, emotional and cognitive tiredness which occurs in the context of cancer or its treatment. Compared with the tiredness felt by a healthy individual, CrF is perceived as being of greater magnitude, disproportionate to recent activity, and it is not completely relieved by rest, leaving the patient with an overwhelming and distressing sense of exhaustion. Prevalence estimates of CrF range from 20% to 99%, depending on the sample, the time and the method of assessment. The CrF symptoms can be temporary, but can also persist up to 15 years after successful treatment completion. Having a profound effect on the patient´s life, CrF is much more than just being tired: Patients are often unable to engage in their usual activities of daily living (up to the inability to perform one's job and earn a living), social relationships are lost and quality of life is impaired. Thus, CrF clearly poses a problem not only for the patients themselves, but also for the persons around them.

Although evidence-based therapies for CrF are available and though the National Comprehensive Cancer Network (NCCN, 2015) recommends that CrF should be “recognized, evaluated, monitored, documented and treated promptly for all age groups, at all stages of disease, prior to, during, and following treatment”, CrF is not sufficiently perceived by physicians: Individuals are often reluctant to report fatigue, and cancer-care providers frequently do not screen for it because they are uncertain about how to treat it. However, patients are suffering, and since fatigue is related not only to cancer, but also to other diseases and symptoms (e.g. underlying comorbidities, emotional distress, anemia, sleep disturbance, pain, or adverse events of medication ), diagnosis and differential diagnosis are a significant precondition for an adequate causal and/ or symptomatic treatment of CrF.

This further education provides an overview about adequate screening methods and diagnostic procedures (e.g. anamnesis, suitable self report questionnaires) as well as about symptomatic treatment options supported by evidence from systematic reviews, meta-analyses and randomized controlled clinical trials (the latter with CrF as a primary endpoint), such as physical training, patient education, cognitive behavior therapy, energy conservation, phytotherapy, (psycho-)stimulants or corticosteroids.

Disclosure: No conflict of interest disclosed.

Freier Vortrag

Stammzellen I

V341 - Effective mobilization of hematopoietic stem cells with a small molecule α9β1/α4β1 integrin antagonist

Grassinger J.1, Cao B.2,3, Zhang Z.2, Klatt S.1, Williams B.2,3, Mueller G.1, Hart C.1, Schelker R.1, Herr W.1, Nilsson S.K.2,3

1University Hospital Regensburg, Internal Medicine III, Regensburg, Germany, 2Commonwealth Scientific and Industrial Research Organization, Manufacturing Flagship, Melbourne, Australia, 3Monash University, Australian Regenerative Medicine Institute, Melbourne, Australia

Introduction: Mobilization of sufficient numbers of hematopoietic stem cells (HSC) is a prerequisite for a successful engraftment in peripheral blood stem cell transplantation (PBSCT). To date, granulocyte-colony stimulating factor (G-CSF) is the most effective mobilization agent used clinically. However, G-CSF has substantial nonspecific effects on peripheral blood (PB) and bone marrow (BM) cells and, according to our data, mainly expands hematopoietic stem and progenitor cells (HSPC) within the central BM region. We recently showed that biologically superior HSC are located at the endosteal BM region and therefore propose that mobilization of these HSC will result in superior engraftment than G-CSF mobilized HSC.

Methods: A small molecule α9β1/α4β1 integrin antagonist (N-(Benzene-sulfonyl)-L-prolyl-L-O-(1-Pyrrolidinylcarbonyl)-tyrosine; BOP) and the fluorescent analogue (R-BC154) was synthesized and shown to disrupt the binding of HSC to extracellular ligands within the endosteal niche. In vitro and in vivo analysis was performed to investigate the binding capacity of BOP and HSC mobilization efficiency.

Results: Using R-BC154, we show that this class of antagonists preferentially bind mouse and human HSC via intrinsically activated α9β1/α4β1 integrins within the endosteal niche. BOP rapidly mobilizes HSC with long-term multi-lineage engraftment potential in mice. Additive augmentation of the engraftment of PB-HSC was observed when BOP was co-administered with a SDF-1 antagonist (Plerixafor). Impressively, this combination effectively outcompeted PB-HSC mobilized with 4 days of G-CSF treatment. The enhanced mobilization observed with the small molecule combination was recapitulated in humanized NSG mice, where a significant increase in PB CD34+ HSPC was observed after treatment with BOP and Plerixafor.

Conclusions: Dual α9β1/α4β1 integrin inhibitors effectively mobilize murine and human HSC in mice. The combination of BOP and Plerixafor might allow efficient single dose mobilization strategies with reduced side effects as compared to multiple G-CSF injections. Moreover, mobilization of aberrant hematopoietic cells in acute and chronic myeloid leukemia using BOP might enhance treatment efficiency and is currently under investigation.

Disclosure: No conflict of interest disclosed.

V342 - The role of the transcription factor trps1 an its relation to evi1 in zebrafish hematopoietic development

Alghisi E.1, Konantz M.1, Lengerke C.1,2

1University Hospital Basel, Department of Biomedicine, Basel, Switzerland, 2University Hospital Basel, Clinic for Hematology, Basel, Switzerland

Introduction: The EVI1 (ecotropic viral integration site 1) gene is a zinc finger transcriptional regulator that is expressed in hematopoietic stem cells (HSC) and our lab has shown that it regulates their emergence during development via up-regulation of the NOTCH pathway in the aorta-gonado-mesonephros (AGM) region However, the precise molecular mechanism of EVI1-mediated NOTCH-induction is unclear (Konantz et al., unpublished). Here we study the trichorhinophalangeal syndrome 1 (trps1) gene, a GATA family transcriptional factor known for its role in skeletal development, with respect to its role in evi1-mediated hematopoiesis. TRPS1 has been reported to induce epithelial-to-mesenchymal transition and angiogenesis in breast carcinoma (BC) cells through activation of its direct target gene VEGFA. Previous data from our lab report that VEGF overexpression can rescue HSC defects in evi1-morphants.

Methods: Genetic modifications were obtained by injection of inhibiting morpholino oligonucleotides or mRNA into the ZF zygote or via heat-shock inducible transgenic lines. Hematopoiesis was analysed by in situ hybridization for trps1, runx1, cmyb, rag1, notch1b and notch3 in the AGM, the caudal hematopoietic tissue (CHT) and the thymus.

Results: Zebrafish embryos show trps1 expression in the brain, the jaw mesenchyme and potentially in the AGM, and later in development also in the kidney marrow. Knockdown of trps1 reduced the formation of cmyb/runx1-double positive HSC in the AGM region as well as the formation of rag1 positive lymphoid cells in the thymus, indicating a functional role in definitive hematopoiesis. Importantly, trps1 morphants showed down-regulation of NOTCH pathway genes in the dorsal aorta and AGM regions suggesting an upstream role of trps1 in the vegf-notch axis in developmental hematopoiesis. Co-injection of evi1 mRNA was not able to rescue runx1 expression and morphants showed a normal evi1 expression compared to controls, indicating that indeed trps1 may act as an evi1 downstream target. Rescue experiments of evi1 morphants with trps1 mRNA further exploring this hypothesis are underway.

Conclusion: Taken together, our data indicate an unsuspected role of trps1 in hematopoiesis and suggest this transcription factor as a possible downstream target of evi1 during blood stem cell development. We are currently exploring trps1 as a direct target of evi1.

Disclosure: No conflict of interest disclosed.

V343 - Wnt5a expressed by the niche maintains migratory properties of hematopoietic stem cells through the planar cell polarity pathway

Schreck C.1, Istvanffy R.1, Ziegenhain C.2, Gärtner F.3, Grziwok S.1, Pagel C.1, Henkel L.4, Schiemann M.4, Götze K.1, Massberg S.3, Peschel C.1, Enard W.2, Oostendorp R.A.J.1

1III. Medizinische Klinik, Klinikum rechts der Isar, Technische Universität München, München, Germany, 2Department of Biology II, Ludwig-Maximilians-Universität, Anthropology and Human Genetics, München, Germany, 3Ludwig-Maximilians-Universität, I. Medizinische Klinik, München, Germany, 4Technische Universität München, Institute of Medical Microbiology, Immunology, and Hygiene, München, Germany

We previously identified Wnt5a as a secreted factor that maintains HSCs in vitro (Buckley, et al., Exp Hematol. 2011) and as a factor responsible for decline of HSC function during aging (Florian et al., Nature. 2013). We here studied in detail how Wnt5a-deficiency affects the niche and its ability to maintain long-term repopulating HSC. The BM niche in Wnt5a-deficient mice show increased number of CD31+ endothelial cells. The number of mature and immature mesenchymal cells was not different, but multipotent stromal cells (MSCs) showed increased proliferation and calcification upon differentiation. Chimeric transplantation experiments showed that Wnt5a± HSCs engraft similarly to wild-type Wnt5a+/+ (WT) HSCs up to tertiary transplantations. To assess whether niche Wnt5a affects engraftment, and/or self-renewal capacity of HSCs, wild-type (WT) HSCs were transplanted in Wnt5a-deficient recipients. We found that in primary recipients, engraftment was similar to that in WT recipients. Unexpectedly, however, WT donor LSK cells from the Wnt5a-deficient recipients completely failed to engraft in secondary recipients. Although the phenotype of the WT donor cells engrafted in either WT or Wnt5a± environments was similar, the sorted LSK cells from Wnt5a± recipients showed defective, apolar distribution of Cdc42 and F-actin, suggesting possible defects in migratory properties. Indeed, RNAseq of sorted donor LSK cells from Wnt5a± recipients showed highly divergent expression of molecules involved in heterotrimeric G-protein assembly and small GTPase-mediated planar cell polarity (PCP) signal transduction. Further experiments showed that WT donor LSK cells from Wnt5a± recipients showed defective lamellipodia formation, and only poorly homed to the bone marrow of recipient WT mice. In addition, mature T and B lymphocytes failed to migrate towards CXCL12.

In conclusion, our experiments show that a Wnt5a-deficient niche allows for initial engraftment, but fails to maintain the ability of engrafted HSCs to properly home and engraft secondary recipients, due to defective PCP-regulated migratory properties of HSCs and mature lymphoid cells.

Disclosure: No conflict of interest disclosed.

V344 - Allogeneic T cells disrupt medullary thymic epithelial cell formation and indirectly lead to chronic graft-vs-host disease

Müller A.M.1,2, Florek M.2, Min D.3, Burnett C.2, Weinberg K.3, Shizuru J.A.2

1UniSpital Zürich, Hämatologie, Zürich, Switzerland, 2Stanford University, BMT Medicine, Stanford, United States, 3Stanford University, Stanford, United States

Graft-vs-host disease (GVHD) remains a major complication of allogeneic hematopoietic cell transplantation (HCT). Acute (a) GVHD results from direct donor T cell (TC) damage of organs. In contrast, the biology of chronic (c) GVHD with its various autoimmune-like manifestations remains poorly understood.

We studied the pathophysiology of cGVHD in an MHC-matched mouse model (C57BL/6 à BALB.B) using lethal irradiation and HCT of pure hematopoietic stem cells (HSC; cKIT+Thy1.1loSca1+Lin-) or HSC + TC. Recipients of pure HSC remained healthy, whereas mice given HSC+TC developed aGVHD (diarrhea, weight loss) with a mortality of ~30%. Survivors stabilized around d+45, but developed clinical chronic GVHD after 6-12m with sclerodermatous skin excoriations, cataracts, and liver fibrosis/cirrhosis.

Further, thymuses were a major target of aGVHD resulting in severe hypocellularity and disrupted organ architecture. During the months of lymphoid reconstitution thymuses of HSC+TC-recipients showed markedly lower expression of cytokeratin 5 (CK5) than HSC recipients. CK5 marks medullary thymic epithelial cells (mTEC) that provide a specialized microenvironment for survival, proliferation, and differentiation of immature TC. Expression of AIRE, a transcription factor in the thymic medulla that controls negative selection during TC maturation was low in all transplanted groups. During aGVHD only TH1-donor TC, but not CD4+IL17+ (TH17)-cells were detectable. Starting at 2m TH17 cells emerged, first in intestines then liver and skin (all typical GVHD target organs) increasing until 6-12 months post-HCT. Of note, TH17 cells originated from donor-HSC, not adaptively transferred mature TC. Even recipients of pure HSC showed increasing proportions of TH17 cells over time, and could manifest discrete signs of cGVHD.

From our model we hypothesize that the thymus is damaged by transplant conditioning and alloreactive donor T cells. Disruption of medullary thymic epithelial cells (mTEC) may lead to impaired restoration of the epithelial network. As a consequence, negative selection of nascent HSC-derived donor T cells is compromised and emergence of IL-17 secreting donor T cells in target organs of chronic GVHD could be due to misguided T-cell maturation in a disrupted thymic microenvironment. Subclinical GVH-reactions within the thymus could thereby contribute to chronic GVHD in an autoreactive fashion.

Disclosure: No conflict of interest disclosed.

V345 - Specific transcripts of DNMT3A modulate differentiation of hematopoietic progenitor cells

Božić T.1, Frobel J.1, Raić A.1, Goecke T.2, Jost E.3, Wagner W.1

1Helmholtz-Institute for Biomedical Engineering, Stem Cell Biology and Cellular Engineering, Aachen, Germany, 2RWTH Aachen University Hospital, Department of Obstetrics and Gynecology, Aachen, Germany, 3RWTH Aachen University Hospital, Clinic for Oncology, Hematology, and Stem Cell Transplantation, Aachen, Germany

Introduction: DNA-methyltransferase 3A (DNMT3A) is a de novo DNA-methyltransferase that is alternatively spliced in a tissue- and disease-specific manner, but the functional relevance of these transcripts is hardly known. DNMT3A is frequently mutated in patients with acute myeloid leukemia (AML) and we have recently demonstrated that mutations in DNMT3A can be mimicked by aberrant hypermethylation within the DNMT3A sequence: about 40% of AML patients carry this “epimutation” that is associated with shorter overall survival. Notably, DNMT3A mutations as well as its “epimutations” seem to have impact on the expression of different DNMT3A transcripts. Therefore, we aimed for a better understanding of the functional role of individual DNMT3A splice variants in hematopoiesis.

Methods: Specific DNMT3A transcripts (transcripts 1+3, transcript 2 or transcript 4) were knocked down by lentiviral expression of individual short-hairpin RNAs (shRNAs) in CD34+ progenitor cells isolated from human cord blood. Subsequently, we evaluated the impact on colony formation potential (CFU assay), proliferation (CFSE assay), and the immunophenotype (CD34+ and CD133+). DNA methylation profiles of individual knockdowns were generated with the Infinium HumanMethylation450 BeadChip platform and are under current analysis.

Results: Knockdown efficiency of DNMT3A transcripts 1+3, transcript 2, and transcript 4 was validated by qRT-PCR (30%, 55% and 70% of basal expression, respectively). Downregulation of either transcript 2 or 4 reduced the proliferation rate significantly (N = 3, p < 0.05). Notably, cells maintained a more primitive phenotype expressing CD34+ for more cell divisions upon knockdown of transcript 2 (N = 3; p < 0.05), whereas CD133+ expression continuously declined in all knockdowns and the control. Downregulation of transcript 4 had significant effect on the CFU potential, leading to a bias for erythroid colonies, together with an increase in the total number of colonies.

Conclusions: Our results indicate that individual DNMT3A transcripts have unique regulatory functions during the differentiation process of blood progenitors that might be relevant for development of AML.

Disclosure: Tanja Božić: No conflict of interest disclosed. Wolfgang Wagner: Employment or Leadership Position: Wolfgang Wagner is involved in the company Cygenia that provides service for analysis of the DNMT3A epimutation (www.cygenia.com).; Honoraria: RWTH Aachen has applied for a patent for the DNMT3A epimutation.

V346 - Signal transduction and functional effects of G protein-coupled receptors in hematopoietic stem- and progenitor cells

Manz P.1, Krauß U.1, Kanz L.1, Möhle R.1

1Med. Univ.-Klinik II, Tübingen, Germany

Introduction: G protein-coupled receptors (GPCR), e.g. the chemokine receptor CXCR4, the leukotriene receptor CysLT1, and the sphingolipid receptor S1P1, represent important regulators of the “stem cell niche,” which supports maintenance and survival of hematopoietic stem and progenitor cells (HPC) in the bone marrow microenvironment. However, the differential effects of these GPCR and their individual roles are poorly understood.

Methods: We analyzed signal transduction pathways of CXCR4, CysLT1, and S1P1 in CD34+ HPC and cell lines (THP-1), particularly calcium signaling by flow cytometry and MAP-kinase/Erk phosphorylation by western blot and flow cytometry. Adhesion of HPC to stromal cells and fibronectin was analayzed in adhesion assays. Proliferation of HPC with and without coculture with stromal cells was also assessed.

Results: We found that after stimulation with their respective ligands, CysLT1 induced the strongest response in calcium signaling, while CXCR4 and particulary S1P1 only moderately mediated intracellular calcium release in HPC. Using a quantitative flow cytometric assay, strong MAPK phosphorylation was mediated by CXCR4, while a modest increase was induced by CysLT1, and only a weak response observed after stimulation of S1P1. Stimulation of both CXCR4 and CysLT1 synergistically increased MAPK phosphorylation, which was not further augmented by activation of S1P1. However, only combined inhibition of all three GPCR resulted in reduced proliferation of HPC in vitro. Interestingly, a converse effect on HPC adhesion was induced by S1P1, which inhibited adhesion of CD34+ HPC to fibronectin, compared to CXCR4 and CysLT1, which both mediated an increased HPC attachment.

Conclusions: We conclude that the G protein-coupled receptors CXCR4, CysLT1, and S1P1 induce differential signaling and functional effects in HPC. While CXCR4 elicited the most prominent effect on calcium signaling, MAPK phosphorylation and signaling related to cell proliferation was increased particularly by CXCR4. Our results suggest that CXCR4 and CysLT1 support lodging and proliferation of HPC in the hematopoietic microenvironment, while only weak effects of S1P1 on calcium and MAPK signaling and even reduced adhesion of HPC fits to the recently recognized opposite role of S1P1 supporting HPC egress and mobilization. Thus, different GPCR play diverse roles in the dynamic regulation of the HPC niche.

Disclosure: No conflict of interest disclosed.

Freier Vortrag

Nicht maligne Hämatologie

V350 - Data from German centers in the global PNH Patient Registry- analysis of the translation of the DGHO-guidelines for the treatment of Paroxysmal Nocturnal Hemoglobinuria (PNH) in real life therapy

Höchsmann B.1, Leichtle R.1, Röth A.2, Panse J.3, Haferlach T.4, Borchmann P.5, Aulitzky W.E.6, Dengler J.7, Port M.8, Platzbecker U.9, Klausmann M.10, Steinmetz T.11, Becker M.12, Schmidt B.13, Schrezenmeier H.1

1Institut für klinische Transfusionsmedizin und Immunogenetik Ulm, DRK Blutspendedienst Baden-Württemberg-Hessen und Universität Ulm, Ulm, Germany, 2Universitätklinik Essen, Abteilung Hämatologie, Essen, Germany, 3Universitätsklinikum Aachen, Klinik für Onkologie, Hämatologie und Stammzelltransplantation (Medizinische Klinik IV), Aachen, Germany, 4MLL Münchner Leukämielabor GmbH, München, Germany, 5Universitätsklinikum Köln, Klinik I für Innere Medizin, Köln, Germany, 6Robert Bosch Krankenhaus Stuttgart, Hämatologie und Onkologie, Stuttgart, Germany, 7Medizinische Universitätsklinik Heidelberg, Abteilung Innere Medizin V, Heidelberg, Germany, 8Medizinische Hochschule, Abteilung Hämatologie und Onkologie, Hannover, Germany, 9Universitätsklinik Dresden, Abteilung Hämatologie und Onkologie, Dresden, Germany, 10Gemeinschaftspraxis Drs Klausmann und Dr Welslau, Aschaffenburg, Germany, 11Gemeinschaftspraxis Hämatologie und Onkologie, Köln, Germany, 12Onkologische Praxis Minden/Porta, Minden, Germany, 13Onkologische Praxis Pasing, München-Pasing, Germany

Introduction: The International PNH Patient Registry was started to enhance the understanding of this orphan disease and includes patients (pts) independent of disease severity or treatment. Complement inhibition with eculizumab is capable to improve the clinical symptoms and overall survival of PNH pts. An expert panel of the DGHO defined guidelines for diagnosis and treatment of PNH (https://www.dgho-onkopedia.de). According to recent data confirming a strong correlation between clinical symptoms and prognosis eculizumab is recommended for symptomatic pts with hemolytic PNH.

Methods: To study the adherence to the DGHO guidelines we analysed the rate of symptomatic PNH pts in the PNH Patient Registry which collects pseudonymized patient data after obtaining an informed consent. Data were collected at baseline and the last follow up (FU) in all pts and in the subgroups of pts ever or never treated with eculizumab. Furthermore, the worsening of the disease defined as the new occurrence of symptoms from baseline to the last FU was investigated. Baseline is defined as the start date of eculizumab for treated pts or registry enrollment date for never treated pts. Last FU is defined by the first reported date of death, BMT, withdrawal from the Registry, or discontinuation of eculizumab (in treated pts). In pts not meeting any of these criteria, last FU is the date of last contact from the Registry.

Results: At enrollment the mean age was 45.1 ± 18.42 yrs, mean GPI-deficient granulocyte population was 52.8 ± 34.32, mean LDH was 3.9 ± 3.55 xULN, The mean follow up duration was 2.2 ± 2.2 yrs in the complete pts group, 2.5 ± 2.3 yrs in the pts ever treated with eculizumab and 1.9 ± 2.1 yrs in the pts never treated with eculizumab. The table below shows the data of the PNH related symptoms according to the treatment with complement inhibition.


Conclusion: A substantial subgroup is not treated with eculizumab despite symptoms. Status deterioration, including new thromboembolic events, was more pronounced in the subgroup never treated with eculizumab. We will present further analyses of the German PNH Patient Registry regarding combination of clinical signs and therapeutic decisions to enable an further improvement of the treatment strategy in symptomatic PNH.

Disclosure: Britta Höchsmann: Advisory Role: ja; Financing of Scientific Research: ja; Expert Testimony: ja Hubert Schrezenmeier: Advisory Role: ja; Financing of Scientific Research: ja; Expert Testimony: ja.

V351 - Red cell calcium-channelopathies - novel approaches and mechanism

Kaestner L.1, Wang J.1, Hertz L.1, Ruppenthal S.2, Lipp P.2, Birnbaumer L.3, Freichel M.4

1Research Centre for Molecular Imaging and Screening, Homburg/Saar, Germany, 2Institute for Molecular Cell Biology, Homburg/Saar, Germany, 3National Institute of Environmental Health Sciences, Research Triangle Park, United States, 4Institute for Pharmacology, Heidelberg, Germany

The molecular origin of hereditary anaemia is sometimes known but often obscure. However, in both situations, the molecular mechanism leading to the symptoms of the disease remain in many cases elusive.

We present single-cell based methods that allow the investigation of functional properties of red blood cells (RBCs) upon exposure to pharmacological and physical stimulation. We investigated in particular the molecular basis of calcium entry by video imaging of intracellular calcium combining pharmacological inhibitors and transgenic animal models (mice).

