Intervirology 2008;51:173–181

Development of Human Single-Chain Antibodies against SARS-Associated Coronavirus

Leung K.M.a, c · Feng D.X.b · Lou J.e · Zhou Y.e · Fung K.P.c · Waye M.M.Y.c · Tsui S.K.W.c · Chan P.K.S.d · Marks J.D.e · Pang S.F.a · Kan Y.W.b
aCK Life Sciences International Inc., Hong Kong, SAR, China; bDepartment of Laboratory Medicine, University of California, San Francisco, Calif., USA; cDepartment of Biochemistry, Croucher Laboratory for Human Genomics, and dDepartment of Microbiology, Chinese University of Hong Kong, Hong Kong, SAR, China; eDepartment of Anesthesia and Pharmaceutical Chemistry, University of California, San Francisco General Hospital, San Francisco, Calif., USA
email Corresponding Author

 goto top of outline Key Words

  • Antibody
  • Severe acute respiratory syndrome
  • Single-chain variable fragment
  • Immunology

 goto top of outline Abstract

The outbreak of severe acute respiratory syndrome (SARS), caused by a distinct coronavirus, in 2003 greatly threatened public health in China, Southeast Asia as well as North America. Over 1,000 patients died of the SARS virus, representing 10% of infected people. Like other coronaviruses, the SARS virus also utilizes a surface glycoprotein, namely the spike protein, to infect host cells. The spike protein of SARS virus consists of 1,255 amino acid residues and can be divided into two sub-domains, S1 and S2. The S1 domain mediates the binding of the virus to its receptor angiotensin-converting enzyme 2, which is abundantly distributed on the surface of human lung cells. The S2 domain mediates membrane fusion between the virus and the host cell. Hence two strategies can be used to block the infection of the SARS virus, either by interfering with the binding of the S1 domain to the receptor or by blocking the fusion of the virus with the cell membrane mediated by the S2 domain. Several antibodies against the S1 domain have been generated and all of them are able to neutralize the virus in vitro and in vivo using animal models. Unfortunately, point mutations have been identified in the S1 domain, so that the virus isolated in the future may not be recognized by these antibodies. As no mutation has been found in the S2 domain indicating that this region is more conserved than the S1 domain, it may be a better target for antibody binding. After predicting the immunogenicity of the epitopes of the S2 domain, we chemically synthesized two peptides and also expressed one of them using a recombinant DNA method. We screened a phage displaying library of human single-chain antibodies (ScFv) against the predicted epitopes and obtained a human ScFv which can recognize the SARS virus in vitro.