Lysophosphatidic acid (LPA) induces a concentration dependent calcium influx in RBCs. We found that this influx happens via a bifurcated signal pathway. The signal transduction starts with the common LPA-receptor agonist binding and the consecutive activation of the Gi-Protein, which triggers the two branches: (i) activation of MEK/MAPK and PI3 kinase followed by an ω-Agatoxin TK sensitive calcium-entry and (ii) activation of the non selective cation channel TRPC6. In detail the LPA induced TRPC6 cascade is: LPA -> Gi -> PLCε -> PIP2 -> DAG -> PKCα -> TRPC6. However, it was unclear, how PKC phosphorylation can activate TRPC6. We could generate evidence that the phosphorylation of TRPC6 by PKCα initially leads to an inactivation of the channel. This inactivation enables TRPC6 channels to form an assembly platform with FK binding protein and calcineurin. The dephosphorylation of TRPC6 by calcineurin then activates TRPC6 and such enables an influx of calcium into the RBC. Furthermore we could show that the function of this pathway is increased in RBCs of sickle cell disease patients through a higher abundance of LPA-receptor 4 in sickle cells.

The LPA signaling pathway and the resulting activation of channels place them into the focus as pharmaceutical targets for the treatment of severe symptoms of anaemia, such as the vaso-occlusive crisis in sickle cell disease. This holds particularly true, as calcium has been identified as a trigger for RBCs self aggregation and for their adhesion to endothelial cells.

Disclosure: No conflict of interest disclosed.

V352 - Artesunate therapy for vaso-occlusive crisis with multi-organ failure due to severe plasmodium falciparum infection in a patient with HbSC trait

Trummer A.1, Thol F.1, Ganser A.1

1Hannover Medical School, Hematology, Hemostasis, Oncology & Stem Cell Transplantation, Hannover, Germany

Introduction: Hemoglobin sickle cell disease (HbSC) is the second most frequent hemoglobinopathy after homozygous sickle cell anemia (SCA). Its clinical course often differs from SCA as it is mostly complicated by thrombotic complications due to hyperviscosity rather than hemolytic-related vasculopathy. Like SCA, HbSC has been associated with a significant risk reduction of malaria. Nevertheless, malaria chemoprophylaxis is often recommended as malaria can be a significant cause of morbidity and mortality in sickle cell patients.

Case: We report the case of 45 year old west-african man with HbSC trait who had only been transfused with RBC once before. He had initially reported to our outpatient clinic due to ongoing bone pain for about 6 months. However, his diagnostic work-up showed only mild hemolysis and no signs of osteonecrosis. Hemoglobin levels were around 14 g/dl with a reticulocytosis of 170/nl, and hemoglobin electrophoresis revealed 51% HbS, 46% HbC and 3% HbA.

As the patient refused phlebotomy, we initiated a therapy with hydroxyurea (500mg/day) and analgetics, which led to a significant clinical improvement. However, the patient left Germany for a 3 month stay in Togo, taking chloroquine for malaria prophylaxis. Again, he developed severe bone pain during the last 3 weeks of his stay and, finally, he reported to our department having fever with chills. Ebola infection was ruled out, but the patient worsened dramatically and rapidly developed multi-organ failure with massive haemolysis, liver cell necrosis, acute kidney failure requiring haemodialysis, paralytic ileus, osteonecrosis and acute myocardial infarction. A thick blood smear revealed infection with Plasmodium falciparum (about 2‰) and a therapy with atovaquone/proguanil as well as RBC exchange transfusions was initiated. However, as the blood smear remained positive after 48 hours, therapy was switched to artesunate/doxycycline and the blood smear was found negative after another 48 hours. The patient slowly recovered and, more than 6 months later, almost all organ functions returned to normal.

Conclusions: Despite a significant risk reduction in HbSC, malaria infection can cause life-threatening hemolysis and vaso-occlusive disease and clinicians need to be aware that even chemoprophylaxis does not ensure sufficient protection. Artesunate/doxycycline appears to be a rapid and efficient therapy for patients irresponsive to other malaria treatments.

Disclosure: No conflict of interest disclosed.

V353 - Increasing number of patients with hemoglobinopathies in the Ruhr metropolitan area

Distelmaier L.1, Dickerhoff R.2, Dührsen U.1

1University Hospital Essen, Department of Hematology, Essen, Germany, 2University Hospital Duesseldorf, Department of Pediatric Oncology, Hematology and Clinical Immunology, Duesseldorf, Germany

Introduction: Due to migration hemoglobinopathies are becoming more frequent in Germany. Especially in industrial areas the number of patients with thalassemia syndromes and sickle cell disorders is increasing while there is a lack of experienced physicians. This leads to incorrect diagnosis and treatment.

Methods: We summarize the numbers of patients we have seen in our hemoglobinopathy clinic at the university hospital in Essen since 2011 and evaluate the patients´ histories and former treatments. The term “patients” is used for all individuals referred to us, both asymptomatic heterozygous and patients with clinical disease.

Results: There has been a significant increase of patients with hemoglobin disorders from 2011 to 2015. Starting with 5 patients in the beginning of 2011, the clinic increased to 19 patients by the end of 2012, 41 in 2013, 71 in 2014 and by April 2015 it reached a total number of 95 patients. 52 of the patients were gene carriers of either HbS, thalassemia-syndromes or hemoglobins such as HbE, HbC and others, while 42 patients had a clinical significant hemoglobinopathy that needed treatment. Within the group of significant hemoglobinopathies, sickle cell disease was the most frequent disorder with 25 patients. Furthermore we found that 45 of the 95 patients had received incorrect treatments before they came to our clinic. 15 beta-thalassemia gene carriers where erroneously diagnosed as iron deficiency and received multiple treatments with iron. 6 HbS gene carriers received treatments meant for homozygous sickle cell disease. At least 10 patients with sickle cell disease had complications related to incorrect treatments. 7 patients with beta-thalassemia major or intermedia suffered from ineffective erythropoiesis or iron overload due to an insufficient transfusion program or iron chelation. The other 7 patients were a heterogeneous group that either came with a wrong diagnosis or had received various not indicated treatments.

Conclusions: Since 2011 the number of patients with hemoglobin disorders and hemoglobinopathy gene carriers that presented in our clinic has constantly increased. In Germany few physicians are experienced in recognizing and treating adults with hemoglobin disorders. For this reason many grown-up patients are still treated in pediatric clinics or are lost altogether. There is a strong need for more awareness and further education of physicians to avoid misdiagnoses and treatment-related complications.

Disclosure: No conflict of interest disclosed.

V354 - Claudia Pechstein's doping ban - reanalysis of the blood counts which led to the ban

Gassmann W.1

1St. Marienkrankenhaus, Siegen, Germany

Claudia Pechstein has been found guilty of blood doping on the basis of increased reticulocyte counts at World Championships in Hamar in 2009 and suspicious counts in the years before.

Blood counts from 2000 to 2009: This analysis comprises all 95 blood counts of Claudia Pechstein taken by anti-doping officials from 2000 to 2009. Repeatedly high reticulocyte counts may be the result of repeated use erythropoiesis stimulating agents or the result of shortened erythrocytic survival time.

Aside from high retics, high MCHC values were observed in Mrs. Pechstein at many occasions suggesting an erythrocytic abnormality. Mean value was 35.7 g/dl (normal range up to 36) with values above 37 g/dl at eleven occasions.

In case of blood doping, one would expect high haemoglobin levels at top events. However, mean haemoglobin levels and retics were the same at championships (mean 14.6 g/dl, 2.1%) compared to world cup competitions (14.5 g/dl, 1.9%) and compared to tests taken during training phases (14.6 g/dl, 2.0%).

The haemoglobin concentration should increase about 1 g/dl or more in case doping with the use of erythropoiesis stimulating agents was the reason for the elevated reticulocytes. This has never been observed.

The situation at Hamar in February 2009: Mrs. Pechstein had had five doping controls in January 2009. The first three controls were performed at the European Championship in Heerenveen. She won that championship with haematocrit levels of 39, 40 and 39% showing reticulocyte counts of 1.7%, 2.2% and 2.0%.

Further checks were done at a world cup race on January 30th and 31st with haematocrit levels as in Heerenveen and reticulocyte counts of 2.4% and 2.2%.

The first doping test at the Hamar championship was done on February 6th and showed a very high reticulocyte count of 3.5%. This count was confirmed at the following day immediately after a 3000m race. Haematocrit levels were determined as 41%, 39%, and 37%.

Another high reticulocyte at another championship in 2011: A high reticulocyte count of 3.8% and 3.1%, respectively, was observed at the world championship at Inzell in 2011. Doping controls including blood counts had been done at seven of the last ten days prior to that high reticulocyte count. No evidence of doping was found in the samples of blood and urine showing that high reticulocyte counts can occur in Mrs. Pechstein without doping.

The data available do not and did never support the suggestion of blood doping.

Disclosure: No conflict of interest disclosed.


Porphyrie / Stoffwechselkrankheit

V359 - Acute porphyrias - diagnosis and therapy

Petrides P.E.1, Bronisch O.1, Beykirch M.1

1Europäisches Porphyriezentrum, Hämatologisch onkologische Schwerpunktpraxis, München, Germany

Several errors of porphyrin metabolism can cause the acute porphyrias, which are characterized by a 50% reduction of the activity of particular enzymes of heme biosynthesis. Under normal conditions supply of porphyrins is sufficient in spite of this impairment. However, when higher amounts are required as for instance through the increased production of heme containing cytochrome P450 enzymes, the bottleneck of metabolism leads to a flood of porphyrin precursors (delta-aminolaevulinic acid (dALA) and porphobilinogen (PBG)) in the body. These precursors are the toxic molecules, which cause the acute attack and are therefore also of prime diagnostic relevance.

Cytochrome P450 enzymes are produced in the liver under the influence of certain drugs, so that drugs and also hormones are the most important (if, however, not the only ones) inducers of an acute attack.

The acute attack is often characterized by the symptom triad of abdominal pain, cerebral symptoms and a peripheral neuropathy. A red discoloration of the urine - in particular after extended storage - occurs in 30% of the patients and can lead to the correct diagnosis.

Severe attacks - often accompanied by a hyponatremia - can cause tetraplegias with the necessity of intubation and also lead to coma. When epileptic seizures occur, MRI analysis shows PRES (posterior reversible encelopathy syndrome), i.e. a vasogenic edema in the posterior cerebral hemispheric region.

When an acute porphyria is suspected the determination of PBG and dALA in a spontaneous urine sample is mandatory. When the values are elevated the presence of an acute porphyria is very likely. Further diagnosis is carried out by plasma fluorescence peak analysis, PBG-deaminase activity determination in red blood cells (in acute intermittent porphyrias) and fecal porphyrin analysis. Final proof is obtained through the identification of the disease causing mutation (genetic test).

Therapy of choice is the identification of toxic substances such as drugs and their immediate discontinuation, the intravenous application of high dose glucose and - more efficiently - the application of heme arginate which is available for intravenous infusion.

In this seminar, the relevant diagnostic and therapeutic aspects of the acute porphyrias will be illustrated using various examples from our cohort of nearly 50 patients with acute porphyrias which we have taken care of for the last 15 years.

Disclosure: No conflict of interest disclosed.



V360 - Standard treatment of gastric cancer - cases discussions

Lordick F.1

1Universitäres Krebszentrum Leipzig (UCCL), Leipzig, Germany

According to the International Agency for Research in Cancer (http://globocan.iarc.fr/Default.aspx), gastric cancer continues to be the fifth most common form of cancer from worldwide: 952,000 new patients are diagnosed with gastric cancer / year and 723,000 people die from gastric cancer. The Eurocare-5 study showed that the mean age-standardized 5-year relative survival for adult patients with gastric cancer diagnosed in 2000-2007 was only 25%, making it one of the most aggressive cancers in Europe (de Angelis, Lancet Oncol 2014).

In this Expert Seminar we will discuss different disease scenarios which are typical for multimodal and potentially curative but also for palliative treatment decisions. The five cases to be discussed are:

1. 44-year-old woman with localized signet ring cell cancer of the stomach corpus, staged uT3 N0 M0. What are the next steps: Laparoscopy? Primary resection? Perioperative chemotherapy?

2. 69-year-old man with localized adenocarcinoma at the esophago-gastric junction, intestinal subtype (Lauren), uT3 N+ M0: What are the next steps: PET-CT? Primary resection? Perioperative chemotherapy? Neoadjuvant radiochemotherapy?

3. 57-year-old man with oligometastatic adenocarcimoma of the esophago-gastric junction: uT2 N+ M1 (lymphatic): What are the next steps: Palliative chemotherapy? Curative resection? Multimodal therapy?

4. 75-year old man with metastatic gastric cancer, mixed type according to Lauren, multiple metastases in the liver and bone. What's next: molecular testing? Chemotherapy first-line? Which choice of first-line treatment?

5. 57-year old man with gastric cancer, diffuse type according to Lauren, treatment with fluorouracil, leucovorin, oxaliplatin, docetaxel (FLOT) first-line for 6 months. Then: progression with massive ascites. What's next: intraperitoneal therapy? Systemic treatment? Which choice of second-line treatment?

I invite you to join this expert seminar in order to solve these difficult as well as interesting cases!

Disclosure: Florian Lordick: Employment or Leadership Position: keines; Advisory Role: Ganymed, Biontech, Roche, Taiho, Lilly, Boston Biomedical; Financing of Scientific Research: an die Institution des Autors; Expert Testimony: GSK, Fresenius Biotech; Other Financial Relationships: Reisekostenterstützung für Kongresse: Roche, Bayer, Taiho, Merck.


NHL aggressiv - Konzepte bei B- und T-Zell Lymphomen

V362 - Treatment of aggressive B-cell lymphomas: R-CHOP or more?

Pfreundschuh M.1, Deutsche Studiengruppe für Hochmaligne  Non-Hodgkin-Lymphome

1Universität des Saarlandes, Innere Medizin I, Homburg, Germany

Young patients with no risk factor according to the age-adjusted (aa) IPI and no bulky disease have been shown in the MInT study to have a high event-free and nearly 100% overall survival after 6 cycles of R-CHOP-21, suggesting that some of these patients are overtreated. For the first time in the history of DLBCL treatment prospective trials like the FLYER study evaluate reduction of treatment from 6 to only 4 cycles of CHOP with 6 applications of rituximab. For young patients with aaIPI = 1 and/or bulky disease, best results have been achieved with 6xR-CHOP-21 plus involved-field radiotherapy to bulky disease or the more aggressive and more toxic R-ACVBP program without radiotherapy. For young poor-prognosis patients (aaIPI = 2,3) best results have been achieved with 8xR-CHOEP-14, with a 3-year overall survival of aaIPI = 2 patients of roughly 90%, demonstrating that this subpopulation is no high-risk population any more in the rituximab era. For young aaIPI = 3 patients there is still room for improvement and ongoing trials evaluate primary high-dose chemotherapy with stem cell transplantation or the addition of new drugs to a CHOP(E)P-14 backbone. Elderly patients who represent up to two thirds of all DLBCL patients do best with 8 cycles of R-CHOP-21 or 6 cycles of R-CHOP-14 plus 2 additional rituximab administrations. Since further intensification of chemotherapy is hardly possible in the elderly population, improvement strategies evaluate new doses and schedules of rituximab based on the recent demonstration that rituximab is suboptimally dosed in all DLBCL patients except elderly females. Other improvement strategies include vitamin D substitution in vitamin D insufficient and deficient DLBCL patients, because vitamin D deficiency impairs ADCC, the most important mechanism of action of rituximab, as well as new drugs targeting specific molecules involved in the signal transduction pathway of the B-cell receptor. There are several novel concepts in clinical trials, the most promising being BTK and BCL2 inhibitors, with BARs (B-cell receptor antigens for reverse targeting) providing an ultimate specificity restricted to the malignant B-cell clone. While the rate of patients dying from DLBCL has been cut into half during the last decade, these new approaches should further improve the cure rate of DLBCL in the near future.

Disclosure: Michael Pfreundschuh: Advisory Role: Advisory Board.

Wissenschaftliches Symposium


V365 - Cellular origin of CLL

Küppers R.1

1Universität Duisburg-Essen, Institut für Zellbiologie (Tumorforschung), Essen, Germany

In chronic lymphocytic leukemia (CLL), about half of the cases carry mutated immunoglobulin V region genes, and the other half are unmutated. Gene expression profiling studies revealed that both groups show a highly similar gene expression pattern. The cellular origin of CLL is much discussed and still not fully resolved. A derivation from CD5-positive B cells had originally been proposed. However, also marginal zone B cells, B cells activated in T independent immune responses and memory B cells are considered as more likely cellular origin of this most frequent leukemia in adults. We performed a refined gene expression profiling study of CLL cells in comparison to naive, mature CD5+, IgM+IgD+CD27+, IgM-only, class-switched memory and splenic marginal zone B cells. The evaluation of the data indicates that the gene expression program of both groups of CLL is most similar to mature CD5+ B cells, suggesting that CLL derive from these cells. We also identified a small subset of CD5+ B cells with somatically mutated immunoglobulin VH genes, which is characterized by expression of the memory B cell marker CD27. The mutated CLL tended to be more similar in their gene expression to these mutated CD5+ B cells than to unmutated CD5+ B cells, suggesting that mutated CLL originate from these germinal center-experienced CD5+ memory B cells. The proposed origin of CLL from mature CD5+ B cells is also supported by the detection of stereotyped immunoglobulin VH gene rearrangements among normal CD5+ B cells. Moreover, our study revealed a distinct gene expression pattern for mature CD5+ B cells, indicating that these lymphocytes represent a distinct B cell differentiation stage, and perhaps have specific functions in immune responses.

Disclosure: No conflict of interest disclosed.

V367 - Activation of autonomous signaling in CLL by mutual BCR interaction

Jumaa H.1

1Universitätsklinikum Ulm, Institut für Immunologie, Ulm, Germany

Introduction: Chronic lymphocytic leukemia (CLL) is a heterogeneous malignancy and is defined as a monoclonal expansion of morphologically mature but functionally incompetent B-lymphocytes with a distinct phenotype. Several mechanisms that mediate the malignant phenotype of CLL cells have been identified and suggest that both cell intrinsic factors as well as signals from the microenvironment contribute to the pathogenesis and prognosis of CLL. For instance, typical CLL-associated genetic alterations, which usually affect genes involved in cell cycle regulation or apoptosis, determine the course of the disease and subsequently more aggressive behavior. Increasing evidence strongly suggests an important role of the B cell receptor (BCR) in CLL etiology and since BCR signaling is usually activated by engagement through cognate antigen, it was assumed that external antigens induce BCR signaling and determine the survival of CLL B cells. We have recently shown that CLL-derived BCRs possess the unique capacity of autonomous signaling, which is mediated by mutual BCR interaction on the same cell.

Methods: We have performed crystallographic analyses of CLL-derived BCRs and found that the analyzed receptors undergo a specific and direct interaction with each other. We determined and subsequently mutated the crucial amino acid residues and expressed the resulting constructs in our TKO cellular reconstitution system described previously.

Results: We found that the original “wild-type” CLL-derived BCR is capable of inducing autonomous signaling in the reconstitution system for BCR expression, independent of external antigens. In contrast, however, the derivatives in which the amino acid residues which are crucial for interaction according to crystallography were mutated lost the ability for autonomous signaling. Treatment with anti-BCR antibody induced the signaling of the mutated version.

Conclusion: The autonomous signaling capacity is induced by direct interaction of BCR molecules in the membrane of the same cell and is characteristic for CLL. Identification of the crucial residues for this interaction might the design of specific reagent that may interfere with the autonomous signaling capacity and prevent CLL progression.

Disclosure: No conflict of interest disclosed.

Wissenschaftliches Symposium

Frühes Prostatakarzinom

V369 - Risk-adapted PSA screening - a way to reduce the harms of prostate cancer screening

Recker F.1, Randazzo M.1,2, Wyler S.F.1, Huber A.3, Grobholz R.4, Manka L.5, Kwiatkowski M.1,5

1Kantonsspital Aarau AG, Klinik für Urologie, Aarau, Switzerland, 2Universitätsspital, Zürich, Switzerland, 3Kantonsspital Aarau AG, Zentrum für Labormedizin, Aarau, Switzerland, 4Kantonsspital Aarau AG, Institut für Pathologie, Aarau, Switzerland, 5Akademisches Lehrkrankenhaus, Klinik für Urologie, Braunschweig, Germany

Introduction: PSA-Screening using a static PSA-cutoff for prostate biopsy (PBx) reduces prostate cancer (PCa)-specific mortality but is associated with a high rate of overdiagnosis of 50%. Treatment of overdiagnosed PCa may reduce men`s quality of life, known as “screening harms“. This in turn underlines the unmet need for a risk-based PSA-screening strategy. The current work shows the development of the Swiss PCa risk calculator ProstateCheck© in order to perform a “step-by-step” risk-adapted PSA-Screening.

Methods: From 1998 to 2012, 4932 men were prospectively observed (ERSPC Aarau, Switzerland). PSA-screening was undertaken every 4 years with a value of ≥3.0 ng/ml as indication for PBx. At baseline, men with 1-2.99 ng/ml and free-to-total PSA-Ratio (f/t-PSA) = < 20% also underwent PBx.

Primary endpoint was PCa diagnosis. Aggressive PCa was defined as intermediate and high-risk PCa according to d`Amico classification. Hosmer-Lemeshow-Test was used to count for model variables. The area under the curve (AUC) was calculated. The model was divided into a diagnostic (PSA > = 3 ng/ml) as well as a prognostic (PSA < 3 ng/ml) component.

Results: The best prediction model was achieved by using the following combined predictors: age, PSA-value, f/t-PSA, family history, prostate volume and digital rectal examination (DRE). The AUC was 0.74 (95% CI 0.69-0.80) for overall PCa diagnosis. For detection of aggressive PCa, AUC increased as follows: 0.72 (PSA), 0.80 (PSA+f/t-PSA), 0.82 (PSA+DRE), 0.87 (PSA+DRE+f/t-PSA), 0.88 (PSA+DRE+f/t-PSA+Age), 0.89 (PSA+DRE+f/t-PSA+age+family history+ prostate volume). Among men with baseline PSA < 3ng/ml, the incidence of aggressive PCa was recorded during follow-up. Using these frequencies, re-test intervals for subsequent PSA-screening according to baseline PSA were calculated for an individualized screening algorithm.

Conclusion: The Swiss PCa risk calculator ProstateCheck© can predict the probability of aggressive PCa with a high AUC (0.89) as compared to other risk tools (AUC of PCPT- /San Antonio 0.63, AUC Rotterdam 0.82). Aside from its diagnostic abilities, an individualized re-screening interval can be proposed which can save up to 70% of unnecessary PSA-tests and therefore reduce overdiagnosis. With this refinement, both screening and decision to perform PBx can be adjusted according to a man`s individual risk constellation leading to a more personalized medicine.

Disclosure: Franz Recker: No conflict of interest disclosed. Maciej Kwiatkowski: Advisory Role: Astellas und Myriad.

V371 - The PREFERE trial - the desire to prove the efficacy of treatment options for low risk prostate carcinoma

Weissbach L.1

1Stiftung Männergesundheit, Berlin, Germany

The German S3 guideline recommends in treating low risk carcinoma of the prostate: prostatectomy, irradiation, LDR brachytherapy, active surveillance. Considering these recommendations insufficiently substantiated, the initiators of PREFERE started the trial stating that ‘the recommendations of the S3 guideline appear to suggest that the four alternative treatments are equally effective in tumour control. This is definitively not the case. There are no conclusive data proving this assumption' (Michael Stöckle, principal investigator). However, the epidemiology of low risk prostate carcinoma as well as results of already published or other initiated RCTs comparing treatment options of localized prostate carcinoma may raise doubts if the clinical superiority of one or the other option can, or even need to be proven.

The mortality of low risk PCa (2% within 15 years subsequential early detection of the tumor) is very low, and reliable data concerning the mortality of prognostically particularly favourable low volume tumours are still lacking. Even a study population of 7,600 men is thus unlikely to yield statistically significant differences with PCa mortality as primary endpoint. In addition, results of PIVOT (though methodically not entirely flawless) show no significant advantage of radical prostatectomy as compared to no treatment. As PREFERE sets out to include only men with PSA ≤10 ng/ml the hope to find a survival benefit with one of the treatment modalities in comparison to any other of the available options seems highly questionable.

Experiences of already initiated studies indicate considerable recruitment difficulties: PIVOT recruited only 700 men instead of 5,000 planed for, START had to be terminated for lack of sufficient numbers, and PROTECT succeeded in recruiting only half of the anticipated study population. Obviously, available information about benefits and risks of available treatment options are sufficient to prompt potential candidates to refuse randomisation.

PREFERE's problems with recruiting patients do not imply a general impossibility to conduct RCTs nor do they exclude the potential of other study designs to answer questions of efficacy. They do however indicate the fact that potential patients as well as some treating urologists may have different views than the initiators of PREFERE concerning the need of comparative therapy studies of low risk PCa - an experience common to this and other large trials investigating its treatment

Disclosure: No conflict of interest disclosed.


Geriatrische Onkologie Hämatologische Erkrankungen im Alter 80+

V377 - Elderly patients with diffuse large B-cell-lymphoma

Zettl F.1, Hohloch K.2, Trümper L.3

1Klinikum Traunstein, Hämatologie/Onkologie und Palliativmedizin, Traunstein, Germany, 2Department Hämatologie und Onkologie, Kantonsspital, Chur, Switzerland, 3Klinik für Hämatologie und Medizinische Onkologie und Stammzelltransplantation, Universitätsmedizin, Göttingen, Germany

Introduction: Aggressive Lymphoma is a frequent disease in elderly patients, the median age at diagnosis is 66 years. Therapeutic decisions for elderly patients are difficult to make and often influenced by other factors like social status and physicians' experience and preferences. Raising numbers of comorbidities are complicating the treatment and management of elderly patients with aggressive lymphoma.

Results/Discussion: Poor risk factors identified in a SEER Database analysis in elderly patients are age above 80 years, pre-existing heartfailure and an increased comorbidity score. Nevertheless in elderly and comorbid patients the intention of treatment must be curative, the only curative treatment approach with a 5-year overall survival of approx. 75% is a combined immuno-chemotherapy including anthracyclines. The cumulative amount of antracylines given directly correlates with prognosis; however comorbidities are often the limiting factor of a full dosed CHOP chemotherapy. A dose reduced CHOP (mini-CHOP) in combination with rituximab has shown acceptable efficacy with reduced toxicity in patients not qualifying for fulldose therapy. The decision if such a regimen is feasible can only be made after a prephase therapy in individual patients. In addition, comorbidities and socioeconomic factors must be incorporated into the treatment plan in this patient population. Geriatric assessments may aid in the establishment of rational approaches. In common practice, Bendamustine is frequently being used in frail patients not qualifying for an antracyline based therapy. There is limited prospective data concerning the efficacy of this approach. Studies are ongoing.

Conclusion: Therapeutic options in elderly patients are very limited. Antracyclines are essential for a curative treatment strategy. Probably standardized geriatric assessments can help identify patients who are feasible for a more aggressive approach. The final choice of treatment has to be made by the treating physician in close interaction with the patient and his relatives.

Disclosure: Florian Zettl: Expert Testimony: Drittmittelföderung durch Roche und Mundipharma. Lorenz Trümper: Expert Testimony: Drittmittelföderung durch Roche und Mundipharma.

V378 - Patients with Multiple myeloma

Kleber M.1

1Clinic for Internal Medicine, University Hospital, Basel, Switzerland

Multiple myeloma (MM) is a disease which primarily affects elderly patients with a median age of 70 years. Over the last decade, novel therapeutic agents and enhanced supportive care improved survival in MM. Though, this benefit in survival appears to be confined to the young: the 5-year relative survival for MM patients younger than 65 years improved by more than 17% between 1998 and 2002 and between 2003 and 2007, but only 3.3% in the elderly (≥75 years). To date, the choice of therapy in MM patients is primarily based on chronologic age and performance status (PS). However, recent studies in MM highlight that the biological fitness of elderly patients shows relevant variations and assessment strategies are essential to define functional status, comorbidities and vulnerability to adverse events, which may increase treatment toxicity and drug discontinuation. A number of prognostic scores for evaluating comorbidity in cancer patients are available. Especially in MM specific comorbidity scores are proposed. E.g. the Freiburg Comorbidity Index (incl.: Karnofsky PS, renal and pulmonary disease status) is a reliable tool and highly predictive for outcome. Nevertheless, a systematic and evidence-based way of defining the global health and functional status is desirable to guide and objectify treatment decisions. In cancer patients a Comprehensive Geriatric Assessment (CGA) is considered to be the most appropriate way to evaluate the functional status, frailty and cognitive impairment. In MM patients the CGA is not routinely performed. Recently, Italian authors performed a brief geriatric assessment (GA) in 869 elderly patients with newly diagnosed MM. The GA consisted of 3 tools (ADL, IADL and Charlson Comorbidity Index). The results showed that a GA combined in a frailty score including age, comorbidities, cognitive and physical conditions can identify 3 groups of fit, intermediate fitness and frail patients with different 3-year OS of 84%, 76% and 57%, regardless of staging and treatment administered. In addition, frailty was associated with an increased risk of nonhematologic AEs and treatment discontinuation. Further studies in MM to validate determinants of therapeutic decision-making, in elderly-specific MM therapeutic and interventional trials with relevant outcomes beyond survival (e.g. effects of long-term toxicity on quality of life, independence, etc.) are needed. Algorithms for optimal therapy allocation in older patients will then be warranted.

Disclosure: No conflict of interest disclosed.

Wissenschaftliches Symposium

Stem Cell Biology - New Insights

V385 - Mathematical modelling of benign and malignant hematopoiesis: Potentials and insights

Roeder I.1

1Medizinische Fakultät Carl Gustav Carus an der Technischen Universität Dresden, Institut für Medizinische Informatik und Biometrie, Dresden, Germany

Systems biology, i.e. the application of mathematical models to quantitatively describe complex biological systems, is still a nascent discipline. However, even in the context of translational and clinical applications, the use of computational methods is becoming more and more accepted.

In my presentation I will discuss the potentials of mathematical modeling and simulation studies to broaden our understanding of hematopoietic stem cell organization. This comprises processes on the cellular level, such as cell proliferation or migration within the so called bone marrow stem cell niche, the tissue level, such as clonal competition, and last but not least, on the patient level, such as the prediction of treatment effects in chronic myeloid leukemia.

Using selected examples of modeling results, I will demonstrate how theoretical approaches can help to disentangle competing hypotheses about biological processes and to support the design of optimal experimental and/or clinical strategies. On the other hand, I will also point to potential pitfalls and limitations of in silico studies.

Disclosure: Ingo Roeder: Financing of Scientific Research: Bristol-Myers Squibb; Expert Testimony: Bristol-Myers Squibb.

V386 - Designer nuclease mediated gene modulation in stem cells

Cathomen T.1

1Universitätsklinikum Freiburg, Institut für Zell- und Gentherapie, Freiburg, Germany

Targeted genome editing with designer nucleases has become increasingly popular. The most commonly used designer nuclease platforms are meganucleases, zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs) and the clustered regularly interspaced short palindromic repeat (CRISPR)/Cas system. These powerful tools have greatly facilitated the generation of plant and animal models for basic research, and harbor an enormous potential for applications in biotechnology and regenerative medicine. However, the application of designer nuclease technology in human stem cells requires special considerations. Given that off-target cleavage activity of customized nucleases can potentially induce genotoxic side-effects, including the transformation of the target cell type, particular attention has to be paid to the specificity of designer nucleases. In my talk I will present data on preclinical applications of designer nucleases, highlight parameters that affect specificity of these nucleases, and end with a personal view on what aspects should be contemplated before moving into the clinic.

Disclosure: Toni Cathomen: Advisory Role: Beratungstätigkeit für TRACR Hematology Ltd.

Freier Vortrag

Kolon-/Rektumkarzinom I

V390 - Next-generation sequencing liquid biopsy strategy to monitor the emergence of EGFR antibody-resistant tumor subclones in gastrointestinal cancer

Braig F.1, März M.1, Schieferdecker A.1, Schulte A.2, Voigt M.1, Stein A.1, Grob T.3, Alawi M.4, Indenbirken D.5, Kriegs M.6, Engel E.7, Vanhoefer U.8, Grundhoff A.5, Loges S.1,9, Riecken K.10, Fehse B.10, Bokemeyer C.1, Binder M.1

1Universitätsklinikum Hamburg-Eppendorf, Onkologisches Zentrum, II. Medizinische Klinik und Poliklinik, Hamburg, Germany, 2Universitätsklinikum Hamburg-Eppendorf, Kopf- und Neurozentrum, Klinik und Poliklinik für Neurochirurgie, Hamburg, Germany, 3Universitätsklinikum Hamburg-Eppendorf, Zentrum für Diagnostik, Institut für Pathologie, Hamburg, Germany, 4Universitätsklinikum Hamburg-Eppendorf, Bioinformatics Service Facility (BIS), Hamburg, Germany, 5Heinrich-Pette-Institut, Leibnitz-Institut für experimentelle Virologie, Hamburg, Germany, 6Universitätsklinikum Hamburg-Eppendorf, Onkologisches Zentrum, Klinik und Poliklinik für Strahlentherapie und Radioonkologie, Hamburg, Germany, 7Hämatologisch-onkologische Praxis Altona (HOPA), Hamburg, Germany, 8Marienkrankenhaus, Zentrum für Innere Medizin, Hamburg, Germany, 9Universitätsklinikum Hamburg-Eppendorf, Zentrum für Experimentelle Medizin, Institut für Tumorbiologie, Hamburg, Germany, 10Universitätsklinikum Hamburg-Eppendorf, Onkologisches Zentrum, Klinik für Stammzelltransplantation (KMT), Hamburg, Germany

Introduction: The emergence of epidermal growth factor receptor (EGFR) ectodomain mutations as well as activating RAS mutations represents a clinical challenge in EGFR targeting of colon cancer. However, screening for such mutant subclones hasn't entered routine clinical practice yet.

Methods: 21 solid tumor samples of patients suffering from gastrointestinal cancer treated with chemotherapy +/- EGFR antibodies cetuximab or panitumumab were screened by next generation sequencing (NGS) for mutations in KRAS exon 2/3/4, NRAS exon 2/3/4 and the overlapping epitopes of both EGFR antibodies. Results were validated using circulating tumor DNA (ctDNA) of an independent cohort of 27 patients. Binding, signaling and drug sensitivity studies were performed in Ba/F3 cells to functionally characterize the EGFR G465R mutation.

Results: In these two patient cohorts that were in their majority still responsive to the combination of chemotherapy with EGFR antibodies, we found the EGFR G465R mutation in 7% and activating RAS mutations in 21% of cases. In line with previous data, this suggests that these mutations may be picked up even before overt clinical resistance occurs. NGS screening of ctDNA seemed to be more sensitive than NGS of post-treatment tumor material in the detection of such mutations, most likely because ctDNA more broadly reflects tumor heterogeneity. The EGFR G465R mutation was characterized for loss of cetuximab and panitumumab binding and for induction of cross-resistance in EGFR G465R transfected Ba/F3 cells.

Conclusions: We show that EGFR ectodomain mutations may also emerge in patients with gastrointestinal cancers other than colorectal cancer (e.g. cholangiocellular carcinoma) under the selective pressure of EGFR antibodies. These mutations can be easily screened for and monitored by a next-generation sequencing liquid biopsy strategy using ctDNA. This technology may compare favorably to BEAMing in terms of cost-effectiveness. Prospective studies should address what minimal mutational loads are required to faithfully predict lack of response to EGFR antibodies.

Disclosure: No conflict of interest disclosed.

V391 - Molecular profiling of single circulating colon cancer cells (liquid biopsy)

Liebs S.1, Keilholz U.2, OncoTrack Consortium

1Deutsches Konsortium für Translationale Krebsforschung (DKTK), Deutsches Krebsforschungszentrum (DKFZ), Partnerstandort Berlin, Berlin, Germany, 2Charite Universitätsmedizin Berlin, Berlin, Germany

Characterization of circulating tumor cells (CTCs) from tumor patient`s blood (liquid biopsy) represents a non-invasive approach for cancer profiling and treatment monitoring. We established a protocol for single CTC isolation based on the distinction of CD45-positive leukocytes and tumor cells expressing the epithelial markers EpCAM and/or carcinoembryonic antigen (CEA). The method is successfully implemented into the EU OncoTrack Project designed to identify novel biomarkers for colon cancer.

Prior to surgery, 50-60 ml of peripheral blood from patients with primary or metastatic colon cancer was collected. After red blood cell lysis, negative enrichment of CTCs was performed using the slightly modified EasySep(TM) human CD45 Depletion Kit protocol (StemCell). Immunofluorescent staining was used to detect and aspirate viable (LIVE/DEAD-negative), EpCAM/CEA-positive and CD45-negative CTCs by microscopic-assisted micromanipulation. Linear RNA amplification of the cell lysate and subsequent RT-qPCR was performed using CD45 and cytokeratin 19 (CK19), an intracellular tumor marker that is not downregulated during epithelial-to-mesenchymal transition.

Application of the protocol allowed CTC detection in 21 of 39 (54%) blood samples of patients with primary colon cancer and in 15 of 25 (60%) with metastases. A CTC detection rate of 88% was achieved in blood samples from patients undergoing surgery of the primary and metastatic tumor. The number of isolated cells varied between 1-4 cells (primary tumor) and 1-7 CTCs (metastatic disease) per blood sample. TaqMan and HybProbe PCR showed inconsistent results probably due to more successful amplification of smaller target regions on fragmented sample cDNA (e.g. CK19 amplicon length of 90 bp and 150 bp, respectively). Based on their positive CK19 and housekeeping gene status, 10 of 32 (31%) TaqMan or HybProbe-analyzed cells were defined as CTCs. Only 1 of 26 TaqMan-analyzed cells showed CD45 expression proving the accuracy of the cell picking. A panel deep sequencing protocol for targeted mutational analysis is under development.

CTC analysis holds tremendous potential for the identification of new drug targets and resistance-associated markers. Therefore, the introduced method represents a promising approach, which is not only adaptable to other tumor entities but also allows a variety of downstream applications.

Disclosure: No conflict of interest disclosed.

V392 - AIO-KRK-0109: A randomized phase II trial of panitumumab plus FOLFOXIRI or FOLFOXIRI alone as 1st-line treatment in RAS-wild-type metastatic colorectal cancer (mCRC)

Martens U.1, Weßendorf S.2, Riera Knorrenschild J.3, Büchner-Steudel P.4, Florschütz A.5, Atzpodien J.6, Keller R.7, Greeve J.8, Kanzler S.9, Ettrich T.10, Lindig U.11, Egger M.12, Hebart H.13, Geißler M.2, AIO

1SLK-Kliniken Heilbronn GmbH, Medizinische Klinik III, Heilbronn, Germany, 2Klinikum Esslingen GmbH, Klinik für Allgemeine Innere Medizin, Onkologie/Hämatologie, Gastroenterologie und Infektiologie, Esslingen, Germany, 3Universitätsklinikum Gießen und Marburg GmbH, Hämatologie, Onkologie und Immunologie, Marburg, Germany, 4Universitätsklinikum Halle (Saale), Uniklinik und Poliklinik für Innere Meidzin I, Halle, Germany, 5Städtisches Klinikum Dessau, Klinik für Innere Medizin, Dessau-Roßlau, Germany, 6Franziskus Hospital Nils-Stensen-Kliniken, Klinik für Interne Onkologie und Hämatologie, Georgsmarienhütte, Germany, 7AIO-Studien-gGmbH, Berlin, Germany, 8St. Vincenz Krankenhaus Paderborn, Medizinische Klinik I und Vincenz Darmzentrum, Paderborn, Germany, 9Leopoldina-Krankenhaus d. Stadt Schweinfurt GmbH, Medizinische Klinik II, Schweinfurt, Germany, 10Universitätsklinikum Ulm, Klinik für Innere Medizin I, Ulm, Germany, 11Universitätsklinikum Jena / Institut für Psychosoziale Medizin und Psychotherapie, Klinik für Innere Medizin I, Jena, Germany, 12Ortenau Klinikum, Hämatologie / Onkologie, Lahr, Germany, 13Stauferklinikum Schwäbisch Gmünd, Klinik für Innere Medizin, Mutlangen, Germany

Background: Patients with mCRC belonging to ESMO groups 1 and 2 are recommended to be treated with the most active applicable chemotherapy aiming on rapid and effective tumor responses. Group 1 patients may have the chance of a curative secondary resection of liver/lung metastases while group 2 patients have tumor symptoms and/or the presence of an aggressive tumor biology. The current study evaluates the efficacy of FOLFOXIRI plus panitumumab compared to FOLFOXIRI alone.

Methods: Prospective open-label 2:1 randomized Fleming's single stage phase II study using Irinotecan 150 mg/m2 d1; Oxaliplatin 85 mg/m2 d1; Leucovorin 200 mg/m2 d1;5-FU 3000 mg/m2 cont. 48 h; Panitumumab 6.0 mg/kg q2w (arm A) and Irinotecan 165 mg/m2 d1; Oxaliplatin 85 mg/m2 d1; LV 200 mg/m2 d1; 5-FU 3200 mg/m2 cont. 48 h q2w (arm B) up to 12 cycles. Primary endpoint was objective RR; secondary endpoints were secondary tumor resection rate, DCR, PFS, TTR, OS, toxicity, QL. To date 72/93 patients are recruited.

Results: A planned interim analysis was performed after 30 patients treated in Arm A to demonstrate a CR/PR in >18 of treated patients. 10 (33.3%) and 20 (66.7%) patients belonged to ESMO groups 1 and 2, respectively. Median age was 55.5 years (31-76 years). A median of 8.5 cycles FOLFOXIRI+Panitumumab were applied. The objective RR was 86.7% (n = 26). DCR was achieved in 30 patients (100%). A preplanned safety analyses after 10 patients in arm A did result in a dose adaption of irinotecan from 165 to 150 mg/m2 and 5-FU from 3200 mg to 3000 mg/m2. With this dosage no severe safety signals were observed compared to FOLFOXIRI alone although there was an increased number of SAE events..

Discussion: After dose modification of the FOLFOXIRI regimen the addition of panitumumab had a tolerable safety profile. A strong efficacy signal regarding RR and DCR was observed suggesting that FOLFOXIRI+panitumumab may be a prefered treatment for selected RAS wild-type patients belonging to ESMO groups 1+2. Further data will be presented.

Disclosure: No conflict of interest disclosed.

V393 - The colorectal carcinoma - treatment research and treatment reality in oncology practices

Valdix A.-R.1, Zaun S.2, Göttel R.3, Tessen H.-W.4,5

1Onkologische Schwerpunktpraxis, Schwerin, Germany, 2Sanofi-Aventis Deutschland GmbH, Berlin, Germany, 3rgb Onkologisches Management GmbH, Sarstedt, Germany, 4Onkologische Kooperation Harz, Goslar, Germany, 5Projektgruppe Internistische Onkologie (PIO), Goslar, Germany

Introduction: Which contribution does a continuous, systematic case documentation and evaluation provide to treatment research? What is the significance of new substances such as Aflibercept?

Methods: Data from oncology practices from 2003-2015 (PIO, ONCOReg).

7,802 histories of diseases comprising 12,623 palliative treatments originating from 123 oncology practices from 16 federal states are available for analysis. 12,446 treatments have been concluded so far and a response has been recorded in 11,655 of them.

Results: Patient characteristics

Gender: m: 60%, f: 40%

Distant metastatic diseases: 5,151 patients (85% lung/liver)

KRAS mutation status (n = 1,957): 63% wild type, 37% mutant

Median age at start of 1st-line therapy: 68 (19-92) years

Palliative chemotherapy (n = 5,076):


Median duration of treatment (n = 11,446): 119 days


Survival PFS/OS as of start of 1st-line treatment: 9.5/24.7 mths.

PFS/OS as of start of 2nd-line treatment: 6.4/15.1 mths.

27 practices treated 98 patients with aflibercept (87% with FOLFIRI).

Median age at start of 1st-line treatment: 74 years.

32% of patients received 2nd-line, 22% 3rd-line, 20% 4th-line.

97% of patients had been pretreated with oxaliplatin.

Median duration of aflibercept-based therapy (n = 65): 73 days


Conclusions: The data collected over a long period of time depict very precisely the reality of treatment in oncology practices in Germany. The analysis presented provides an important contribution to the complex of treatment research and answers a series of patient-related questions.

Aflibercept is applied according to its approved use. Current data will be presented.

Disclosure: No conflict of interest disclosed.

V394 - Implementation of RAS testing in patients with metastatic colorectal cancer in German outpatient cancer centers - data from the clinical TKK registry

Schnell R.1, Trarbach T.2, Broszeit-Luft S.3, Musch R.4, Frank M.2, Kopfmann S.2, Jänicke M.2, Marschner N.5, for the TKK Registry Group

1Praxis Internistischer Onkologie und Hämatologie, Frechen, Germany, 2iOMEDICO, Freiburg, Germany, 3Onkologische Praxis, Lehrte, Germany, 4Krebsheilkunde Lichtenberg, Berlin, Germany, 5Praxis für Interdisziplinäre Onkologie & Hämatologie, Freiburg, Germany

Introduction: Since 2008 KRAS mutation testing is mandatory before applying an anti-EGFR monoclonal antibody (mab) in patients (pts) with metastatic colorectal cancer (mCRC). Since 2014 expanded RAS mutation testing including exons 2-4 of the KRAS gene as well as exons 2-4 of the NRAS gene is required.

Methods: Starting in 2006, the Tumor Registry Colorectal Cancer (TKK) prospectively documents treatment of pts with CRC by office-based medical oncologists in Germany. Pts receive standard treatments according to physician's choice. The registry collects data on biomarker testing, all systemic treatments, outcome, treatment decision-making as well as pts and tumor characteristics. Currently (March 2015), 166 sites have recruited over 5500 pts with CRC, of which about 3700 have mCRC. Information on mutation testing has been collected for KRAS status since 2008 (n = 2264) and for RAS since 2014 (n = 226).

Results: The number of pts tested for KRAS mutation before the start of first-line treatment has been increasing since 2008 (47% in 2008, 86% in 2015). NRAS status was determined in 58% of pts in 2014, and in 63% in early 2015. These results correlate with the increasing number of pts tested for both RAS mutations (58% in 2014, 62% in 2015).

Pts who are not tested for RAS mutation are older (70 vs. 67 years) and more often affected by a concomitant disease (83% vs. 72%) compared to pts tested for RAS mutation.

The main reason for not testing pts' RAS status before the start of first-line treatment is “no intention to use an anti-EGFR mab”, according to the attending oncologists.

While 54% of pts tested were KRAS wild-type (wt) in 2014, this number slightly decreased to 48% in 2015, presumably because of more exons are being tested. This trend is also seen in pts tested for NRAS mutation (89% (n = 156) wt in 2014 and 83% (n = 46) in 2015). Of the 226 pts tested for both, NRAS and KRAS mutation, 69% were RAS wt in 2014 and 57% in 2015.

Of all pts with RAS wt status, 51% were subsequently treated with anti-EGFR mab-based regimen as first-line treatment, most frequently with FOLFIRI and cetuximab (38%). The 49% of pts not treated with an anti-EGFR mab, most frequently received FOLFIRI and bevacizumab (28% of RAS wt pts).

Conclusion: Testing for RAS mutation has been rapidly implemented in German outpatient cancer centers with almost all pts being tested for KRAS and an increasing number of pts being tested for NRAS.