Copyright © 2008 S. Karger AG, Basel

 goto top of outline References
  1. Ksiazek TG, Erdman D, Goldsmith CS, Zaki SR, Peret T, Emery S, Tong S, Urbani C, Comer JA, Lim W, Rollin PE, Dowell SF, Ling AE, Humphrey CD, Shieh WJ, Guarner J, Paddock CD, Rota P, Fields B, DeRisi J, Yang JY, Cox N, Hughes JM, LeDuc JW, Bellini WJ, Anderson LJ: A novel coronavirus associated with severe acute respiratory syndrome. N Engl J Med 2003;348:1953–1966.
  2. Kuiken T, Fouchier RA, Schutten M, Rimmelzwaan GF, van Amerongen G, van Riel D, Laman JD, de Jong T, van Doornum G, Lim W, Ling AE, Chan PK, Tam JS, Zambon MC, Gopal R, Drosten C, van der Werf S, Escriou N, Manuguerra JC, Stohr K, Peiris JS, Osterhaus AD: Newly discovered coronavirus as the primary cause of severe acute respiratory syndrome. Lancet 2003;362:263–270.
  3. Drosten C, Preiser W, Günther S, Schmitz H, Doerr HW: Severe acute respiratory syndrome: identification of the etiological agent. Trends Mol Med 2003;9:325–327.
  4. Cheng Y, Wong R, Soo YO, Wong WS, Lee CK, Ng MH, Chan P, Wong KC, Leung CB, Cheng G: Use of convalescent plasma therapy in SARS patients in Hong Kong. Eur J Clin Microbiol Infect Dis 2005;24:44–46.
  5. Marks JD, Hoogenboom HR, Bonnert TP, McCafferty J, Griffiths AD, Winter G: By-passing immunization. Human antibodies from V-gene libraries displayed on phage. J Mol Biol 1991;222:581–597.
  6. Leonard JP, Coleman M, Ketas JC, Chadburn A, Ely S, Furman RR, Wegener WA, Hansen HJ, Ziccardi H, Eschenberg M, Gayko U, Cesano A, Goldenberg DM: Phase I/II trial of epratuzumab (humanized anti-CD22 antibody) in indolent non-Hodgkin’s lymphoma. J Clin Oncol 2003;21:3051–3059.
  7. Hayashi T, Treon SP, Hideshima T, Tai YT, Akiyama M, Richardson P, Chauhan D, Grewal IS, Anderson KC: Recombinant humanized anti-CD40 monoclonal antibody triggers autologous antibody-dependent cell-mediated cytotoxicity against multiple myeloma cells. Br J Haematol 2003;121:592–596.
  8. Juweid M: Technology evaluation: epratuzumab, Immunomedics/Amgen. Curr Opin Mol Ther 2003;5:192–198.
  9. Poul MA, Becerril B, Nielsen UB, Morisson P, Marks JD: Selection of tumor-specific internalizing human antibodies from phage libraries. J Mol Biol 2000;301:1149–1161.
  10. Gallagher TM, Buchmeier MJ: Coronavirus spike proteins in viral entry and pathogenesis. Virology 2001;279:371–374.
  11. Li W, Moore MJ, Vasilieva N, Sui J, Wong SK, Berne MA, Somasundaran M, Sullivan JL, Luzuriaga K, Greenough TC, Choe H, Farzan M: Angiotensin-converting enzyme 2 is a functional receptor for the SARS coronavirus. Nature 2003;426:450–454.
  12. Sui J, Li W, Murakami A, Tamin A, Matthews LJ, Wong SK, Moore MJ, Tallarico AS, Olurinde M, Choe H, Anderson LJ, Bellini WJ, Farzan M, Marasco WA: Potent neutralization of severe acute respiratory syndrome (SARS) coronavirus by a human mAb to S1 protein that blocks receptor association. Proc Natl Acad Sci USA 2004;101:2536–2541.
  13. Rota PA, Oberste MS, Monroe SS, Nix WA, Campagnoli R, Icenogle JP, Penaranda S, Bankamp B, Maher K, Chen MH, Tong S, Tamin A, Lowe L, Frace M, DeRisi JL, Chen Q, Wang D, Erdman DD, Peret TC, Burns C, Ksiazek TG, Rollin PE, Sanchez A, Liffick S, Holloway B, Limor J, McCaustland K, Olsen-Rasmussen M, Fouchier R, Gunther S, Osterhaus AD, Drosten C, Pallansch MA, Anderson LJ, Bellini WJ: Characterization of a novel coronavirus associated with severe acute respiratory syndrome. Science 2003;300:1394–1399.
  14. Castilla J, Sola I, Enjuanes L: Interference of coronavirus infection by expression of immunoglobulin G (IgG) or IgA virus-neutralizing antibodies. J Virol 1997;71:5251–5258.
  15. Huie MA, Cheung MC, Muench MO, Becerril B, Kan YW, Marks JD: Antibodies to human fetal erythroid cells from a nonimmune phage antibody library. Proc Natl Acad Sci USA 2001;98:2682–2687.
  16. Sheets MD, Amersdorfer P, Finnern R, Sargent P, Lindquist E, Schier R, Hemingsen G, Wong C, Gerhart JC, Marks JD: Efficient construction of a large nonimmune phage antibody library: the production of high-affinity human single-chain antibodies to protein antigens. Proc Natl Acad Sci USA 1998;95:6157–6162.
  17. Poul MA, Marks JD: Targeted gene delivery to mammalian cells by filamentous bacteriophage. J Mol Biol 1999;288:203–211.
  18. Jacobsson K, Rosander A, Bjerketorp J, Frykberg L: Shotgun phage display – selection for bacterial receptins or other exported proteins. Biol Proced Online 2003;5:123–135.
  19. Zhang H, Wang G, Li J, Nie Y, Shi X, Lian G, Wang W, Yin X, Zhao Y, Qu X, Ding M, Deng H: Identification of an antigenic determinant on the S2 domain of the severe acute respiratory syndrome coronavirus spike glycoprotein capable of inducing neutralizing antibodies. J Virol 2004;78:6938–6945.

 goto top of outline Author Contacts

K.M. Leung
Room 610, Mong Man Wai Building
Department of Biochemistry, Chinese University of Hong Kong
Hong Kong, SAR (China)
Tel. +852 9420 4960, Fax +852 2651 9324, E-Mail

 goto top of outline Article Information

Received: January 30, 2008
Accepted after revision: May 8, 2008
Published online: August 25, 2008
Number of Print Pages : 9
Number of Figures : 7, Number of Tables : 2, Number of References : 19

 goto top of outline Publication Details

Intervirology (International Journal of Basic and Medical Virology)

Vol. 51, No. 3, Year 2008 (Cover Date: September 2008)

Journal Editor: Liebert U.G. (Leipzig)
ISSN: 0300–5526 (Print), eISSN: 1423–0100 (Online)

For additional information:

Copyright / Drug Dosage / Disclaimer

Copyright: All rights reserved. No part of this publication may be translated into other languages, reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying, recording, microcopying, or by any information storage and retrieval system, without permission in writing from the publisher or, in the case of photocopying, direct payment of a specified fee to the Copyright Clearance Center.
Drug Dosage: The authors and the publisher have exerted every effort to ensure that drug selection and dosage set forth in this text are in accord with current recommendations and practice at the time of publication. However, in view of ongoing research, changes in goverment regulations, and the constant flow of information relating to drug therapy and drug reactions, the reader is urged to check the package insert for each drug for any changes in indications and dosage and for added warnings and precautions. This is particularly important when the recommended agent is a new and/or infrequently employed drug.
Disclaimer: The statements, opinions and data contained in this publication are solely those of the individual authors and contributors and not of the publishers and the editor(s). The appearance of advertisements or/and product references in the publication is not a warranty, endorsement, or approval of the products or services advertised or of their effectiveness, quality or safety. The publisher and the editor(s) disclaim responsibility for any injury to persons or property resulting from any ideas, methods, instructions or products referred to in the content or advertisements.