Disclosure: No conflict of interest disclosed.

V395 - Relevance of neo- or adjuvant pretreatment in patients with metastatic colorectal cancer treated with FOLFIRI plus bevacizumab as first-line treatment - data from the prospective TKK registry

Schnell R.1, Trarbach T.2, Broszeit-Luft S.3, Musch R.4, Wetzel N.2, Kopfmann S.2, Jänicke M.2, Marschner N.5, for the TKK Registry Group

1Praxis Internistischer Onkologie und Hämatologie, Frechen, Germany, 2iOMEDICO, Freiburg, Germany, 3Onkologische Praxis, Lehrte, Germany, 4Krebsheilkunde Lichtenberg, Berlin, Germany, 5Praxis für interdisziplinäre Onkologie & Hämatologie, Freiburg, Germany

Background: A large number of patients (pts) diagnosed with metastatic colorectal cancer (mCRC) have received neo- or adjuvant treatment for initially localized CRC. The influence of such pretreatment on the outcome after diagnosis of metastasis is still under debate. While clinical trials recruit highly selected pts, clinical registries collect data on treatment in routine practice, representing all pts and therapies. This allows analysis of pretreatments and their impact on outcome in real-life pts.

Methods: Starting in 2006, the Tumor Registry Colorectal Cancer (TKK) prospectively documents treatment of pts with CRC by office-based medical oncologists in Germany. Pts receive standard treatments according to physician's choice. A broad set of data including pts and tumor characteristics, all systemic treatments and outcome are collected. At the time of this interim analysis (March 2014), 122 sites had recruited 5024 pts with CRC; 2466 pts received treatment for mCRC.

Here, outcome data such as progression free survival (PFS) and overall survival (OS) of pts treated with the most frequently applied first-line treatment FOLFIRI and bevacizumab (FOLFIRI+BEV, 24% [n = 588]) were estimated. Progression was reported by study sites as per local standard. The outcome was compared between pts pretreated with neo- or adjuvant chemotherapy (pretreated pts) and treatment naïve pts (naïve pts).

Results: At the start of first-line treatment, 36% (n = 211) of pts receiving FOLFIRI+BEV were pretreated (64% naïve). Pts characteristic were comparable between pretreated and naïve pts: 65% of pts were male (vs. 68%), mean age was 64 years (vs. 64 years), 71% of pts had a concomitant disease (vs. 62%).

According to the attending physician, 50% of pretreated pts had a complete or partial response (vs. 56%). Median first-line PFS of pretreated pts was 10.9 months (95% confidence interval [CI] 10.1-11.6) and median OS was 24.7 months (95% CI 19.8-29.7). Without neo- or adjuvant pretreatment, median first-line PFS was 10.8 months (95% CI 9.8-11.7) and median OS was 23.4 months (95% CI 21.0-25.8).

Conclusions: In routine practice, median OS and PFS of mCRC pts treated first-line with FOLFIRI+BEV do not differ between pts pretreated with neo- or adjuvant chemotherapy and pts who were not. Median OS and PFS of FOLFIRI+BEV are comparable to data from clinical trials although pts are older and more often affected by comorbidities.

Disclosure: Roland Schnell: No conflict of interest disclosed. Norbert Marschner: Financing of Scientific Research: Vortragshonorare von Bayer AG, Merck Pharma GmbH, Roche Pharma AG.


Transplantation Spendersuche GvHD Nachsorge

V399 - Acute and chronic GVHD: State of the art therapy

Greinix H.T.1

1Medizinische Universität Graz, Hämatologie, Graz, Austria

GVHD has remained a serious and life-threatening complication of allogeneic hematopoietic cell transplantation (HCT).Calcineurin inhibitors (CNI) and methotrexate (MTX) or mycophenolate mofetil (MMF) have been the established GVHD prophylaxis for many years. Sirolimus and CNI demonstrated equal protection from GVHD with less mucosal side effects. Use of antithymocyte globulin in combination with CNI and MTX resulted in significantly reduced incidences of acute GVHD grades III-IV and chronic GVHD with no difference in survival. Recently, post-transplant cyclophosphamide was successfully investigated not only after haploidentical but also HLA-identical related and unrelated donor HCT. Established first-line therapy of acute GVHD consists of corticosteroids at 2 mg/kg/day and no advantages of higher steroid doses have been reported. However, in acute GVHD grade IIa steroid doses of 0.5 mg/kg/day were effective whereas in grades ≥IIb doses of 1 mg/kg/day increased the need for secondary immunosuppressive treatment and thus, are not recommended. Response to first-line therapy assessed on day 14 is significantly correlated to NRM and survival. Second-line therapy of acute steroid-refractory GVHD is indicated after 3 days with progression, 1 week with persistent unimproving grade III GVHD or after 2 weeks with persistent unimproving grade II GVHD. Currently, no standard salvage treatment options for steroid-refractory acute GVHD patients are available and choice of therapy should be guided by potential toxicity, infections and interactions with other agents. Due to its efficacy and excellent safety profile ECP is a valuable second-line treatment option. Patients with NIH-defined moderate or severe chronic GVHD should receive systemic immunosuppressive treatment whereas in mild chronic GVHD topical therapy is preferable. Established first-line treatment of chronic GVHD consists of corticosteroids with or without CNIs. In case of progression under 1 mg/kg/day of steroids for 2 weeks or stable disease on ≥0.5 mg/kg/day for 4 to 8 weeks salvage therapy should be considered. ECP is recommended for second-line treatment based on efficacy in prospective studies and lack of general immunosuppression. Other salvage therapies include mTOR-inhibitors, MMF, imatinib and rituximab.Supportive care including prophylaxis of infections and osteoporosis, vaccinations, psychosocial and dietary support and rehabilitation are of major importance for patients' outcome.

Disclosure: Hildegard Greinix: Financing of Scientific Research: Therakos.


AYA Adoleszente und junge Erwachsene - was ist anders?

V400 - What are the psychosocial challenges after cancer acute treatment for adolescent and young adults?

Leuteritz K.1

1Universitätsklinikum Leipzig AöR, Department für Psychische Gesundheit, Abteilung für Medizinische Psychologie und Medizinische Soziologie, Leipzig, Germany

Introduction: In Germany, about 3% of all cancer diagnoses per year are among adolescent and young adults (AYA). AYAs are dealing with a serious disease at a complex psychosocial stage in life, including development tasks like detachment from the parental home, establishing financial and social independence, the formation and consolidation of a partnership and family as well as starting a professional career. Workingship addresses many aspects of cancer patients´ life satisfaction. The psychosocial impact of cancer in this age group is rarely represented.

Methods: A total of N = 354 diagnosed young adult cancer patients who were aged 18-39 years at the time of survey and had completed their acute treatment (range 0 to 6 month) answered a questionnaire, including standardized instruments and self-developed items focusing on life satisfaction, the supply situation and the need for support in addition to medical, sociodemographical and work specific variables. Life satisfaction was measured with the questionnaire for life satisfaction (FLZ) who captures 10 relevant aspects of life. For analysis we considered patients who were “at work/school” before diagnosis (N = 190) and not unemployed/disabled.

Results: Over half of the responding AYAs working/in school before diagnosis (N = 190, age: 28,8 years, men: 29,6%) are satisfied with their life situation (51,7%). They experience most changes after cancer acute treatment in areas of healthiness (61%) followed by changes in terms of leisure activities (49%), professional life (48%) and family planning (46%)as well as financial situation (42%). Dissatisfaction among AYA patients especially concerns areas of family planning (43%), financial situation (42%), sexuality (35%) and professional life (33%). Most satisfaction exists regarding apartment situation, relatives, friends and partnership. Higher life satisfaction is associated with not having operation, working part time before diagnosis, less changes in income since diagnosis as well as low psychological distress and having social support. Sex, diagnosis, number of medical therapies had no significant impact (R²= 0,79).

Conclusions: Despite a high life satisfaction after cancer acute treatment more than every third AYA patient reports about dissatisfaction in issues like family planning, financial situation, sexuality and professional life. A future task must be to develop specific interventions in these areas for this age group and their specific problems in Germany.

Disclosure: No conflict of interest disclosed.

V402 - Back to normal life - should a specialized rehabilitation be offered?

König V.1

1Klinik Bad Oexen, Bad Oeynhausen, Germany

Intensified treatment strategies are used in cancer therapy of adolescents and young adults (AYA) to improve treatment results. Therefore, functional impairments and therapy-related disorders are often more severe than those observed in older cancer patients. Cancer incidence rates are lower in AYA, therefore young adults are underrepresented in acute care hospitals compared to older adults. In AYA, psychological distress is higher after cancer therapy as professional training and / or partnership planning are incomplete. These factors should be taken into account in aftercare planning. In-patient rehabilitation in a group setting promises several advantages: Physical recovery is enhanced by the motivational effects among the group members competing against one another. Under professional supervision, the direct exchange among peers is promoted to the advantage that the psychological group meetings have co-therapeutic effects. Additionally, one-on-one consultations are available for topics that have to be dealt with individually, for example therapy-related infertility. Moreover, reintegration into work life, necessary change of occupation with additional career guidance etc. can be addressed specifically in this setting. Thus, age-specific group rehabilitation within the framework of a specialist clinic is an additional therapeutic tool in the aftercare treatment of juvenile cancer patients providing accelerated recovery from cancer.

Disclosure: No conflict of interest disclosed.


Ösophaguskarzinom Therapiemodalitäten State of the Art

V403 - State of the art - Treatment of localized oesophageal carcinoma according to tumor stage

Stahl M.1

1Kliniken Essen-Mitte, Internistische Onkologie und Hämatologie, Essen, Germany

Introduction: Patients with carcinomas of the oesophagus and oesophago-gastric junction (OGJ) presenting with localized disease are potentially curable. But cure rates and intensity of therapy are different and strictly dependent on tumour extension.

Statements: (1) Carcinomas limited to the mucosa (T1): In tumours without risk factors (signet ring carcinoma, G3, ulceration) complete endoscopic resection is the therapy of choice with high cure rates in mucosal (m1-m3) cancers and also superficially submucosal (sm1 < 500müm) adenocarcinomas (G1/2, <20 mm, L0, V0). (2) Localized early cancer (T1sm-2): sm2-3 carcinomas have a risk of lymphatic spread in up to 50%. This is even higher in tumours of T2-category. These tumours should therefore be treated with surgical resection. However, recurrences will occur in up to 70% after complete resection if lymph nodes proved to be involved (pN1-3). Preoperative chemoradiotherapy (CRT) showed to significantly improve local tumour control but not overall survival. However, this procedure is limited by the operative risks particularly in patients with squamous cell cancer (SCC), where relevant numbers of the patients may not tolerate transthoracic oesophagectomy. In these patients definitive chemoradiotherapy can be a treatment option with curative intent. Even more, if the primary tumour is located in the cervical or upper intrathoracic part of the oesophagus. (3) Locally advanced cancer (T3-4): Multimodality approaches are reflecting the state of the art in this clinical situation with surgery proved superiority to other approaches regarding local tumour control and survival. But operative mortality of about 10% must be realized in SCC patients. Therefore, definitive chemoradiation showed similar survival in randomized trials, although local tumour control was hampered by omitting surgery. It remains a matter of debate in adenocarcinomas of the OGJ whether and when preoperative chemoradiotherapy should be preferred to perioperative chemotherapy. Adjuvant therapy appears to be restricted to patients with understaged tumours of pT3-4pN1-3 category where primary surgery has been performed erroneously.

Disclosure: No conflict of interest disclosed.

V404 - Induction chemotherapy - pro arguments

Ruhstaller T.1

1Onkologie, St. Gallen, Switzerland

Induction chemotherapy is defined as two cycles of chemotherapy before the neoadjuvant or definitive chemoradiation. The specific question of an induction chemotherapy before CRT in oesophageal cancer has hardly been investigated in clinical studies so far, however, this approach has been used in a lot of studies with other focus and it is daily practice in a lot of centres.

There are several arguments which militate in favour of induction chemotherapy. The treatment can usually be started suddenly after the diagnosis ( no waiting time for planning the radiotherapy). Most patients are presenting by diagnosis with relevant dysphagia. A direct start with radiotherapy is usually worsening the dysphagia and makes often feeding tubes necessary. With induction chemotherapy the dysphagia is often very fast improving in such a way, that patients are able to eat quite normally when the CRT has to start. As a conclusion less patients need feeding tubes during radiottherapy. Last but not least, most patients die of systemic disease and we hope to better target microscopic disease with induction chemotherapy, however, this has not yet be proven.

For this reason we are eagerly awaiting new trials with specific focus on induction chemotherapy.

Disclosure: No conflict of interest disclosed.

V405 - Inductionchemotherapy prior to radiochemotherapy and operation in esophageal cancer - the CONTRA position

Thuss-Patience P.1

1Charite Universitätsmedizin Berlin, Med. Klinik m S Hämatologie, Onkologie und Tumorimmunologie, Berlin, Germany

Due to the dismal prognosis of esophageal cancer we feel the need to maximize therapy as long as possible with the hope to also maximize efficacy. The rationale for induction-chemotherapy prior to radiochemotherapy and operation is based on the hope to maximize the effect on micrometastases which are not covered in the radiation field. In my talk I will critically check whether there are any reliable data which support this feeling and whether induction chemotherapy really translates into any clinical benefit for the patient.

Disclosure: Peter Thuss-Patience: Advisory Role: Roche, Lilly, Nordic; Expert Testimony: GSK.

V406 - What is the optimal chemotherapy for neoadjuvant and definitive chemoradiotherapy?

Eisterer W.1

1Medizinische Universität Innsbruck, Univ.-Klinik für Innere Medizin I, Internistische Onkologie, Innsbruck, Austria

With an increasing incidence and overall 5-year survival of about 15%, the prognosis of esophageal cancer (EC) patients remains poor. Neoadjuvant chemoradiation (nCRT) has shown to be superior compared with surgery alone leading to the current standard procedure in medically fit esophageal carcinoma with curative resectable EC. In patients who are not eligible for curative intended surgery due to technical reasons or medically unfit for resection definitive chemoradiotherapy (dCRT) respresents an alternative option.

The most commonly used regimens are those consisting of cisplatinum in combination

with 5-fluorouracil (5FU) or paclitaxel combined with carboplatin.

Current guidelines in the United States and Europe recommend the combination of cisplatinum with 5-FU as standard combined with 50.4 Gy radiation therapy [1] while the carboplatin/paclitaxel regimen is frequently used in patients with extensive comorbidity [2].

A comparison of two nCRT regimens in patients with potentially curable EC proved that the carboplatin/paclitaxel/41.4 Gy regimen caused less toxicity compared with the cisplatin/5-FU/50.4 Gy regimen, with an insignificant difference in response rates and long-term survival [3].

In the Dutch CROSS Trial [4] paclitaxel and carboplatin weekly for 5 weeks with 41.4 Gy radiotherapy led to a significant survival benefit compared to surgery alone; nCRT resulted in 23% pathologic complete Response, less locoregional relapse and no compromise of the surgical resection.

A retrospective analysis of dCRT with carboplatin/Paclitaxel versus cisplatin/5FU in 102 patients [5] showed similar survival between the two regimes. Carboplatin/paclitaxel was completed more frequently (82% vs 57%) and showed less hematologic and non-hematologic toxicity (grade 3 or higher 4% vs 18%).

Conclusion: Carboplatin/paclitaxel represents a valid alternative to cisplatin/5FU. Further development of taxane-based CRT schedulesare warranted.


1 Stahl M et al.: Ann Oncol 2010; 21(Suppl 5):v462.

2 National Comprehensive Cancer Network. Clinical Practical Guidelines in Oncology. Esophageal and Esophagogastric Junction Cancers Version 2.2013. 2013:ESOPH-E 6.-v49.

3 Blom RL et al.: Dis Esophagus 2014;27:380-387.

4 van Hagen P et al.: N Engl J Med 2012;366:2074-2084.

5 Honing J et al.: Ann Oncol 2014;25:638-643.

Disclosure: No conflict of interest disclosed.



V408 - Medication safety from a pharmaceutical perspective

Jaehde U.1

1Institute of Pharmacy, University of Bonn, Clinical Pharmacy, Bonn, Germany

Cancer patients are exposed to a variety of therapy-associated risks when treated with anticancer drugs. Due to the high toxicity and complexity of most therapeutic regimens there is a high incidence of drug-related problems such as adverse effects, drug-drug interactions, and non-adherence. Therefore, there is a strong need for specific interventions assuring medication safety. The provision of a structured medication management is a novel approach to reduce the incidence of adverse events and enhance medication adherence.

In our working group, we have developed several interventions to increase the medication safety of cancer patients. For example, we have shown that a structured medication management can reduce the incidence of nausea and emesis in patients with breast and ovarian cancer [1]. Moreover, a prospective two-arm cohort study revealed the potential of an adherence management for patients receiving the orally administered anticancer drug capecitabine [2]. Recently, we have developed a novel modular medication management for outpatient

cancer care in a multiprofessional quality circle consisting of oncologists, pharmacists and nurses defining the tasks and responsibilities of each profession. All care modules include evidence-based recommendations for supportive care, written patient information, and an algorithm illustrating the care process. Care modules were developed for medication review and interaction check, malnutrition, and for the management of four common adverse events: nausea/emesis, mucositis, fatigue, and pain. They can be applied individually for each patient according to medication and anticipated toxicity. The feasibility of the model has recently been evaluated in a randomized two-arm interventional trial [3].

In conclusion, a structured medication management can reduce the incidence of drug-related problems and hence improve medication safety. Multiprofessional approaches, including at least oncologists, pharmacists and nurses, are most promising with regard to implementation in daily routine.


1 Liekweg A, Westfeld M, Braun M, Zivanovic O, Schink T, Kuhn W, Jaehde U. Support Care Cancer 2012;20:2669-2677.

2 Krolop L, Ko YD, Schwindt PF, Schumacher C, Fimmers R, Jaehde U. BMJ Open 2013;3:e003139.

3 Wilmer A, Jansen C, Ko YD, Schmidt-Wolf I, Jaehde U. FORUM 2014;29:324-330.

Disclosure: No conflict of interest disclosed.

V410 - Learning from errors: Clinical risk management in the oncology

Paula H.1

1Inselspital, Universitätsspital Bern, Ärztliche Direktion, Qualitätsmanagement, Bern, Switzerland

Introduction: Learning from errors is one of the most common principles in the clinical risk management. Indeed it is undisputable important to detect and understand the reasons why things are going wrong. Especially in high-risk disciplines the knowledge about these facts is the key to avoid adverse events. Usually in hospitals the emergency medicine, operating theatres, anaesthesia and intensive care are reckoned to this category. The hazardous nature of oncology is frequently underestimated in this context. Highly potential therapies and vulnerable patients are a very dangerous mixture. Therefore it is no exaggeration to declare oncology also as a high-risk discipline.

Methods: In health care the most activities are based on human decisions and performances. In the case of an adverse event it is thus on the first sight mostly no problem to identify one or more persons which have caused an error. This leads into ignoring other important facts. At a closer look it is feasible to detect error promoting elements in the environment or in the organisational structure. There are a few established and approved methods to achieve this goal. More than mortality and morbidity conferences or root-cause-analyses incident reporting is widely-used. CIRS (Critical Incident Reporting System) means to collect incidents, which have caused no harms (“near misses”) with the purpose of the identification of error promoting factors.

Results: Despite of the worldwide distribution of incident reporting systems a lot of the referred problems are still unresolved. Theoretically enough information are available to manage the reported risks. In reality there are still a lot of unanswered questions. One factor might be the conventional way of interpreting critical incidents. Perhaps it is more promising not to ask what was going wrong, but to ask which factors are usually responsible for an error-free treatment.

Conclusions: In the foreseeable future incident reporting remains as an important tool. However, the system should be critically scrutinized on existing opportunities for improvements. One possible approach is to focus on factors that are normally ensuring the error free treatment. This so called “Safety II” (according to Hollnagel et al) might be a useful strategy to enhance patient safety. It is quite possible to use CIRS as an information source in this context. But it is necessary to direct the focus on other facts and to ask questions that never have been asked before.

Disclosure: No conflict of interest disclosed.


Hämostaseologie II

V412 - Screening for occult cancer after idiopathic VTE - not too much and not too little

Matzdorff A.1

1Asklepios Klinikum Uckermark, Mediz. Klinik II, Hämatologie, Onkologie, Schwedt, Germany

Cancer can trigger thromboembolism. There is a 4-10% chance of finding an asymptomatic occult cancer in patients with idiopathic venous thromboembolism. So far guidelines recommend limited cancer screening with history taking, physical examination, and screening examinations as recommended by age. Studies show that a more extensive screening program, including endoscopy and computed tomography finds more cancers at an early stage with a better chance of cure.

While extensive cancer screening detects more cancers it has, as of yet, not been shown to reduce overal mortality. More arguments against extensive cancer screening are the increased risk of false-positive results, which lead to additional diagnostic interventions with more side effects and complications and higher costs.

It is highly unlikely that there will be another randomized study on limited vs. extensive cancer screening large enough to show an effect on mortality or cost-effectiveness. Considering the clinical need for advice and guidance on this topic and the consequences for the individual patient when missing a cancer diagnosis the following screening program for patients with idiopathic VTE is suggested:

Medical history including risk factors for cancer,

Physical examination including rectal exam,

Complete blood count, electrolytes, liver and renal function tests, urinalysis, and with atypically localized thromboses (e.g. abdominal vein thromboses) also JAK2 and CALR mutation screening,

Women: breast exam*, mammography*, pelvic exam*, PAP-smear*,

Men: prostate exam*, transrectal ultrasound*, PSA* (PSA never alone, always combined with the aforementioned exams),

Tumormarker-Screening (CEA, CA 12-5, αFP, never alone, only following positive findings from other studies to narrow the differential diagnosis),

New recommendation: gastroscopy and colonoscopy*,

New recommendation: CT or - if available - MRI of abdomen and pelvis,

New recommendation: low-dose chest CT, (* if not already received as part of prior cancer screening.)

The pros and contras of this approach will be discussed in the presentation.

Disclosure: No conflict of interest disclosed.

V413 - Anticoagulation in liver cirrhosis

Langer F.1

1Universitätsklinikum Eppendorf, II. Medizinische Klinik und Poliklinik, Hamburg, Germany

The coagulopathy of liver cirrhosis is characterized by downregulation of both procoagulant (e.g. fibrinogen and clotting factors II, V, VII and X) and anticoagulant factors such as antithrombin (AT), protein C and protein S. Consequently, even under conditions of severe hepatic dysfunction, the hemostatic system in patients with chronic liver disease is typically balanced with regard to pro- and anticoagulant pathways. Commonly used coagulation tests such as the prothrombin time (PT) and the activated partial thromboplastin time (APTT), however, do not assess the function of coagulation inhibitors. In particular, the PT is not influenced by a defect in the activated protein C pathway, and the spontaneously prolonged PT in patients with liver cirrhosis has therefore long been interpreted as a laboratory sign of systemic hypocoagulability. More global coagulation tests such as calibrated automated thrombography (CAT) in the presence of soluble thrombomodulin, however, have clearly shown that thrombin generation in plasmas from cirrhosis patients is not significantly decreased when activation of the protein C system is enabled. In fact, plasma thrombin generation capacity may even be increased in patients with liver cirrhosis, and vascular thrombotic events are increasingly recognized in this patient population. Although cirrhosis patients may experience severe bleeding symptoms, mainly as a result from portal hypertension, endothelial dysfunction, infection and/or uremia, the coagulopathy of liver cirrhosis in itself does not preclude systemic anticoagulation, especially, since accompanying thrombocytopenia is typically counterbalanced by massively increased plasma levels of von Willebrand factor. The use of vitamin K antagonists (VKAs) in cirrhosis patients is hampered by the spontaneously prolonged PT, which is partially due to clotting factor V deficiency. The INR may therefore not adequately reflect the antithrombotic effects of VKAs. In this regard, the direct oral anticoagulants rivaroxaban, apixaban and dabigatran may have several pharmacological advantages, but their use in patients with severe hepatic and/or renal dysfunction is limited. For this reason, low-molecular heparins still play a major role in the prevention and treatment of thromboembolic events in cirrhosis patients, but sufficiently high AT plasma levels are required for these indirect agents to exert there anticoagulant effects.

Disclosure: Florian Langer: Advisory Role: Baxter, Bayer, Bristol-Myers Squibb, CSL Behring, Daiichi-Sankyo, LEO, Novartis, Novo Nordisk, Pfizer; Financing of Scientific Research: Baxter, Bayer, Boehringer Ingelheim, Bristol-Myers Squibb, CSL Behring, Daiichi-Sankyo, LEO, Novartis, Novo Nordisk, Pfizer, Sanofi; Expert Testimony: CSL Behring, Pfizer.

Freier Vortrag

Lymphome indolent

V418 - Bendamustine plus Rituximab (B-R) versus CHOP plus Rituximab (CHOP-R) as first-line treatment in patients with indolent lymphomas or mantle cell lymphomas (MCL) - 7-year updated results from the StiL NHL1 study

Rummel M.1, Maschmeyer G.2, Ganser A.3, Heider A.4, von Grünhagen U.5, Losem C.6, Heil G.7, Welslau M.8, Balser C.9, Kaiser U.10, Weidmann E.11, Dürk H.12, Balló H.13, Stauch M.14, Barth J.1, Blau W.1, Burchardt A.1, Kauff F.1, Brugger W.15, StiL (Studiengruppe indolente Lymphome)

1Universitätsklinik, Med Kl IV, Hämatologie, Gießen, Germany, 2Ernst von Bergmann Klinikum, Potsdam, Germany, 3Medizinische Hochschule Hannover, Hannover, Germany, 4Klinikum Leverkusen gGmbH, Leverkusen, Germany, 5Schwerpunktpraxis, Cottbus, Germany, 6Facharztzentrum, Neuss, Germany, 7Klinikum Lüdenscheid, Lüdenscheid, Germany, 8Phase drei, Hämato-Onkologischer Studienkreis am Klinikum Aschaffenburg, Aschaffenburg, Germany, 9Gemeinschaftspraxis, Marburg, Germany, 10St. Bernward Krankenhaus, Hildesheim, Germany, 11Krankenhaus Nordwest, Frankfurt/Main, Germany, 12St. Marien-Hospital Hamm gGmbH, Hamm, Germany, 13Gemeinschaftspraxis, Offenbach, Germany, 14Hämatologisch-Onkologische Schwerpunktpraxis, Kronach, Germany, 15Schwarzwald-Baar Klinikum, Villingen-Schwenningen, Germany

Introduction: This multicenter, randomized, phase III study compared B-R and CHOP-R as first-line treatment in patients (pts) with indolent lymphomas or MCL and was published in The Lancet in 2013. The final published analysis at a median follow-up of 45 months demonstrated a significantly prolonged progression-free survival (PFS) in the B-R group, compared to the CHOP-R group (p < 0.001). Median PFS was 69.5 vs. 31.2 months, respectively. In the current analysis, we present updated results for overall survival (OS), time-to-next-treatment (TTNT), and secondary malignancies (sNPL) with a median follow-up of 87 months.

Methods: 549 pts with indolent lymphomas or MCL were randomized to receive B-R or CHOP-R for a maximum of 6 cycles. The primary endpoint was PFS; secondary endpoints included OS, TTNT, and sNPL.

Results: 514 randomized pts were evaluable (261 B-R; 253 CHOP-R). Patient characteristics were well balanced between arms; median age was 64 years.

Fewer pts treated initially with B-R needed second-line treatments due to disease progression compared to CHOP-R treated pts: 93 pts (36%) in the B-R group received salvage treatment compared with 140 pts (55%) in the CHOP-R group. Of those in the CHOP-R group, 69 pts (49%) received B-R as salvage. TTNT was significantly prolonged with B-R compared with CHOP-R (p < 0.001). Median TTNT was not yet reached in the B-R group vs. 42.3 months in the CHOP-R group.

The difference in complete response (CR) rates (independent of treatment arms) between male (n = 272, median age 63 years) and female (n = 242, median age 64 years) pts was statistically significant: 28.6% for male pts versus 42.1% for female pts (p = 0.0016). Female pts had a longer median TTNT compared to male pts (not yet reached vs. 52.2 months, respectively; p = 0.006). The achievement of a CR was associated with significantly prolonged OS, with an estimated 10-year survival rate of 72.6% for pts with a CR and 63.6% for pts with a partial response (p = 0.006).

The difference in OS between the treatment arms was not statistically significant. The estimated 10-year survival rates were 67.4% for B-R and 60.1% for CHOP-R (p = 0.262). In pts with indolent lymphomas (without MCL), there was a trend toward a longer survival for the B-R group compared with the CHOP-R group, with 43/215 pts (20.0%) in B-R and 58/205 pts (28.3%) in CHOP-R. The estimated 10-year survival rates for pts with indolent lymphomas were 71.9% for B-R and 61.5% for CHOP-R (p = 0.076). No difference in OS was found in the subgroup of pts with MCL (n = 95; p = 0.429).

Twenty sNPL were observed in the B-R group compared with 23 in the CHOP-R group, with 1 hematological malignancy in each group to date.

Conclusions: In pts with previously untreated indolent lymphomas, and in elderly pts with MCL, B-R demonstrates a PFS and TTNT benefit over CHOP-R. Additionally, treatment with B-R resulted in a trend toward a survival benefit in the group of pts with indolent lymphomas.

Disclosure: Mathias Rummel: Financing of Scientific Research: Amgen, Celgene, Gilead, Janssen, Mundipharma, Roche; Expert Testimony: Grifols, Mundipharma, Roche. Wolfram Brugger: Advisory Role: Ja; Financing of Scientific Research: Mundipharma, Roche Pharma; Other Financial Relationships: Teilnahme an wissenschaftlichen Kongressen.

V419 - Bendamustine plus Rituximab versus Fludarabine plus Rituximab in patients with relapsed follicular, indolent, or mantle cell lymphomas - 8-year follow-up results of the randomized phase III study NHL 2-2003 on behalf of the StiL (Study Group Indolent Lymphomas, Germany)

Rummel M.1, Balser C.2, Kaiser U.3, Balló H.4, Stauch M.5, Heider A.6, Welslau M.7, Losem C.8, Weidmann E.9, Blau W.1, Burchardt A.1, Barth J.1, Kauff F.1, Brugger W.10, StiL (Studiengruppe indolente Lymphome)

1Universitätsklinik, Med Kl IV, Hämatologie, Gießen, Germany, 2Gemeinschaftspraxis, Marburg, Germany, 3St. Bernward Krankenhaus, Hildesheim, Germany, 4Gemeinschaftspraxis, Offenbach, Germany, 5Hämatologisch-Onkologische Schwerpunktpraxis, Kronach, Germany, 6Klinikum Leverkusen gGmbH, Leverkusen, Germany, 7Phase drei, Hämato-Onkologischer Studienkreis am Klinikum Aschaffenburg, Aschaffenburg, Germany, 8Facharztzentrum, Neuss, Germany, 9Krankenhaus Nordwest, Frankfurt/Main, Germany, 10Schwarzwald-Baar Klinikum, Villingen-Schwenningen, Germany

Introduction: No standard exists for patients (pts) with relapsed/refractory follicular lymphoma (FL), other indolent lymphoma (oiNHL), or mantle cell lymphoma (MCL). Treatment with fludarabine containing regiments were common. In 2003 we initiated a multicenter, randomized phase III study to compare the efficacy and safety of bendamustine plus rituximab (B-R) versus F-R for pts with relapsed FL, oiNHL or MCL to further improve the treatment.

Methods: 230 pts with relapsed FL, oiNHL, or MCL in need of treatment were randomized to rituximab 375 mg/m² (day 1) plus either bendamustine 90 mg/m² (days 1+2) or fludarabine 25 mg/m² (days 1-3) q 28 days for a maximum of 6 cycles. Prophylactic use of antibiotics or G-CSF was not generally recommended; however, in cases of severe granulocytopenia, G-CSF use was permitted. The primary endpoint was progression-free survival (PFS). Secondary endpoints were overall survival (OS), overall response rate (ORR), and complete response rate (CR).

Results: A total of 219 pts were evaluable for the analysis (114 B-R; 105 F-R). There were no significant differences between arms for patient characteristics. Most pts had stage IV (71.6% B-R; 60.6% F-R) or stage III disease (21.1% B-R; 25.3% F-R). Median pts age was 68 yrs (range 38-87). Pts had received a median of 1 prior therapy (range 1-7). Histological subtypes were distributed equally between the B-R and F-R arms. A median of 6 cycles were given in both treatment arms, with 75.2% and 53.4% of B-R and F-R pts receiving 6 cycles, respectively. At the time of this analysis, the median observation time was 96 months. The ORR was significantly higher with B-R than with F-R (83.5% vs. 52.5%, respectively; p < 0.0001). The CR rate with B-R was also significantly higher than that with F-R (38.5% vs. 16.2%; p = 0.0004). Median PFS was significantly prolonged with B-R compared with F-R (34 vs. 12 months p < 0.0001). The longer PFS translated into a survival benefit with a significantly longer median overall survival in the B-R group than in the F-R group (110 vs. 49 months; p = 0.0125) comprising 55 and 71 deaths in the B-R and F-R groups, respectively.

There were no significant differences in the rates of alopecia, stomatitis, erythema, allergic reactions, peripheral neuropathy, or infectious episodes between groups. Hematologic toxicities were also similar between arms. The overall incidence of serious adverse events was similar for the B-R and F-R groups (17.4% and 22.2%, respectively). 17 pts (14.9%) developed a secondary neoplasia after B-R compared with 16 pts (15.2%) after F-R. Of these, 5 pts in the B-R group, and 3 pts in the F-R group developed a secondary hematological neoplasia.

Conclusions: B-R was more effective than F-R in this setting of relapsed FL, other indolent lymphomas and MCL due to higher overall and complete response rates, a longer PFS, and an improved OS. These data confirm the high anti-lymphoma activity of B-R.

Disclosure: Mathias Rummel: Financing of Scientific Research: Amgen, Celgene, Gilead, Janssen, Mundipharma, Roche; Expert Testimony: Grifols, Mundipharma, Roche. Wolfram Brugger,: Advisory Role: Ja; Financing of Scientific Research: Mundipharma, Roche Pharma; Other Financial Relationships: Teilnahme an wissenschaftlichen Kongressen.

V420 - Treatment of refractory hairy cell leukemia by BRAF-inhibition

Dietrich S.1, Pircher A.2, Andrulis M.A.3, Peyrade F.4, Wendtner C.-M.5, Gaehler A.6, Follows G.7, Dyer M.8, Elter T.9, Zeiser R.10, Herrmann M.11, Herold M.11, Dearden C.12, Haferlach T.13, Hallek M.14, Hüllein J.15, Matutes E.12, Dürig J.16, von Kalle C.15, Glimm H.15, Fröhling S.15, Abdel-Wahab O.17, Hutter B.18, Steurer M.2, Ho A.3, Zenz T.15

1Universitätsklinik Heidelberg, Medizinsiche Klinik V, Heidelberg, Germany, 2Universität, Innsbruck, Austria, 3Universitätsklinik, Heidelberg, Germany, 4University, Nice, France, 5Krankenhaus Schwabing, München, Germany, 6Kantonspital, Luzern, Switzerland, 7University of Cambridge, Cambridge, United Kingdom, 8University of Leicester, Leicester, United Kingdom, 9Universitätsklinikum, Köln, Germany, 10Universitätsklinikum, Freiburg, Germany, 11Helios Klinikum, Erfurt, Germany, 12Royal Marsden Hospital, London, United Kingdom, 13MLL Münchner Leukämie Labor, München, Germany, 14Universität, Köln, Germany, 15NCT, Heidelberg, Germany, 16Universitätsklinikum, Essen, Germany, 17MSKCC, New York, United States, 18DKFZ, Heidelberg, Germany

Background: An activating mutation of the BRAF serine/threonine protein kinase (BRAF V600E) is the key genetic lesion in hairy cell leukemia patients (HCL), suggesting oncogene dependence and opportunities for therapeutic targeting.

Methods: We have collected and analyzed clinical course, molecular studies and follow up of 21 HCL patients treated with the BRAF inhibitor vemurafenib. In three of these patients we performed whole exome sequencing to better understand treatment refractory disease course.

Results: BRAFi treatment led to improved blood counts and clinical response in all patients. The overall response rate (ORR) was 95% with 8 CRs (38%) and 12 PRs (57%). A forth of the standard dose of vemurafenib was sufficient (240 mg bid, n = 12/21) to completely abrogate p-ERK signaling of hairy cells in the bone marrow, including patients with PR. Time to blood count improvement (e.g. median time to platelet > 100/nl 28 days) and decrease of sCD25 serum levels (80% within the first two weeks) was very fast and no difference between patients with low (≤240 mg bid) and higher vemurafenib doses was observed. Off drug relapses occurred in 11/21 patients including patients with CR or treated upfront with vemurafenib after a median of 17 months since treatment cessation. Nine patients were retreated and responded to a second course of BRAFi (n = 6) and monoclonal antibodies with and without purine analogues (n = 3).

To better understand disease biology and course of HCL treated with BRAFi, we performed whole exome sequencing in 3 patients. In addition to the disease defining BRAFV600E mutations, we identify EZH2, ARID1A mutations and recurrent inactivating mutations of the cell cycle inhibitor CDKN1B (p27). Targeted deep sequencing of CDKN1B in a larger cohort of HCL patients identify deleterious CDKN1B mutations in 16% of patients with HCL (n = 13/81). In 11 of 13 patients the CDKN1B mutation was clonal, implying an early role of CDKN1B mutations in the pathogenesis of HCL.

Conclusion: Our data provide clinical proof of the critical dependence on active BRAF signaling in HCL. We provide conclusive evidence that anti-tumor effect and side effects are observed at fractions of the standard vemurafenib dosing, but off drug relapses occur inevitably. Exome sequencing reveals that additional cooperating recurrent genetic events occur and may contribute to the disease biology of HCL.

Disclosure: No conflict of interest disclosed.

V421 - Advanced stage follicular lymphomas with and without breaks in the BCL2 locus do not differ with respect to clinical features and outcome

Leich E.1, Unterhalt M.2, Wartenberg M.3, Siebert R.4, Klapper W.5, Engelhard M.6, Stuhlmann-Laeisz C.5, Koch K.5, Puppe B.7, Horn H.8,9,10, Bernd H.-W.11, Feller A.C.11, Hummel M.12, Lenze D.12, Stein H.13, Hartmann S.14, Hansmann M.L.14, Möller P.15, Hiddemann W.2, Dreyling M.2, Hoster E.16, Ott G.17, Rosenwald A.3, German Low Grade Lymphoma Study Group

1Institute of Pathology,University of Würzburg, Würzburg, Germany, 2Department of Internal Medicine III, University Hospital, Munich, Germany, 3Institute of Pathology, University of Würzburg, Würzburg, Germany, 4Institute of Human Genetics, Christian-Albrechts-University Kiel and University Hospital Schleswig-Holstein, Kiel, Germany, 5Institute of Pathology, Hematopathology Section and Lymph Node Registry, University Hospital Schleswig-Holstein, Kiel, Germany, 6Department for Radiotherapy, University Hospital, Essen, Germany, 7Pathologisches Institut/Universität, Würzburg, Germany, 8Dr. Margarete Fischer-Bosch Institute of Clinical Pharmacology, Stuttgart, Germany, 9Robert Bosch Krankenhaus, Stuttgart, Germany, 10University, Tuebingen, Germany, 11Institute of Pathology, University Hospital Schleswig-Holstein, Lübeck, Germany, 12Institute of Pathology, Campus Benjamin Franklin, Charité Universitätsmedizin, Berlin, Germany, 13Pathodiagnostik, Berlin, Germany, 14Institute of Pathology, University Hospital, Frankfurt a.M., Germany, 15Institute of Pathology, University Hospital, Ulm, Germany, 16Institute of Medical Informatics, Biometry, and Epidemiology, University of Munich, Munich, Germany, 17Department of Clinical Pathology, Robert-Bosch-Krankenhaus, Stuttgart, Germany

Introduction: Approximately 15% of follicular lymphomas (FL) lack chromosomal breaks in BCL2. Clinical features of this subgroup remain largely undefined. The aim of this study, therefore, was to improve knowledge on molecular and clinical features of breakpoint negative FL.

Participants and Methods: We studied the presence or absence of chromosomal breaks in BCL2, BCL6 and MYC genes by fluorescence in situ hybridization in large clinical trial cohorts of FL stages III/IV (n = 540) and I/II (n = 117) patients enrolled in the German Low Grade Lymphoma Study Group trials, treated with chemotherapy regimens with or without rituximab and radiation, respectively. The protein expression status of BCL2, BCL6, MUM1, CD10 and P53 and the proliferation status of Ki67 was determined by immunohistochemistry. Finally, we compared the biological and clinical features of FL grades 1-3A with and without chromosomal breaks in BCL2.

Results: Chromosomal breaks in BCL2 were detected in 86% of FL stages III/IV and in 53% of FL stages I/II. BCL2 was expressed in almost all t(14;18)-positive FL and surprisingly also in 86% and 69% of t(14;18)-negative FL stages III/IV and I/II, respectively. CD10 expression was significantly reduced in BCL2-break negative FL of all stages. However, there was high similarity between FL with and without BCL2-break with respect to the other molecular features and the clinical parameters investigated. Most importantly, median overall survival and time to treatment failure did not differ significantly between stage III/IV FL with and without breaks in BCL2. Likewise, no significant difference for FLIPI score and performance-status was observed.

Conclusion: The results of this large retrospective study suggest that FL with and without breaks in BCL2 show a high degree of similarity with regard to most molecular markers investigated and in particular with regard to their clinical behaviour. This supports the notion that FL with and without breaks in BCL2 represent two subgroups within the same entity.

Disclosure: No conflict of interest disclosed.

V422 - Gastric marginal zone B-cell lymphoma of the mucosa-associated lymphoid tissue (MALT lymphoma) in the course of time: The Vienna University experience

Kiesewetter B.1, Dolak W.2, Wöhrer S.3, Mayerhoefer M.E.4, Simonitsch-Klupp I.5, Chott A.6, Jäger U.7, Raderer M.1

1Medizinische Universität Wien / Universitätsklinik für Innere Medizin I, Onkologie, Wien, Austria, 2Medizinische Universität Wien / Universitätsklinik für Innere Medizin III, Gastroenterologie und Hepatologie, Wien, Austria, 3Medizinische Universität Wien / Universitätsklinik für Innere Medizin I, Knochenmarktransplantation, Wien, Austria, 4Medizinische Universität Wien, Radiologie, Wien, Austria, 5Medizinische Universität Wien, Pathologie, Wien, Austria, 6Wilhelminenspital, Pathologie und Mikrobiologie, Wien, Austria, 7Medizinische Universität Wien / Universitätsklinik für Innnere Medizin I, Hämatologie und Hämostaseologie, Wien, Austria

Introduction: Gastric MALT lymphoma, most prominent for its association with Helicobacter pylori (HP), has been well characterized in the last three decades. However, changes in clinicopathological features and incidence have recently been suggested by scattered reports.

Methods: We report our experience of 320 consecutive MALT lymphoma patients diagnosed and followed at our institution 1999-2015. Patients with gastric lymphoma were reassessed for clinicopathological characteristics including treatment and outcome.

Results: 116/320 patients (36%) were of gastric origin, while the majority of patients had extragastric MALT lymphoma. Only 4/116 patients (3%) showed an additional diffuse-large B-cell lymphoma component. Using a standardized staging protocol we found 25 patients (22%) to have disseminated disease, with only 3 (3%) having bone marrow involvement, while 78% had localized disease. Out of 99 patients with a fully tested HP-status, 34 (34%) were negative. Interestingly, 9 of them had been diagnosed before 2004 (9/34, 26%) and 25 between 2004-2015 (74%). 40% of patients had monoclonal gammopathy and plasmacytic differentiation was present in 31%. In addition, 29% were found to have an underlying autoimmune disease, with chronic thyroiditis being most common. In terms of treatment, 76 patients (66%) had antibiotics as first line therapy, 31 systemic therapy (27%), while the remaining had surgery, radiation or wait and see. After an impressive median FUP of 63m, 85% (99/116) are alive, while 17 (15%) have died. HP-status was not associated with relapse after first line therapy (p = 0.07), and there was no difference in time to progression between the cohorts. 14 HP-negative patients were treated with antibiotics alone, resulting in 5 CR and one PR (46%), 5 SD and 3 progressions. Results of antibiotic treatment in HP negative patients were not significantly different for time to progression (p = 0.1) and progression following first line (p = 0.38). Transformation to DLBCL was only seen in one patient while two developed clonally unrelated DLBCL.

Conclusion: Our findings suggest, that the rate of extragastric MALT lymphoma appears to be higher than stated in the literature and might have been underestimated. In addition, cautious interpretation of our data proposes an increase of HP-negative gastric lymphoma over the years and that also this collective can be managed safely with antibiotics. Our data also show that the risk of DLBCL transformation is minimal.

Disclosure: No conflict of interest disclosed.

V423 - Treatment of low grade B-cell-lymphoma cell lines with arylindolylmaleimide derivates PDA-66 and PDA-377 induces antiproliferative effects

Eschenburg A.1, Roolf C.1, van der Wall K.1, Sklarz L.-M.1, Jaenecke C.1, Sekora A.1, Murua Escobar H.1, Rolfs A.2,3, Pews-Davtyan A.4, Beller M.4, Junghanss C.1

1Universitätsmedizin Rostock, Klinik III, Hämatologie, Onkologie, Palliativmedizin, Rostock, Germany, 2Universitätsmedizin Rostock, Albrecht-Kossel-Institute for Neuroregeneration, Rostock, Germany, 3Centogene AG, Rostock, Germany, 4University of Rostock, Leibniz-Institute for Catalysis, Rostock, Germany

Introduction: B-cell lymphomas are the most common type of non-Hodgkin lymphomas (NHL). Curative options for indolent lymphoma patients diagnosed in advanced stages are currently limited. The novel arylindolylmaleimide PDA-66 has been shown to inhibit the polymerisation of microtubules inducing antiproliferative effects in acute lymphoblastic leukemia cells. Here, we evaluated the effects and potency of PDA-66 and its derivate PDA-377 on B-cell NHL cell lines.

Methods: NHL cell lines (SC-1, MINO and U-266) were treated with increasing concentrations of PDA-66 and PDA-377 (0.5 µM-5 µM) and incubated for 72 h. Metabolic activity was determined by WST-1 assay. Furthermore, gene expression changes were investigated after 2 and 24 h treatment with PDA-66 using human signal transduction pathway finder qPCR assay. Further, 1 µM PDA-66 was tested in combination with 25 nM cytarabine (AraC), 50 nM doxorubicin (Doxo) or 10 nM dexamethasone (Dexa) on SC-1 cells, all in IC-80 concentrations. The applied cytostatics were added either at the time of cell seeding simultaneously with PDA-66, or 24 h before, or after PDA-66 treatment, respectively.

Results: Metabolic activity was significantly reduced in a dose dependent manner in all cell lines starting at a concentration of 1 µM PDA-66 and 2.5 µM PDA-377. Treatment with 5 µM PDA-66 decreased metabolic activity to 34.4% (SC-1), 7.9% (MINO) and 29.7% (U-266) compared to control cells treated with DMSO (100%). Incubation with 5 µM PDA-377 reduced metabolic activity to 39.4% (SC-1), 51.1% (MINO) and 40.5% (U-266) respectively. Gene expression analysis showed that PDA-66 influenced NFkB pathway by down regulation of TNF gene. Key genes of other investigated pathways like Wnt, JAK/STAT, p53 and NOTCH were not affected.

Treatment with PDA-66 demonstrates synergistic effects on proliferation inhibition when combined with AraC, Doxo or Dexa using Bliss statistics. The observed combined effect was independent from PDA-66/cytostatic application showing no enhanced impact on the metabolic activity.

Conclusions: Our data indicates that application of PDA-66 alone or in combination with cytostatic drugs shows distinct effects on B-NHL cells impacting the NFkB signaling pathway.

Disclosure: No conflict of interest disclosed.

Freier Vortrag

AML klinisch

V424 - Prolonged survival in patients ≥60 years with first relapsed/refractory acute myeloid leukemia treated with vosaroxin plus cytarabine vs placebo plus cytarabine: Results from the phase 3 VALOR study

Krug U.1, Ravandi F.2, Ritchie E.K.3, Sayar H.4, Lancet J.E.5, Craig M.D.6, Vey N.7, Strickland S.A.8, Schiller G.J.9, Jabbour E.2, Erba H.P.10, Pigneux A.11, Horst H.-A.12, Recher C.13, Klimek V.M.14, Cortes J.2, Roboz G.J.3, Craig A.R.15, Fox J.A.15, Ward R.15, Smith J.A.15, Acton G.15, Mehta C.16, Kantarjian H.M.2, Stuart R.K.17

1Universitätsklinikum Münster, Leverkusen, Germany, 2University of Texas MD Anderson Cancer Center, Houston, United States, 3Weill Cornell Medical Center, New York, United States, 4Indiana University Cancer Center, Indianapolis, United States, 5Moffitt Cancer Center, Tampa, United States, 6West Virginia University, Morgantown, United States, 7Institut Paoli-Calmettes and Aix-Marseille University, Marseille, France, 8Vanderbilt-Ingram Cancer Center, Nashville, United States, 9University of California, Los Angeles, United States, 10University of Alabama, Birmingham, United States, 11Université de Bordeaux, CHU de Bordeaux, Bordeaux, France, 12II. Medizinische Klinik und Poliklinik im Städtischen Krankenhaus, Kiel, Germany, 13Institut Universitaire du Cancer de Toulouse Oncopole, Université de Toulouse III, CHU de Toulouse, Toulouse, France, 14Memorial Sloan-Kettering Cancer Center, New York, United States, 15Sunesis Pharmaceuticals, Inc., South San Francisco, United States, 16Cytel, Inc. and Harvard School of Public Health, Cambridge, United States, 17Medical University of South Carolina, Charleston, United States

Introduction: Prognosis for older patients with relapsed or refractory (R/R) acute myeloid leukemia (AML) is poor and treatment options are limited. Vosaroxin is a first-in-class anticancer quinolone derivative active in AML. VALOR, a phase 3, randomized, double-blind, placebo-controlled trial, evaluated vosaroxin plus cytarabine (vos/cyt) vs placebo plus cytarabine (pla/cyt) in patients with R/R AML (NCT01191801).

Methods: Patients with refractory disease after primary induction treatment or in first relapse were randomized 1:1 to receive cytarabine (1 g/m2 IV over 2 h, d 1-5) plus either vosaroxin (90 mg/m2 IV over 10 min d 1 and 4; 70 mg/m2 in subsequent cycles) or placebo. Primary endpoints: overall survival (OS) and 30- and 60-day mortality. Here, we report results of a predefined subgroup analysis in patients ≥60 y of age.

Results: Of 711 patients randomized, 63% (n = 451) were ≥60 y (n = 226 randomized to vos/cyt and n = 225 to pla/cyt). At final analysis, OS was improved with vos/cyt (7.1 mo vs 5.0 mo with pla/cyt; HR = 0.75; P = 0.003) (Figure). Complete remission (CR) was achieved in 31.9% of patients treated with vos/cyt vs 13.8% with pla/cyt (P< 0.0001). Responses were durable; median leukemia-free survival in patients who achieved CR was 10.3 mo with vos/cyt vs 6.5 mo with pla/cyt (P = 0.20).

Thirty- and 60-day all-cause mortality was similar between treatment arms (30-day: 10.2% vs 9.0%; 60-day: 20.4% vs 22.6% with vos/cyt vs pla/cyt, respectively). Serious AEs were more common with vos/cyt (57% vs 33% with pla/cyt); most common serious AEs were febrile neutropenia (9.3% with vos/cyt vs 5.9% with pla/cyt), sepsis (9.3% vs 5.0%), pneumonia (7.5% vs 4.5%), bacteremia (7.5% vs 2.3%), and stomatitis (4.4% vs 1.8%). Serious and non-serious cardiac, renal, neurologic, and hepatic AEs were comparable between treatment groups.

Conclusions: Addition of vosaroxin to cytarabine improved OS and increased CR rates without increasing early mortality in patients ≥60 y with R/R AML. The toxicity observed in the vosaroxin arm was acceptable in light of the benefit received. VALOR results support the use of vos/cyt as a potential standard of care in patients ≥60 y with R/R AML.


Disclosure: Utz Krug: Advisory Role: Sunesis Pharmaceuticals, Roche, Gilead Sciences, Novartis, Catapult Cell Therapeutics, Celgene, Clavis Pharma; Expert Testimony: Boehringer Ingelheim; Other Financial Relationships: Travel support: Mundipharma, Novartis; data monitoring board fees: German-Austrian AML Study Group. Robert K. Stuart: Advisory Role: Sunesis Pharmaceuticals; Expert Testimony: Sunesis Pharmaceuticals; Other Financial Relationships: Travel support: Sunesis Pharmaceuticals.

V425 - Clinical outcome in older AML patients not fit for intensive chemotherapy: Results of the population-based AMLSG-BiO (NCT01252485) registry study of the German-Austrian AML study group

Schlenk R.F.1, Paschka P.2, Krauter J.3, Lübbert M.4, Brossart P.5, Salih H.R.6, Theobald M.7, Fiedler W.8, Wulf G.9, Kirchner H.10, Hertenstein B.11, Heidel F.12, Westermann J.13, Grießhammer M.14, Wattad M.15, Götze K.16, Girschikofsky M.17, Kündgen A.18, Koller E.19, Rudolph C.2, Heuser M.20, Thol F.20, Goehring G.20, Teleanu V.2, Weber D.2, Gaidzik V.I.2, Döhner K.2, Ganser A.20, Döhner H.2, German-Austrian Acute Myeloid Leukemia Study Group (AMLSG)

1Uniklinik Ulm, Klinik für Innere Medizin III, Ulm, Germany, 2Uniklinik, Ulm, Germany, 3Klinikum, Braunschweig, Germany, 4Uniklinik, Freiburg, Germany, 5Uniklinik, Bonn, Germany, 6Uniklinik, Tübingen, Germany, 7Uniklinik, Mainz, Germany, 8Uniklinik, Hamburg, Germany, 9Uniklinik, Göttingen, Germany, 10Klinikum Frankfurt-Höchst, Frankfurt, Germany, 11Klinikum Bremen Mitte, Bremen, Germany, 12Uniklinik, Magdeburg, Germany, 13Charité - Universitätsmedizin, Berlin, Germany, 14Klinikum, Minden, Germany, 15Klinikum, Essen, Germany, 16Uniklinik München r.d.I, München, Germany, 17Krankenhaus der Elisabethinen, Linz, Austria, 18Uniklinik, Düsseldorf, Germany, 19Hanuschkrankenhaus, Wien, Austria, 20Medizinische Hochschule Hannover, Hannover, Germany

Background: Comparative population-based outcome data in older AML patients (pts) unfit for intensive chemotherapy (IC) are rare outside interventional treatment trials and treatment options include best supportive care (BSC), decitabine (DAC), azacitidine (AZA), or low-dose cytarabine (LDAC).

Aims: To assess outcome in pts >60 years (yrs) unfit for IC within the registry/molecular-screening trial AMLSG BiO (NCT01252485).

Methods: A total of 2786 pts (median age 72 yrs, range 60-95) were registered between October 2010 and May 2015. All pts were screened within 48-hours for gene mutations and fusion-genes.

Results: Of the 2786 pts, n = 1338 (48%) received IC, n = 107 (4%) AZA, n = 222 (8%) DAC, n = 281 (10%) LDAC, n = 276 (10%) BSC, and n = 562 (20%) unknown treatment. Of 886 non-intensively treated pts, n = 201 (23%) were treated within clinical trials. In the remaining 685 pts treatment was as follows: n = 144 LDAC, n = 162 DAC, n = 106 AZA, and n = 273 BSC. There was no difference between the groups in terms of age (p = 0.18, median 76 yrs, range 60-92) and cytogenetic risk groups according to ELN recommendations (p = 0.12; high, 27%; intermediate/low, 73%). Significantly more pts with HCT-CI score >2 were present in the LDAC group (p = 0.03) and with ECOG performance status >1 (p < 0.001) in the BSC group. NPM1 mutations were more frequent (p < 0.001) in BSC and LDAC (22%, each) groups than in AZA (5%) or DAC (7%) group. FLT3-ITD and FLT3-TKD were rarely detected (2.5%, 2.7%, respectively) with no difference between the groups (p = 0.69, p = 0.99). WBC at diagnosis was significantly (p < 0.001) higher in BSC and LDAC (26G/l; each) groups compared to AZA (4.5G/l) or DAC (5.8G/l) groups. According to the 4 groups (BSC, AZA, DAC, LDAC), median survival was 0.9, 10.5, 8.6, and 3.6 months, while one-year and two-year survival rates were 15%, 46%, 39%, 30% and 4%, 15%, 6%, 16%, respectively. In a Cox regression model including the groups AZA, DAC and LDAC, survival was significantly influenced by age [hazard ratio (HR) by 10 yrs difference, 1.46; p = 0.03], high-risk cytogenetics (HR, 1.78; p = 0.003), treatment (reference LDAC) with AZA (HR, 0.39; p < 0.001), DAC (HR, 0.43; p < 0.001), ECOG > 1 (HR, 1.55; p = 0.03) but not by NPM1 (p = 0.70) and FLT3-ITD (p = 0.42) mutational status, and HCT-CI > 2 (p = 0.16).

Conclusion: In older AML treatment with AZA or DAC is in the short-term perspective of one to two years superior to LDAC; results beyond two yrs are still dismal for all treatment options.

Disclosure: Richard Schlenk: Advisory Role: IQWIG, Ambit; Financing of Scientific Research: Pfizer, Novartis, Janssen, Celgene, Amgen, Teva; Expert Testimony: Pfizer, Novartis, Janssen, Celgene, Amgen, Teva, Arog. Hartmut Döhner: Advisory Role: Celgene; Financing of Scientific Research: Celgene.

V426 - DNMT3A mutations in exon 23 are associated with improved survival in AML patients treated with Azacytidine plus standard chemotherapy

Tschanter P.1, Krug U.2, Füller M.3, Klein M.3, Göllner S.1, Rohde C.1, Röllig C.4, Thiede C.4, Lilly M.A.4, Haack B.4, Koschmieder A.3, Stelljes M.3, Dugas M.3, Koschmieder S.5, Gerss J.3, Butterfaß-Bahloul T.3, Wagner R.3, Eveslage M.3, Thiem U.6, Krause S.W.7, Kaiser U.8, Kunzmann V.9, Steffen B.10, Noppeney R.11, Herr W.12, Baldus C.D.13, Schmitz N.14, Götze K.15, Reichle A.16, Kaufmann M.17, Neubauer A.18, Schäfer-Eckart K.19, Hänel M.20, Peceny R.21, Frickhofen N.22, Kiehl M.23, Giagounidis A.24, Görner M.25, Repp R.26, Link H.27, Kiani A.28, Naumann R.29, Brümmendorf T.H.5, Serve H.10, Ehninger G.4, Berdel W.E.3, Müller-Tidow C.1, Studien-Allianz Leukämie (SAL)

1Universitätsklinikum, Halle (S.), Germany, 2Klinikum Leverkusen gGmbH, Leverkusen, Germany, 3Uniklinik, Münster, Germany, 4Uniklinik, Dresden, Germany, 5Uniklinik, Aachen, Germany, 6Universität, Bochum, Germany, 7Uniklinikum, Erlangen, Germany, 8St. Bernward Hospital, Hildesheim, Germany, 9Uniklinik, Würzburg, Germany, 10Uniklinik, Frankfurt, Germany, 11Uniklinik, Essen, Germany, 12Universitätsklinikum, Mainz, Germany, 13Charité Campus Benjamin Franklin, Berlin, Germany, 14ASKLEPIOS Klinik St. Georg, Hamburg, Germany, 15Klinikum rechts der Isar (Technische Universität München), München, Germany, 16Universitätsklinikum, Regensburg, Germany, 17Robert-Bosch-Krankenhaus, Stuttgart, Germany, 18Uniklinikum, Marburg, Germany, 19Klinikum Nuernberg Nord, Nürnberg, Germany, 20Klinikum Chemnitz gGmbH, Chemnitz, Germany, 21Klinikum, Osnabrück, Germany, 22Dr.-Horst-Schmidt-Klinik, Wiesbaden, Germany, 23Klinikum, Frankfurt/Oder, Germany, 24St. Johannes Hospital, Duisburg, Germany, 25Städtische Kliniken, Bielefeld, Germany, 26Klinikum am Bruderwald, Bamberg, Germany, 27Westpfalz-Klinikum, Kaiserslautern, Germany, 28Klinikum, Bayreuth, Germany, 29Stiftungsklinikum Mittelrhein, Koblenz, Germany

DNMT3A mutations are among the most common mutations in AML with about half of all mutations found in exon 23 in one hotspot, codon R882. DNMT3A-R882 mutations seem to be associated with specific changes in DNA methylation patterns and worse outcome. A few reports suggest that monotherapy with DNMT inhibitors might be specifically active in AML with DNMT3A mutations. Here, we analyzed the association between DNMT3A mutations and chemotherapy response with or without added azacytidine as DNMT3A inhibitor.

The AML-AZA trial (n = 214 pts) tested whether azacytidine (Aza) in combination with intensive chemotherapy could improve outcome in AML (arm A) compared to patients receiving chemotherapy only (arm B). In the whole study population, event free survival (EFS) and overall survival (OS) did not differ significantly between the two groups. DNMT3A mutations were found in 56 of 167 (34%) patients analysed, predominantly in exon 23 (35/167; 21%). Patients with DNMT3A mutation in exon 23 treated with Aza showed trends to longer EFS (median 8.7 versus 5.6 months, p = 0.069) and OS (median not reached versus 13 months, p = 0.28). Cox-regression analysis was performed to evaluate the interaction between DNMT3A exon 23 mutations and treatment. Presence of DNMT3A mutation was associated with trends for prolonged EFS in the AZA arm (HR = 0.64; 95% CI 0.31-1.31, p = 0.221) but not in the control arm. As a result, the hazard ratio (HR) of Aza versus no Aza was decreased to a factor 0.37 when comparing patients with present versus absent DNMT3A mutation exon 23 (ratio of Hazard Ratios = 0.50/1.36 = 0.37; 95%CI 0.15-0.91, p = 0.03). To complement these findings with mechanistic data, U937 cells were transduced with either DNMT3A-wildtype (WT) or DNMT3A-R882H (MT). U937-MT cells showed reduced growth and colony formation in the presence of azacytidine whereas U937-WT cells were resistant. Very low concentrations of azacytidine in combination with cytarabine were effective in inhibiting colony formation in U937-MT cells. U937-WT cells were less affected.

Taken together, these data suggest that DNMT3A-R882 mutations are especially susceptible to azacytdine in combination with chemotherapy. A confirmatory clinical trial is required to validate these findings.

Disclosure: Petra Tschanter: Financing of Scientific Research: Vortragshonorar von Celgene. Carsten Müller-Tidow: Financing of Scientific Research: Vortragshonorar von Celgene; Expert Testimony: von Celgene und Amgen.

V427 - Association of single nucleotide polymorphisms of innate immunity and invasive fungal disease in patients with acute myeloid leukemia

Schnetzke U.1, Fischer M.1, Spies-Weißhart B.1, Schrenk K.1, Gruh B.2, Wittig S.2, Glaser A.3, Hochhaus A.1, Scholl S.1

1Universitätsklinikum Jena, Klinik für Innere Medizin II, Jena, Germany, 2Universitätsklinikum Jena, Klinik für Kinder- und Jugendmedizin, Jena, Germany, 3Universitätsklinikum Jena, Institut für Humangenetik und Anthropologie, Jena, Germany

Introduction: Induction chemotherapy of acute myeloid leukemia (AML) is associated with a high risk of invasive fungal disease (IFD). Several pattern recognition receptors (PRR) of the innate immune system are highly important for the response to fungal infections. A series of functionally relevant single nucleotide polymorphism (SNPs) of such PPRs represent promising candidates to predict the individual risk of IFD for intensively treated AML patients. We sought to investigate the impact of different SNPs of innate immunity on the occurrence of IFD during AML induction chemotherapy.

Methods: Genotyping of TLR1 (rs5743611, rs5743618, rs4833095), TLR2 (rs5743708), Dectin-1 (rs7309123, rs3901533, rs16910526) and DCsign (rs4804800, rs11465384, rs7248637) was performed by TaqMan method and/ or pyrosequencing in 186 adult AML patients. IFD was classified according to the EORTC/MSG consensus guidelines. Multiple logistic regression analysis was applied to evaluate the association between distinct SNPs or defined SNP clusters and the occurrence of IFD.

Results: Patients carrying either two homozygous TLR1 SNPs (rs5743618, rs4833095), the TLR2 polymorphism, the Dectin-1 SNP rs7309123 or polymorphisms of all analyzed DCsign loci have a significant higher risk for developing pulmonary IFD. Importantly, cluster analyses defined as the detection of polymorphisms in at least two subsets of these SNPs (TLR1, TLR2, Dectin-1 and/ or DCsign) demonstrates positivity in only 6 of 138 AML (4.3%) patients without IFD compared to 14 of 48 (29.2%) patients who fulfilled the criteria of IFD according to the EORTC/MSG guidelines (OR 9.06 (95% CI 3.24-25.32), p < 0.001).

Conclusions: Defined SNP clusters of innate immunity may predict an increased risk of IFD in AML patients undergoing induction chemotherapy.

Disclosure: Ulf Schnetzke: Expert Testimony: Fa. Gilead Förderprogramm Infektiologie 2013. Sebastian Scholl: Expert Testimony: Fa. Gilead Förderprogramm Infektiologie 2013.

V428 - Non-myeloablative conditioning (NMA) followed by hematopoietic stem cell transplantation (HCT) in patients (PTS) with acute myeloid leukemia (AML): Prognostic impact of the European LeukemiaNet (ELN) standardized reporting system

Bill M.1, Jentzsch M.1, Schubert K.1, Knyrim M.1, Grimm J.1, Cross M.1, Vucinic V.1, Franke G.N.1, Pönisch W.1, Behre G.1, Lange T.1,2, Niederwieser D.1, Schwind S.1

1Universität Leipzig, Hämatologie und Internistische Onkologie, Leipzig, Germany, 2Asklepios Klinik Weißenfels, Hämatologie und Internistische Onkologie, Weißenfels, Germany

Introduction: Introduction of NMA rendered older or unfit pts eligible for HCT. The prognostic impact of the ELN system for these pts is unknown.

Patients and methods: We analyzed 215 pts, treated at the University Hospital Leipzig between April 1999 & July 2013 (median age 64 years [y; range 37-76y]). We included de novo (n = 123; 57.2%), secondary (n = 72; 33.5%) or therapy-related (n = 19; 8,8%) AML. Pts received: 2Gy total body irradiation either with fludarabine (30 mg/m² at day -4 to -2; n = 211; 98.1%) or without (n = 4; 1.9%). Donors were HLA-matched related (n = 35; 16.3%), HLA-matched (n = 119; 55.3%) or mismatched (>=1 antigen; n = 61; 28.3%) unrelated. Cytogenetics, presence of FLT3-ITD & NPM1 & CEBPA mutation (mut) status at diagnosis were assessed centrally.

Results: For all pts the median overall survival (OS) after transplantation was 1.98y, with a 3y-OS of 45.1%. Pts were classified according to the ELN system: 20.5% favorable (fav; n = 44; including 18.2% [n = 8] with core-binding factor AML, 34.1% [n = 15] CEBPA mut & 53.3% [n = 23] NPM1 mut/no FLT3-ITD with NK), 24.2% intermediate-I (int-I; n = 52; including 17.3% [n = 9] NPM1 mut/FLT3-ITD, 13.5% [n = 7], NPM1 wild type [wt]/FLT3-ITD, 67.3% [n = 35] NPM1 wt/no FLT3-ITD with NK), 23.1% intermediate-II (int-II; n = 50; including 2.0% [n = 1] with t(9;11)) & 32.1% adverse (adv; n = 69; including 71.0% [n = 49] with complex karyotype; 31.9% [n = 22] with -7; 40.6% [n = 28] with -5/del5q; 2.9% [n = 2] with inv(3)) genetic risk. We analyzed the cumulative incidence of relapse (CIR) according to ELN groups & found a significant separation (p = 0.033; Figure 1A). This was also found by trend for OS (p = 0.088; Figure1B). Analyzing OS among each group, only fav was performing significantly better than int-II (p = 0.043) or adv (p = 0.010).

Conclusion: We analyzed the prognostic impact of the ELN system for AML pts receiving NMA-HCT. Several studies focused on younger (≤60y), chemotherapy-based consolidated pts, showed that fav performed best & adv worst. Int-II had better outcome than int-I. In our set only fav pts performed better than int-II or adv pts. Thus, especially older or unfit pts within the int-I or adv ELN group might benefit from NMA-HCT.


Disclosure: No conflict of interest disclosed.

V429 - Minimal residual disease-guided preemptive treatment in patients with AML and MDS - a retrospective survey within the SAL study group

Bulycheva E.1, Mayer J.2, Krüger W.3, Mies A.1,4, Šustkova Z.2, Račil Z.2, Prochazkova J.2, Hirt C.3, Ringhoffer M.5, Berdel W.6, Serve H.7, Röllig C.1, Sockel K.1, Bornhäuser M.1, Ehninger G.1, Thiede C.1,4, Platzbecker U.1,4

1Universitätsklinikum Carl Gustav Carus, Technische Universität Dresden, Medical Department I, Dresden, Germany, 2University Hospital Brno, Department of Internal Medicine - Hematology and Oncology, Brno, Czech Republic, 3Ernst-Moritz-Arndt-Universität Greifswald, Department of Internal Medicine C, Hematology and Oncology, Marrow Transplantation, Greifswald, Germany, 4German Cancer Consortium, German Cancer Research Center, Heidelberg, Germany, 5Städtisches Klinikum Karlsruhe, Department of Internal Medicine III, Karlsruhe, Germany, 6Universitätsklinikum Münster, Department of Medicine A - Hematology, Oncology and Pneumology, Münster, Germany, 7J. W. Goethe-Universität Frankfurt, Department of Medicine II, Hematology/Oncology, Frankfurt, Germany

Introduction: Detection of minimal residual disease (MRD) in patients with acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS) in complete hematological remission (CR) offers a potential therapeutic window allowing early interventions in order to prevent overt hematological relapse. Since prospective clinical studies are limited we initiated a retrospective survey within the SAL study group and members of the DKTK Consortium to document the current clinical use of the approach and clinical outcomes.

Patients: We analyzed 74 patients (pts) at four centers (m/f = 42/32; median age 54 yrs (19-74)) with either MDS (n = 7) or AML (n = 67) in CR after either conventional intensive chemotherapy (CTx; n = 18) or allogeneic hematopoietic stem cell transplantation (allo-HSCT; n = 56), undergoing preemptive therapy. Markers for MRD monitoring were mutations, cytogenetic aberrations or donor chimerism for transplanted pts. Twenty-eight pts had a normal karyotype; 13 pts - complex; 6 pts carried -7 and/or inv(3); other aberrations were detected in 25 pts. Among molecular aberrations, the most prominent was NPM1 (n = 17) and FLT3-ITD (n = 10), followed by CBFβ/MYH11 (n = 7), RUNX1/RUNX1T1 (n = 7) and MLL/AF6 (n = 3).

Results: The median time to MRD positivity after completion of intensive therapy was 7.5 months (range, 1-97). Subsequent MRD-guided treatment in pts treated only with CTx included additional intensive CTx (13%), hypomethylating agents (HMA, 23%), clofarabine (23%), targeted therapy (gemtuzumab ozogamycin, 10%), low-dose cytarabine (13%), and allo-HSCT (13%). In transplanted patients the most preferred treatment for the molecular relapse was donor lymphocyte infusion (23 pts, 41%; median number of DLI = 2, range, 1-6), alone or in combination (60% with HMA). 26 MRD-based treated pts (38%) did not experience a hematological relapse and were alive (17 pts, median observation time 959 days, range, 297-3647), or died due to another cause (9 pts) with median survival of 359 days (range, 125-954). In 8 pts without hematological relapse more than one MRD recurrence was observed. In the remaining 62% relapsing patients, median time to hematological relapse was 267 days (range, 24-1161) after MRD positivity.

Conclusions: The retrospective analysis demonstrates that MRD-guided therapies have already entered routine practice in patients with MDS/AML. Therefore, clinical trials are needed to better define molecular markers and subsets of patients who might benefit from this approach.

Disclosure: Ekaterina Bulycheva: Other Financial Relationships: Gilead. Uwe Platzbecker: Financing of Scientific Research: Celgene, Novartis.

Freier Vortrag

Mammakarzinom I

V430 - An AKT/SOX2 molecular axis drives breast carcinoma clonogenicity

Schaefer T.1, Wang H.1,2, Mir P.2, Pereboom T.1, Merz B.3, Fehm T.4, Perner S.5, Rothfuss O.3, Kanz L.2, Schulze-Osthoff K.3,6, Lengerke C.1,2,7

1Department of Biomedicine, University Hospital, Basel, Switzerland, 2Department of Internal Medicine II, University Hospital, Tuebingen, Germany, 3Interfaculty Institute of Biochemistry, University of Tuebingen, Tuebingen, Germany, 4Women's Hospital, University Hospital, Duesseldorf, Germany, 5Section for Prostate Cancer Research, Institute of Pathology, Center for Integrated Oncology Cologne/Bonn, University Hospital, Bonn, Germany, 6German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ), Heidelberg, Germany, 7Clinic for Hematology, University Hospital, Basel, Switzerland

Introduction: The embryonic transcription factor SOX2 (Sex determining region Y - Box2) plays key roles in embryonic stem cell self-renewal. More recently SOX2 protein was detected in various types of cancer including breast carcinoma (BC). The molecular regulation of SOX2 in BC remains largely unknown. Here, we describe a functional dependence of SOX2 on the Ser/Thr-kinase AKT, a central mediator of cell growth, proliferation and metabolism, whose deregulation is a common determinant in BC.

Methods: Eight human BC cell lines and 10 patient samples were analyzed for SOX2, AKT and pAKT expression by RT-PCR, immunoblot, and IHC. Cells were cultured under conventional or 3D-conditions enriching for putative cancer stem cells (CSCs) and treated with cisplatin, paclitaxel or respectively MK-2206, AKT1/2 and AKT siRNAs to inhibit AKT activity. SOX2 expression was modulated with lentiviruses containing shSOX2 or an inducible mCherry-SOX2-overexpression construct. A previously described lentivirus containing a DsRed-SOX2 regulatory region 1 reporter element was used to isolate SOX2-positive breast CSCs. Tumor sphere assays were applied to investigate clonogenicity. pAKT and SOX2 subcellular localization was followed by confocal and life-time microscopy.

Results: Both pAKT and SOX2 were induced by culture conditions promoting CSCs and functionally regulated BC cell clonogenicity in tumor sphere assays. Unlike conventional chemotherapies, which promoted the outgrowth of SOX2-positive putative CSCs, AKT-inhibition reduced cell growth without enhancing the frequency of SOX2-reporter positive cells in the surviving fraction. Mechanistically, AKT activity modulated SOX2 subcellular distribution promoting non-canonical localization in the cytosol and subsequent proteasomal degradation. Immunoprecipitation showed direct physical interaction of AKT and SOX2 proteins and permutation of the T116 AKT phosphorylation site fostered cytoplasmic retention of SOX2. Functionally, SOX2 overexpression rescued tumor sphere formation in anti-AKT treated BC cells, indicating SOX2 as a major molecular target mediating AKT-effects on clonogenicity.

Conclusions: We here describe a hitherto unrecognized coupling of AKT and SOX2 proteins in human BC that has utmost significance for clonogencity and therapy resistance. Our investigations reveal important mechanistic details about the promises of AKT-inhibitor therapies that are currently under laboratory and clinical investigation in BC.

Disclosure: No conflict of interest disclosed.

V431 - Mcl-1 confers protection of HER2+ breast cancer cells exposed to hypoxia: Therapeutic implications

Bashari M.H.1, Fan F.1, Vallet S.1, Arn M.2, Cardone M.H.2, Beckhove P.3, Schneeweiss A.1, Sattler M.4, Opferman J.T.5, Jäger D.1,6, Podar K.1

1National Center for Tumor Diseases (NCT), University of Heidelberg, Medical Oncology, Heidelberg, Germany, 2Eutropics Pharmaceuticals, Inc, Cambridge, United States, 3National Center for Tumor Diseases (NCT), University of Heidelberg and German Cancer Research Center (DKFZ), Heidelberg, Germany, 4Dana-Farber Cancer Institute, Harvard Medical School, Boston, United States, 5St. Jude Children's Research Hospital, Memphis, United States, 6German Cancer Research Center (DKFZ), Applied Tumor-Immunity, Heidelberg, Germany

Introduction: Breast cancer (BC) is the most common malignancy and the leading cause of cancer-related mortality in women. HER2 overexpression occurs in 15-20% of BC patients and is associated with poor clinical outcome. Hypoxic conditions develop during cancer progression. Molecular mechanisms, which lead to the adaptation of HER2+ BC cells to hypoxia are unknown. The anti-apoptotic Bcl-2 family member Mcl-1 is frequently amplified in BC; and elevated Mcl-1 levels correlate with poor prognosis. The aim of this study is to evaluate whether Mcl-1 plays a role in the adaptation of HER2+ BC cells to hypoxic conditions and to provide the derived rationale to therapeutically target Mcl-1.

Methods: The molecular role of Mcl-1 in HER2+ BC cells under hypoxic conditions was investigated using genetical as well as pharmacological approaches followed by proliferation, survival, and spheroid forming assays, subcellular fractionation, RT-PCR as well as immunoprecipitation/blotting. Mcl-1(Δ/null) Murine Embryonic Fibroblasts (MEFs) were used to verify the specific anti-Mcl-1 activity of the novel small molecule inhibitor EU-5346 (EUTROPICS, Cambridge).

Results: We demonstrate for the first time a strong correlation between rapid induction of Mcl-1 and HIF-1α under hypoxic conditions in HER2+ BC cells. Genetic depletion of Mcl-1 decreased HER2 and HIF-1α levels followed by inhibition of BC cell survival. Surprisingly, Mcl-1 protein levels did not change after genetic depletion of HER2, indicating a regulatory role of Mcl-1 upstream of HER2. The functional interrelation of Mcl-1 and HER2 under hypoxic conditions was supported by Mcl-1/HER2 co-localization within the mitochondrial fraction as well as the formation of a Mcl-1/HER2- protein complex. Moreover, our data demonstrate that Mcl-1 stabilizes HER2 protein levels via inhibition of ubiquitination. Given its proposed key role in the survival of cancer cells in general, Mcl-1 is the current focus of widespread cancer drug development efforts. Indeed, EU-5346 specifically blocks the interaction of Mcl-1 with pro-apoptotic BH3-only proteins. Similar to genetically targeting Mcl-1 EU-5346 induced cell death and decreased spheroid formation in HER2+ BC cells.

Conclusions: Taken together, our data demonstrate the critical role of Mcl-1 in HER2+ BC cell survival under hypoxic conditions and provide the preclinical framework for the therapeutic use of novel Mcl-1-targeting agents to improve patient outcome in BC.

Disclosure: No conflict of interest disclosed.

V432 - GAIN2: Adjuvant phase III trial comparing an intensified dose-dense adjuvant therapy with EnPC compared to a dose-dense, dose-adapted therapy with dtEC-dtD in patients with primary high risk breast cancer: Results of the second safety interim analysis

Möbus V.1, Lück H.-J.2, Forstbauer H.3, Wachsmann G.4, Ober A.5, Schneeweiss A.6, Christensen B.7, Ekkehard E.8, Grischke E.-M.9, Höffkes H.-G.10, Klare P.11, Ko Y.D.12, Schmatloch S.13, Burchardi N.14, Loibl S.15, von Minckwitz G.14, German Breast Group

1Klinikum Frankfurt Höchst GmbH, Frankfurt am Main, Germany, 2Gynäkologisch-Onkologische Praxis am Pelikanplatz, Hannover, Germany, 3Hämatologisch-Onkologische Schwerpunktpraxis, Troisdorf, Germany, 4Klinikum Sindelfingen-Böblingen, Böblingen, Germany, 5St. Vincenz Krankenhaus, Limburg, Germany, 6Universitäts-Klinikum, Heidelberg, Germany, 7Ruppiner Kliniken GmbH, Neuruppin, Germany, 8Klinikum, Mutlangen, Germany, 9Universitätsklinikum, Tübingen, Germany, 10Klinikum, Fulda, Germany, 11Praxisklinik Krebsheilkunde und Brustzentrum, Berlin, Germany, 12Tumorzentrum, Bonn, Germany, 13Elisabeth-Krankenhaus, Kassel, Germany, 14German Breast Group, Neu-Isenburg, Germany, 15German Breast Group, GBG Forschungs GmbH, Neu-Isenburg, Germany

Introduction: Combined chemotherapy requires compromises in terms of dosage and treatment interval due to toxicities. Sequential administration of monotherapies allows high doses of single substances and dose-dense intervals. Such regimens proved to be very effective in early breast cancer with high risk of recurrence. Nab-paclitaxel (nP) leads to a more favorable toxicity profile and greater efficacy compared with solvent-based taxanes.

Methods: The GAIN2 study compares toxicity and efficacy of a pre-defined intense dose-dense regimen (EnPC; E:150 mg/m2, nP:330 mg/m², C:2000 mg/m²) with a dose-dense regimen, where single doses are adjusted depending on individual hematological and non-hematological toxicities (dtEC-dtD; E:38-120 mg/m², C:450-1200 mg/m², D: 60-100 mg/m²).

Primary endpoint is the invasive disease-free survival in patients with primary node-positive or high-risk node-negative breast cancer (luminal A: ≥LN; luminal B: N+; HER2pos/TNBC: N0/N+)

A first safety interim analysis after 200 with completed chemotherapy has been published (Noeding et al. Ann Oncol 2014). The second safety interim analysis with 900 treated patients will be presented.

Results: In terms of hematological adverse events, the rates of febrile neutropenia grade 3-4 (12% vs. 8%) and thrombopenia grade 3-4 (12% vs. 5%) was significantly higher in the EnPC arm. As for the non-hematological side effects grade 1-4, the rate of increased alkaline phosphatase (59% vs. 40%), increased ALAT (69% vs. 59%), peripheral neuropathy (83% vs. 68%), arthralgia (63% vs. 49%), myalgia (48% vs. 41%), and bone pain (25% vs. 17%) was significantly higher in the EnPC arm whereas the rate of epistaxis (10% vs. 25%), edema (13% vs. 26%), and hand-foot-syndrome (12% vs. 28%) was significantly higher in the dtEC-dtD arm. There were no differences between the treatment arms for the toxicities of special interest (cranial nerves affections, macula degenerations, anaphylactic reactions). In the EnPC arm, 30% required dose-reductions due to hematological toxicities compared to only 10% in the dtEC-dtD arm (p < 0.001). The dose could be escalated to the maximum in more than a third of the patients receiving dtEC-dtD. In 9% of women a reduction was required in the 4th cycle of docetaxel.

Conclusion: Due to similar toxicity profiles, the study will be continued without changes.

Disclosure: No conflict of interest disclosed.

V433 - Routine care reality of patients with metastatic breast cancer 1995-2014: Improvement of survival is restricted to hormone- and HER2-receptor-positive tumors

Weide R.1, Feiten S.2, Friesenhahn V.2, Heymanns J.1, Kleboth K.2, Thomalla J.1, van Roye C.1, Köppler H.1

1Praxisklinik für Hämatologie und Onkologie, Koblenz, Germany, 2Institut für Versorgungsforschung in der Onkologie, Koblenz, Germany

Introduction: Improvements in survival rates of patients suffering from metastatic breast cancer have been found in randomized clinical trials over the past years. Due to necessary selection criteria it is difficult to transfer these results into routine care. The aim of this analysis was to determine overall survival of unselected patients who received their treatment in an oncology group practice.

Methods: Retrospective analysis of all patients with metastatic breast cancer who were seen between 06/1995 and 12/2014. Relevant clinical data were transferred from clinical files into a database and analyzed statistically using SPSS and SURVSOFT.

Results: Data of 767 female patients with a median age of 61 years (31-93) (time of metastasis) were analyzed. 81% were postmenopausal, 23% suffered from primary metastatic disease. Sites of metastases were distributed as follows: 49% visceral, 34% bone, 9% lymph nodes, 4% CNS and 4% others. Mean number of metastatic sites was 1.5 (1-6). 77% of tumors were hormone-receptor-positive, 18% HER2-positive and 9% triple-negative. Median overall survival was 35 months (0-321+), disease specific survival was 37.3 months. Disease specific 5-year survival rate was 34%. Overall survival was significantly correlated with the sites of metastases, number of involved organs, disease-free survival since initial diagnosis and hormone-receptor status. Patients with a hormone-receptor-positive tumor had a median overall survival of 38 months (0-321+) compared to patients with a triple-negative tumor, who showed a median overall survival of 13 months (0-109+). Patients with a HER2-positive tumor had a median overall survival of 39 months (0-189+). Comparisons of disease specific survival with registry data from the US, Great Britain and different German areas revealed a significant better survival after 1 year (86% versus 44-73%) and after 5 years (34% versus 13-25%). A trend, although statistically non-significant, showing an continuous improvement in survival rates could be observed. Patients who received diagnosis between 1995 and 1999 had a median disease specific survival of 30.5 months. Patients who were diagnosed 2010 or later lived 43.0 months in comparison.

Conclusions: Survival rates of patients suffering from metastatic breast cancer with hormone-receptor- and / or HER2-positive tumors can be extended significantly in routine care. This is probably due to the sequential use of targeted therapies.

Disclosure: No conflict of interest disclosed.

V434 - EVI1 promotes breast cancer cell growth via MAPK signaling, but independent of the estrogen

Wang H.1,2, Schäfer T.1, Konantz M.1, Reich S.2, Braun M.3, Perner S.3, Varga Z.4, Moch H.4, Kanz L.2, Schulze-Osthoff K.5,6, Lengerke C.1,2,7

1Department of Biomedicine, University Hospital, Basel, Switzerland, 2Department of Internal Medicine II, University Hospital, Tuebingen, Germany, 3Department of Prostate Cancer Research, Institute of Pathology, University Hospital of Bonn, Bonn, Germany, 4Institute of Surgical Pathology, University Hospital, Zurich, Switzerland, 5Interfaculty Institute of Biochemistry, University of Tuebingen, Tuebingen, Germany, 6German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ), Heidelberg, Germany, 7Clinic for Hematology, University Hospital, Basel, Switzerland

Introduction: The transcriptional regulator EVI1 has been mainly studied in myeloid leukemia, but expression has also been detected in solid tumors. While almost no data exist on EVI1 in breast carcinoma (BC), a previous report suggests that overexpression may indicate poor clinical outcome in estrogen receptor (ER) negative tumors (Patel et al., 2011). Here, we analyze the roles and molecular targets of EVI1 in human BC.

Methods: EVI1 expression was analyzed by qRT-PCR and immunoblot analysis in 8 human BC cell lines and a tissue microarray (TMA) containing 373 patient BC samples surveyed by immunohistochemistry. Lentiviruses containing EVI1-shRNAs were used to generate EVI1-knockdown cells, which were then compared to control-treated cells in various assay formats: cell cycle, BrdU, apoptosis (PI and Annexin-V/7-AAD staining, Caspase 3/7 and PARP activity), immunophenotypic, and qRT-PCR. Xenotransplantation experiments were performed in zebrafish embryos as described (Konantz et al., 2012). Smad3 siRNAs were introduced by transient transfection and β-estradiol (50-100 nM) used for cell culture and zebrafish treatment.

Results: EVI1 was expressed in most breast cancer samples independent of ER status. EVI1 gene expression levels influenced long-term survival in ER- (p = 0.026) but not ER+ patients. In BC cell lines and primary cells, EVI1 knockdown impaired growth by suppressing proliferation and enhancing apoptosis sensitivity. The growth reduction was observed in both EVI1-knockdown ER- as well as ER+ cells, but in the latter β-estradiol could provide a functional rescue. Consistently, in vivo addition of β-estradiol could rescue tumor initiation and growth in zebrafish transplanted with ER+ but not ER- BC cells. On the molecular level, knockdown of EVI1 led to a cell cycle arrest by de-repressing its known target gene SMAD3 and thereby suppressing the cell cycle proteins p21 and p15. siRNA against SMAD3 only partially rescued growth in EVI1-knockdown cells, indicating that additional pathways are also involved. Indeed, EVI1 knockdown strongly inhibited the MAPK-pathway, thereby additionally regulating BC cell proliferation.

Conclusion and discussion:EVI1 is widely expressed amongst breast carcinoma cells where it influences apoptosis and proliferation via SMAD3 repression and activation of the MAPK-pathway. The precise molecular mechanisms underlying the effect of EVI1 on MAPK activity are currently under investigation.

Disclosure: No conflict of interest disclosed.

V435 - Cascade of toxicological tests: breast cancer treatment affects trophoblast cells in 2D and explant culture

Fröhlich K.1, Schmidt A.1, Heger J.1, Lupp A.2, Avemarg S.1, Markert U.R.1

1Friedrich Schiller University, Placenta Laboratories, Department of Obstetrics, Jena, Germany, 2Friedrich Schiller University, Institute of Pharmacology and Toxicology, Jena, Germany

Introduction: Pregnancy and breast cancer is a rare coincidence, but breast cancer is one of the most common malignancies during pregnancy. Guidelines on treatment for breast cancer during pregnancy already exist and demonstrate that pregnancy-associated breast cancer (PABC) can be treated according to recommendations for non-pregnant women with breast cancer. Nonetheless, a decreased birth weight is often observed in newborns which may be due to harmful effects of chemotherapy on trophoblast cells. Therefore, we aimed to determine the toxicity of chemotherapeutic drugs on these cells.

Methods: HTR-8/SVneo and JEG-3 cells cultured as monolayers were grown in 96-well plates. Placental villous tissue explants (PVTE) were obtained after spontaneous delivery or cesarean section (n = 3). Trophoblast cells and PVTE were incubated with doxorubicin, docetaxel, 5-fluorouracil or vincristine for at least 48 hours. The metabolic activity was evaluated via MTS assay. Supernatant analyses of glucose, lactate, lactate dehydrogenase (LDH), human chorionic gonadotropin (hCG), estrogen and progesterone were performed. Data were analyzed with SPSS 22 using a linear mixed model. Furthermore, PVTE were embedded in paraffin for histological examinations.

Results: The metabolic activity of PVTE, as evaluated via MTS assay, was only reduced by doxorubicin or docetaxel, but not by 5-fluorouracil or vincristine treatment. In contrast, the metabolic activity of HTR-8/SVneo and JEG-3 cells was significantly reduced by all tested drugs. Glucose, lactate and LDH as potential markers of toxicity were significantly affected in the supernatant of PVTE treated with the different chemotherapeutics. In contrast, expression of hCG, estrogen and progesterone was not modified. Hematoxylin/eosin staining revealed more morphological anomalies in 5-fluorouracil or vincristine than in doxorubicin or docetaxel treated PVTE.

Conclusion: Toxicological investigations on trophoblast cell lines enable the detection of harmful effects of chemotherapy but fail to simulate in vivo conditions. Therefore, PVTE display a suitable tool for studying effects of different chemotherapeutic drugs during pregnancy. Among the analyzed parameters, only the metabolic markers glucose, lactate and LDH and the MTS assay appear to be useful for detecting harmful effects and are recommended for further investigations. Hematoxylin/eosin staining indicates early toxic morphological changes, even when the MTS assay did not.

Disclosure: No conflict of interest disclosed.


Allogene Transplantation

P436 - Reduced platelet transfusions and earlier platelet engraftment using alemtuzumab based conditioning regimen in allogeneic SCT

Neumann T.1, Schneidewind L.2, Thiele T.3, Schulze M.1, Klenner A.1, Busemann C.1, Pink D.4, Greinacher A.3, Dölken G.1, Krüger W.1

1Universitätsmedizin Greifswald, Klinik für Innere Medizin C - Hämatologie / Onkologie, Greifswald, Germany, 2Universitätsmedizin Greifswald, Klinik für Urologie, Greifswald, Germany, 3Universitätsmedizin Greifswald, Institut für Immunologie und Transfusionsmedizin, Greifswald, Germany, 4HELIOS Klinikum Bad Saarow, Klinik für Hämatologie, Onkologie und Palliativmedizin, Bad Saarow, Germany

In patients undergoing allogeneic stem-cell transplantation (SCT) conditioning regimens containing alemtuzumab instead of antithymocyte globulin (ATG) may result in an earlier platelet engraftment and a reduced number of platelet transfusions. We performed a retrospective, single center, case-control study analyzing time to engraftment and transfusion needs using alemtuzumab in comparison to ATG as part of conditioning protocol. Median values for time to platelet engraftment, number of transfused platelet concentrates and number of transfused red cell concentrates were 12 vs. 19.5 days (p < 0.001), 2 vs. 14 (p < 0.001) and 6 vs. 14.5 (p = 0.003) in the alemtuzumab and ATG group. Time to leukocyte engraftment did not differ significantly with median 15 days in both groups. Patients in the ATG group showed significant lower median platelet counts at the day of stem-cell infusion (38 vs. 95.5 Gpt/l, p = 0.008) and higher values for median C-reactive protein (CRP) after first antibody infusion (69.0 vs. 43.6 mg/l, p = 0.001) compared to alemtuzumab group. Test for significance was done by using Wilcoxon rank sum test. Subgroup analysis considering the type of ATG used (Thymoglobulin vs. ATG Fresenius) revealed that differences between alemtuzumab and ATG group were more due to effects of ATG Fresenius than Thymoglobulin. Median values for time to platelet engraftment, number of transfused platelet concentrates and number of transfused red cell concentrates were 14 vs. 23 days, 5 vs. 21 and 8 vs. 21 in the Thymoglobulin and ATG Fresenius group.

Disclosure: No conflict of interest disclosed.

P437 - CD19+CD21low B-cells as biomarker for the prediction and monitoring of response to immunosuppressive therapy for chronic graft-versus-host disease

Kralj M.1, Kuzmina Z.1, Weigl R.1, Rottal A.2, Körmöczi U.2, Pickl W.2, Greinix H.3

1Department of Internal Medicine I, Medical University of Vienna, Vienna, Austria, 2Institute of Immunology, Medical University of Vienna, Vienna, Austria, 3Department of Internal Medicine, Medical University of Graz, Graz, Austria

Chronic graft-versus-host disease (cGVHD) is a serious, life-threatening complication of allogeneic hematopoietic stem cell transplantation (HCT). Afflicted patients present heterogeneous organ manifestations varying in severity and patients' impairment of quality of life. Here, we investigated cellular biomarkers for the prediction of clinical response to various immunosuppressive agents (ECP, cyclosporine A (CsA), tacrolimus and sirolimus) in the course of cGVHD.

Different B-cell subsets, inluding CD19+CD21low B-cells were analyzed by flow cytometry in the peripheral blood (PB) of cGVHD patients at the onset of immunosuppressive therapy (IS) and again 6 months after start of therapy. At the same time patients were evaluated for the presence of clinical signs and symptoms of cGVHD according to NIH recommendations. Different B-cell subsets were compared overall between responders and non-responders to IS and within each IS treatment group. The study included 144 patients with a median age of 39 (range, 17-70) years. Response to therapy was defined as decrease in severity for at least 1 score and no new organ manifestations of cGVHD. Overall the response rate to IS at 6 months after start of treatment was 54% (78 out of 144 patients). 47 patients received ECP, 51 CsA, 18 tacrolimus and 28 sirolimus.

The overall absolute numbers of CD19+ B-cells in PB at the start of IS therapy were significantly lower in non-responders compared to responders (157 × 106/L vs 307 × 106/L, p = 0.003). Overall, relative numbers of CD19+CD21low B-cells measured at the onset of IS were significantly elevated in patients that later did not respond to therapy compared to the ones that did (17.94% vs 13.55%, p = 0.017). A similar trend was observed within the sirolimus and tacrolimus IS groups. At 6 months after start of IS, percentages of CD19+CD21low B-cells were significantly higher in non-responders compared to responders (20.16% vs 7.55%, p < 0.001). Similar significant observations were seen in all four therapy groups. By comparing relative values of CD19+CD21low B-cells at the onset of IS and 6 months after, we observed a significant decrease in patients who responded to therapy (13.55% vs 7.55%, p < 0.001). In patients who did not respond to IS, these values stayed elevated (17.94% vs 20.16%, p = 0.243).

Our results show that CD19+CD21low B-cells could serve both as predictive biomarker for response to cGVHD treatment as well as for monitoring disease activity in cGVHD.

Disclosure: No conflict of interest disclosed.

P438 - Cytomegalovirus induce apoptosis and alloresponse in acute leukemia cells

Koldehoff M.1, Lindemann M.2, Opalka B.3, Mühlenberg T.4, Ross R.S.5, Elmaagacli A.H.1,6

1University of Duisburg-Essen, Department of Bone Marrow Transplantation, Essen, Germany, 2University of Duisburg-Essen, Institute for Transfusion Medicine, Essen, Germany, 3University of Duisburg-Essen, Department of Hematology, Essen, Germany, 4University of Duisburg-Essen, Department of Medical Oncology, Essen, Germany, 5University of Duisburg-Essen, Institute of Virology, Essen, Germany, 6HELIOS Schwerin, Department of Oncology and Hematology, Schwerin, Germany

Introduction: Cytomegalovirus (HCMV) reactivation occurs frequently after hematopoietic stem cell transplantation and is associated with an increased treatment-related mortality. Induction of apoptosis by HCMV is unusual because HCMV utilizes various strategies to prevent apoptosis in infected cells in order to delay cell death and maintain viral replication.

Cells and Methods: Various acute leukemic cell lines were purchased for in vitro experiments, and further bone marrow or peripheral blood stem cell samples were collected from healthy volunteers after informed consent in accordance with institutional guidelines. For infection we used the AD169 HCMV strain (ATCC-VR-538, American Type Culture Collection, Manassas, VA, USA). Various different cellular and molecular biological studies with HCMV-infected cells were performed.

Results: Here we show that HCMV can infect the acute leukemia cell lines Kasumi-1 (AML) and SD-1 (BCR-ABL-positive ALL), which inhibited their proliferation and induced apoptosis in almost all cells after 14 days. Although HCMV induced a significant up-regulation of the anti-apoptotic gene cFLIP and the anti-stress gene Gadd45a, and simultaneously down-regulated the pro-apoptotic genes p53, Gadd45gamma in Kasumi-1 and SD-1 cells, we found that these anti-apoptotic mechanisms failed in HCMV-infected acute leukemia cells and apoptosis occurred via a caspase-dependent pathway. ELISpot data indicated that peripheral blood mononuclear cells (PBMC) of healthy individuals had significantly higher numbers of interferon-gamma, granzyme B and perforin secreting cells when stimulated with Kasumi-1 with vs. without HCMV infection; indicating that alloresponses were enhanced due to HCMV infection of the Kasumi-1 cell line. In addition, HLA-DR expression as determined by FACS analysis was significantly increased on PBMC of healthy individuals when stimulated with Kasumi-1 with vs. without HCMV infection. Finally, proliferation of Kasumi-1 with vs. without HCMV infection was significantly lower.

Conclusions: We conclude that HCMV can provide anti-leukemic effects in vitro. To determine if this phenomenon may be clinically relevant further investigations will be required.

Disclosure: No conflict of interest disclosed.

P439 - No impact of geographic distance to patient residence on the outcome of allogeneic hematopoietic cell transplantation (alloHCT): Retrospective analysis in a JACIE-accreditated transplant center

Brand S.1, Dietrich S.2, Hegenbart U.2, Bondong A.2, Luft T.2, Schönland S.2, Ho A.D.2, Görner M.1, Dreger P.2

1Städtische Kliniken, Bielefeld, Germany, 2Universitätsklinikum Heidelberg, Innere Medizin V, Heidelberg, Germany

The purpose of this study was to assess the impact of distance to residence on alloHCT outcomes in a JACIE-accreditated center in Germany.

Study design and patients: In a single center retrospective analysis, the distance to primary residence was assessed for all consecutive patients who underwent alloHCT for hematological malignancy 2005 and 2011 and assigned to 5 categories: Local (0-50 km), regional A (>50-100 km), regional B (>100-150 km), national (>150 km), and international. The impact of distance category on non-relapse mortality (NRM), relapse, progression-free survival (PFS), and overall survival (OS) was studied by log-rank comparisons.

Results: A total of 562 patients were assigned to local (229), regional A (166), regional B (76), national (75), and international residence (15). Median age was 53 (18-70) years; diagnoses were myeloid malignancies (53%), lymphoma (16%), myeloma (13%), CLL (10%), and ALL (8%); disease status was refractory at alloHCT in 17%; median EBMT score (Gratwohl) was 2 (0-7). Donors were matched siblings (31%), unrelated donors (68%), and other (1%). Compared to local/regional patients, the national/international group contained significantly less patients with myeloid malignancy (40% vs 56%, p < 0.0001), significantly more patients with myeloma (25% vs. 10%), and had a significantly poorer EBMT score (3 vs 2, p < 0.0001), but was matched for all other baseline variables. There were no differences between local, regional A and regional B patients for any survival endpoint. Similarly, NRM (HR 1.04; 95% CI 0.58-1.87) and OS (HR 0.85; 95%CI 0.61-1.2) were not significantly different between pooled local/regional patients and national/international patients. However, local/regional patients had a significantly lower relapse risk (HR 0.59; 95% CI 0.4-0.89; p 0.011), translating to a significant PFS advantage (HR 0.69; 95% CI 0.49-0-97; p 0.032). This effect disappeared after adjusting for diagnosis. Significant predictors of PFS were disease status (refractory unfavorable), EBMT score, and diagnosis (myeloma unfavorable, CLL favorable).

Conclusion: This study failed to detect a detrimental impact of distance from residence to transplant center on NRM and disease control after alloHCT for hematological malignancies. Albeit further studies are needed, these results suggest that distance to HCT center should not overrule criteria such as JACIE accreditation and centre experience when selecting a transplant center for a specific patient.

Disclosure: No conflict of interest disclosed.

P440 - GTPase Rap1 regulates the shear stress-resistant adhesion of Mesenchymal Stromal Cells (MSCs)

Steinmann J.1, Fleck E.2, Henschler R.2, Fisch P.1, Scheele J.1, Kropshofer H.3

1University Hospital Medical Center Freiburg, Department of Pathology, Freiburg, Germany, 2Institute for Transfusion Medicine and Immune Hematology, German Red Cross Blood Service, Medical Center of the Johann Wolfgang Goethe University, Frankfurt, Germany, 3F. Hoffmann- La Roche Ltd., Pharmaceuticals Division, Basel, Switzerland

Introduction: Mesenchymal stromal cells (MSCs) are increasingly used as an intravenously applied cellular therapy. Rap1 is a major regulator of integrin activation, and therefore a candidate molecule to modulate homing properties of MSCs. We here studied the adhesion of human and murine MSCs to endothelial cells and recombinant integrin ligands and their homing behavior in mice.

Methods: MSCs deficient in Rap1A and/ or Rap1B, mediated via siRNA medicated knockdown or derived from the respective knockout mice were investigated with regards to MSC migration and adhesion in a flow-chamber model.

Results: siRNA mediated knockdown of Rap1A or Rap1B led to decrease in both basal and Stromal Derived Factor-1 (SDF-1) induced firm adhesion of MSCs in a flow-chamber model, whereas knockdown of both Rap1A+B also inhibited arrest of flowing MSCs to inflammatory endothelial cells. Further siRNA experiments indicated that Rap1-Guanosin Nucleotide Exchange Factor (GEF) PDZ-GEF1 is responsible for constitutive adhesion of MSCs to Vascular Cell Adhesion Molecule (VCAM)-1, whereas treatment of MSCs with siRNA against EPAC2 inhibited predominantly the adhesion induced by the chemokine, SDF-1. We next established Rap1 deficient MSCs from Rap1A knockout mice. Whereas homozygous MSCs were not viable in culture, heterozygous Rap1A +/- MSCs recapitulated the adhesion deficit observed in human MSCs. The EPAC stimulator 8-Br-(4-PCT)-cAMP induced Rap1 activation and increased adhesion of MSCs. Homing experiments in tumor bearing mice indicated a shift in the homing pattern of Rap1A deficient mMSCs compared with wt MSCs, away from lung, liver and hematopoietic tissues towards muscle, kidney and lymph nodes.

Conclusions: Together, our findings demonstrate an essential role for Rap1mediated pathways in MSC migration, and provide a strategy to engineer MSC grafts in an effort to improve their therapeutic efficiency.

Disclosure: No conflict of interest disclosed.

P441 - Invasive mycosis after allogeneic stem cell transplantation at the university medical centre of Würzburg - Incidence of the disease as well as morbidity and mortality of patients with the condition

Möller A.-K.1, Einsele H.2, Heinz W.J.2

1Klinikum Heilbronn GmbH, Innere Medizin I, Heilbronn, Germany, 2Universitätsklinikum Würzburg; II. Medizinische Klinik, Würzburg, Germany

Introduction: Invasive fungal infections (IFI) still present an important and life threatening complication especially for recipients of an allogeneic stem cell transplantation (SCT). Despite improved diagnostics, new prophylactic options and therapies, mortality and morbidity of patients with a systemic mycosis remain at a high level. Knowledge about the incidence, timing and impact of IFI is relevant to choose the appropriate diagnostic and antifungal strategy.

Methods: To record the incidence of IFI among patients after allogeneic SCT, a prospective, non-interventional study was conducted. A primary endpoint was to analyse the morbidity and mortality rates of patients with IFI. The clinical course of the patients was actively monitored until 100 days after the start of the conditioning chemotherapy and a follow-up examination was carried out after six months. IFI were classified according to the EORTC-MSG criteria revised in 2008.

Results: Within six months a total of 41 patients were included in the survey. During this study 22% of the patients contracted a probable and 17% a possible invasive mycosis. One probable mycosis was caused by Fusarium spp., the remaining infections were ascribed to Aspergillus spp. (94%). At the end of the first month the risk to develop a probable mycosis was at 17%, which increased to 22% at the end of the sixth month. The patients without evidence of an IFI were hospitalized for SCT for an average of 41.8 days. Patients with IFI spent an average of 55.9 days at the hospital, revealing a significant difference in the length of hospital stay (p = 0.015). Deducting two possible cases with unknown survival, in the first half year after SCT 16% of patients without IFI had passed away while 20% of those with a possible aspergillosis and 66.6% of those with a probable mycosis had died. The estimated cumulative survival time of the patients without systemic mycosis was 170.9 days and of those with IFI was 131.8 days (p = 0.016). The death of the patients with a possible or probable infection occurred an average of 40 days after the initial diagnosis of the IFI.

Conclusion: In the survey presented here, a high incidence of invasive IFI after SCT was detected. The mortality of the affected patients was significantly higher than in SCT recipients without IFI. Improved diagnostic options may increase the rate of diagnosed infections. Prophylactic and therapeutic strategies have to be adapted to the clinical situation.

Disclosure: No conflict of interest disclosed.

P442 - Allogeneic stem-cell transplantation for early relapse of multiple myeloma after autologous stem-cell transplantation

Schmidt V.1, Mügge L.-O.1, Klink A.1, Hilgendorf I.1, Hochhaus A.1, Sayer H.G.1,2

1Klinik für Innere Medizin II, Universitätsklinikum Jena, Hämatologie und internistische Onkologie, Jena, Germany, 24. Medizinische Klinik, HELIOS-Klinikum Erfurt, Hämatologie und internistische Onkologie, Hämostaseologie, Erfurt, Germany

Introduction: Prognosis of patients with early relapse of multiple myeloma (MM) after conditioning with melphalan followed by autologous hematopoietic stem-cell transplantation (autoHSCT) is poor, and treatment options are limited.

Methods: We report a retrospective analysis of 12 MM patients (8male/4female; median age 49 (37-63) years) who received an allogeneic HSCT (alloHSCT) between 03/2008 and 09/2014 at our center. Before alloHSCT, patients had received a median of 5 (range 2-6) therapy lines including a median of 2 (1-3) previous autoHSCTs. Relapse occurred 7 (3-9) months after the last autoHSCT or 12 (12-18) months in 3 patients who received maintenance therapy with bortezomib. Conditioning therapy consisted of treosulfan/fludarabin (9/12), melphalan/fludarabin (2/12) or fludarabin/TBI (1/12). HSCT was performed from a matched related donor (1/12), from an HLA-matched (10/10) unrelated donor (7/12), or a mismatched unrelated donor (4/12). ATG was added in case of unrelated donor HSCT.

Results: After alloHSCT, 2 patients (17%) achieved a complete remission, 3 patients (25%) reached a very good partial remission, and 6 patients (50%) a partial remission. One patient (8%) experienced a graft failure, and despite an autologuous rescue transplantation died due to relapse of MM within 4 months. After a median follow up of 55 months, 7 patients (58%) are still alive and median overall survival has not been reached yet. A relapse occurred in 8 patients (66%). Median progression free survival was 8 months. An acute GvHD (≥ grade III) occurred in 5 patients, but was sensitive to GvHD-directed therapy. Three patients (25%) who developed a mild to moderate chronic GvHD are still alive and in a stable remission 42, 50, and 69 months after alloHSCT, even though they received no further MM-directed therapy and were not in a CR early after alloHSCT. All patients (5/12) with a deletion of chromosome 13 (del13q) together with a translocation involving chromosome 14 [t(4;14) or t(11;14)] died within 31 (median 13) months after alloHSCT. In comparison to the other 7 patients, the difference was significant at p < 0.01 (Fisher exact test).

Conclusion: AlloHSCT is a feasible therapeutic option for heavily pretreated patients with an early relapse of MM after autoHSCT. Especially patients with chronic GvHD may profit from a graft-versus-myeloma effect. The combination of del13q and either t(4;14) or t(11;14) is a strong negative predictor, even after alloHSCT.

Disclosure: No conflict of interest disclosed.

P443 - Steroid-induced hyperglycemia adversely impacts outcome in graft-versus-host disease

Stauber M.1, Aberer F.2, Zebisch A.1, Neumeister P.1, Greinix H.T.1, Sill H.1, Sourij H.2, Wölfler A.1

1Medizinische Universität Graz, Klin. Abteilung für Hämatologie, Graz, Austria, 2Medizinische Universität Graz, Klin. Abteilung für Endokrinologie und Stoffwechsel, Graz, Austria

Introduction: Graft-versus-host disease (GvHD) is a common complication after allogeneic stem cell transplantation. Since the current first-line treatment is based on high-dose glucocorticoid therapy, steroid-induced hyperglycemia develops frequently in these patients. However, little is known about the impact of hyperglycemia on outcome in GvHD.

Methods: We performed a retrospective analysis of 100 patients, who received systemic glucocorticoid therapy due to acute or chronic GvHD and investigated the impact of hyperglycemia on outcome in these patients. For each subject regular blood glucose measurements during glucocorticoid therapy and transplant- as well as GvHD-related parameters, such as underlying disease, time to onset of GvHD after transplantation, number of affected organs, response to glucocorticoids, insulin therapy and (fungal) infectious complications were recorded. For analysis of the impact of hyperglycemia, the median glucose level was taken and patients were divided into 4 groups (median glucose level < 100 mg/dl, 100-120 mg/dl, 121-150 mg/dl and >150 mg/dl).

Results: With a median of 42 blood glucose measurements per patient, only nine patients had a median glucose level <100 mg/dl and were therefore assigned to group 1. In 29 cases the median glucose value was between 101 and 120 mg/dl (group 2), 30 patients were assigned to group 3 and the remaining 32 cases displayed a median glucose level >150 mg/dl. While median overall survival (OS) was not reached in group 1, increasing median glucose levels were associated with decreasing median OS of 20, 17 and 8 months for groups 2, 3 and 4, respectively (p < 0,01). Accordingly, 5-year OS was significantly lower for group 4 when compared to all other groups (p < 0,01) and for group 2 and 3 when compared to group 1 (p < 0,01). In addition, insulin treatment, which was given in 49 patients, was associated with a shorter median OS (8 versus 29 months, p < 0,01). While the number of relapses was low (n = 9) and not affected by hyperglycemia, non-relapse related mortality was responsible for most cases of death (n = 51). In particular, patients with invasive fungal infections had a poor outcome (median OS of 5 months versus 24 months, p < 0,001).

Conclusions: In this retrospective analysis, we observed an adverse impact of glucocorticoid-induced hyperglycemia on outcome in patients with GvHD. Prospective trials testing a stringent glycemic control are therefore urgently needed in this clinical setting.

Disclosure: No conflict of interest disclosed.

P444 - Minimal toxicity conditioning with fludarabine and 2 Gy total body irradiation for allogeneic hematopoietic cell transplantation

Dörfel D.1,2, Mück F.1, Vogel W.1, Wirths S.1, Möhle R.1, Kanz L.1, Faul C.1, Bethge W.A.1

1Universitätsklinik Tübingen, Innere Medizin II - Onkologie, Hämatologie, Klinische Immunologie, Rheumatologie und Pulmologie, Tübingen, Germany, 2Clinical Collaboration Unit Translational Immunology, German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ), Partner site Tuebingen, Tübingen, Germany

Introduction: Non-myeloablative conditioning with fludarabine (FLU, 3×20 mg/m2) and 2 Gy total body irradiation (TBI) is currently the least toxic conditioning regimen used for the treatment of hematologic malignancies with hematopoietic cell transplantation (HCT) primarily in an elderly or comorbid patient population.

Methods: We report retrospectively our single center experience with this regimen in 40 consecutive patients undergoing allogeneic HCT after minimal toxicity conditioning with FLU/2Gy TBI followed by postgrafting immunosuppression using a calcineurininhibitor combined with mycophenolate mofetil with or without sirolimus from 2004 to 2014.

Results: 40 adult patients received allogeneic HCT during this period for the treatment of acute myeloid leukemia (n = 25), non-Hodgkin lymphoma (n = 10), myelodysplastic syndrome (n = 4), and multiple myeloma (n = 1). The median age at time of HCT was 62 (range 25 to 76) years. Peripheral blood stem cells were exclusively used as graft source. Donors were matched related (n = 5), matched unrelated (n = 33) or mismatched unrelated (n = 2). Median follow-up was 13 months with a Kaplan Meier estimated overall survival (OS) for all patients of 55%, 37% and 32% at 1, 3, and 5 years, respectively. There was no significant difference in OS/EFS in different age groups. The non-relapse mortality (NRM) at day 100, 1 year and 3 years was 5%, 20% and 23%. 38% of patients developed a relapse of the underlying disease (AML 10/25 = 40%, NHL 3/10 = 30%, MDS 1/4 = 25%, MM 1/1 = 100%). The cumulative incidence of acute GvHD was 23% (≥II° 13%) and of chronic GvHD was 73% (limited = 43%, extensive = 30%).

Conclusion: In conclusion, our data suggest that RIC with FLU/TBI enables long-term survival even in an elderly and comorbid patient population. However, incidence of chronic GVHD is high, which may have a negative impact on quality of life.

Disclosure: No conflict of interest disclosed.

P445 - Urological complications and BK virus associated hemorrhagic cystitis under allogenic stem cell transplantation

Schneidewind L.1, Neumann T.2, Burchardt M.1, Krüger W.2

1Universitätsmedizin Greifswald, Klinik für Urologie, Greifswald, Germany, 2Universitätsmedizin Greifswald, Klinik für Innere Medizin C, Hämatologie/Onkologie, Greifswald, Germany

Introduction: Every year 50.000 patients receive a stem cell transplantation worldwide, but there is a lack of data focussed on urological complications under this therapy. To fill this gap of knowledge we performed a retrospective analysis of all adult patients undergoing allogenic stem cell transplantation from 01/2011 to 06/2013 in our institution.

Methods: All patient records of adult patients with hematological diseases undergoing their first allogenic stem cell transplantation from 01/2011 to 06/2013 were collected. The statistical analyses was performed with SPSS 22.0. Statistical tests performed were Pearson´s correlation, Qui-Square testing and logistic regression.

Results: We identified 39 patients (22 males and 17 females). Twenty five patients (64.1%) had a urological complication. Most frequent complications were bacterial urinary tract infection (n = 13; 33.3%), acute renal failure (n = 6; 15.4%) and BK virus associated hemorrhagic cystitis (n = 5; 12.8%). We observed an association of creatinine increase (about 20 µmol/l at time of onset of BK viruria) with BK viruria (Pearson`s correlation 0.64; p = 0.01) and BK viruria is significantly linked to acute renal failure (Pearson´s correlation 0.35; p = 0.029). In univariate regression BK viruria is significant linked to urological complication (p = 0.025). We could not identify a significant risk factor for urological complications in multivariate analysis.

Conclusion: The most frequent urologic complications in adult allogenic stem cell transplantation are bacterial urinary tract infection and BK virus associated hemorrhagic cystitis. We suggest that BK virus infection during stem cell transplantation can also lead to BK virus associated nephropathy.

Disclosure: No conflict of interest disclosed.

P446 - Significance of serum galactomannan for diagnosing invasive fungal disease in patients after allogeneic hematopoietic stem cell transplantation

Rieger C.1, Peterson L.1, Lustig D.1, Fiegl M.1, Ostermann H.1

1Ludwig-Maximilians-Universität München, M