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A recombinant, fully human monoclonal antibody with antitumor activity constructed from phage-displayed antibody fragments

Abstract

A single-chain Fv antibody fragment specific for the tumor-associated Ep-CAM molecule was isolated from a semisynthetic phage display library and converted into an intact, fully human IgG1 monoclonal antibody (huMab). The purified huMab had an affinity of 5 nM and effectively mediated tumor cell killing in in vitro and in vivo assays. These experiments show that nonimmunized phage antibody display libraries can be used to obtain high-affinity, functional, and clinically applicable huMabs directed against a tumor-associated antigen.

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Figure 1: Characterization of scFv UBS-54.
Figure 2: Characterization of huMab UBS-54.
Figure 3: Sensorgram of huMab UBS-54 and murine Mabs 17-1A and 323/A3.
Figure 4: Antibody-dependent cellular cytotoxicity using huMab UBS-54 and control murine Mabs.
Figure 5: Mean tumor size in nude mice xenografted with the colon carcinoma cell line LS180 and treated with PBS (), UBS-54 (□) 17-1A (), or 323/A3 (▪).

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References

  1. Shawler, D.L., Bartholomew, R.M., Smith, L.M. & Dillman, R.O. Human immune response to multiple injections of murine monoclonal IgG. J. Immunol. 135, 1530–1535 (1985).

    CAS  PubMed  Google Scholar 

  2. Miller, R.A., Oseroff, A.R., Stratte, P.T. & Levy, R. Monoclonal antibody therapeutic trials in seven patients with T-cell lymphoma. Blood 62, 988–995 ( 1983).

    CAS  PubMed  Google Scholar 

  3. Hakimi, J. et al. Reduced immunogenicity and improved pharmacokinetics of humanized anti-Tac in cynomolgus monkeys. J. Immunol. 147, 1352–1359 (1991).

    CAS  PubMed  Google Scholar 

  4. Stephens, S. et al. Comprehensive pharmacokinetics of a humanized antibody and analysis of residual anti-idiotypic responses. Immunology 85 , 668–674 (1995).

    CAS  PubMed  PubMed Central  Google Scholar 

  5. Riechmann, L., Clark, M., Waldmann, H. & Winter, G. Reshaping human antibodies for therapy. Nature 332, 323– 327 (1988).

    Article  CAS  PubMed  Google Scholar 

  6. Junghans, R.P. et al. Anti-Tac-H, a humanized antibody to the interleukin 2 receptor with new features for immunotherapy in malignant and immune disorders. Cancer Res. 50, 1495–1502 (1990).

    CAS  PubMed  Google Scholar 

  7. Vaughan, T.J., Osbourn, J.K. & Tempest, P.R. Human antibodies by design. Nat. Biotechnol . 16, 535–539 ( 1998).

    Article  CAS  PubMed  Google Scholar 

  8. Khazaeli, M.B. et al. Pharmacokinetics and immune response of 131I-chimeric mouse/human B72.3 (human gamma4) monoclonal antibody in humans. Cancer Res . 51, 5461–5466 ( 1989).

    Google Scholar 

  9. Elliot, M.J. et al. Repeated therapy with monoclonal antibody to tumor necrosis factor alfa (cA2) in patients with rheumatoid arthritis. Lancet 344, 1125–1127 (1994).

    Article  Google Scholar 

  10. Jones, P.T., Dear, P.H., Foote, J., Neuberger, M.S. & Winter, G. Replacing the complementarity-determining regions in a human antibody with those from a mouse. Nature 321 , 522–525 (1986).

    Article  CAS  PubMed  Google Scholar 

  11. Foote, J. & Winter, G. Antibody framework residues affecting the conformation of the hypervariable loops. J. Mol. Biol. 224, 487–499 (1992).

    Article  CAS  PubMed  Google Scholar 

  12. McLaughlin, P. et al. Rituximab chimeric anti-CD20 monoclonal antibody therapy for relapsed indolent lymphoma: half of patients respond to a four-dose treatment program. J. Clin. Oncol. 16, 2825– 2833 (1998).

    Article  CAS  PubMed  Google Scholar 

  13. Pegram, M.D. et al. Phase II study of receptor-enhanced chemosensitivity using recombinant humanized anti-p185 Her2/neu monoclonal antibody plus cisplatin in patients with Her2/neu-overexpressing metastatic breast cancer refractory to chemotherapy treatment. J. Clin. Oncol. 16, 2659– 2671 (1998).

    Article  CAS  PubMed  Google Scholar 

  14. Vincenti, F. et al. Interleukin-2-receptor blockade with daclizumab to prevent acute rejection in renal transplantation. N. Engl. J. Med. 338, 161–165 (1998).

    Article  CAS  PubMed  Google Scholar 

  15. Bruggeman, M. & Neuberger, M.S. Strategies for expressing human antibody repertoires in transgenic mice. Immunol. Today 17, 391–397 (1996).

    Article  Google Scholar 

  16. Mendez, M.J. et al. Functional transplant of megabase human immunoglobulin loci recapitulates human antibody response in mice. Nat. Genet. 15, 146–156 (1997).

    Article  CAS  PubMed  Google Scholar 

  17. Winter G., Griffiths, A.D., Hawkins, R.E. & Hoogenboom, H.R. Making antibodies by phage display technology. Annu. Rev. Immunol . 12, 433–455 ( 1994).

    Article  CAS  PubMed  Google Scholar 

  18. Burton, D.R. & Barbas III, C.F. Human antibodies from combinatorial libraries. Adv. Immunol. 57, 191–280 (1994).

    Article  CAS  PubMed  Google Scholar 

  19. De Kruif, J., Terstappen, L., Boel, E. & Logtenberg, T. Rapid selection of cell subpopulation-specific human monoclonal antibodies from a synthetic phage antibody library. Proc. Natl. Acad. Sci. USA 92, 3938–3942 (1995).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Riethmuller, G. et al. 1994. Randomised trial of monoclonal antibody for adjuvant therapy of resected Dukes = C colorectal carcinoma. Lancet 343, 1177–1183 (1994).

    Article  CAS  PubMed  Google Scholar 

  21. De Kruif, J., Boel, E.Z. & Logtenberg, T. Selection and application of human single chain Fv antibody fragments from a semi-synthetic phage antibody display library with designed CDR3 regions. J. Mol. Biol. 248, 97– 105 (1995).

    Article  CAS  PubMed  Google Scholar 

  22. Gottlinger, H.G., Funke, I., Johnson, J., Gokel, J.M. & Riethmuller, G. The epithelial cell surface antigen CO17-1A, a target for antibody-mediated tumor therapy: its biochemical nature, tissue distribution and recognition by different monoclonal antibodies. Int. J. Cancer 38, 47–53 ( 1986).

    Article  CAS  PubMed  Google Scholar 

  23. Momberg, F., Moldenhauer, G., Hammerling, G.J. & Moller, P. Immunohistochemical study of the expression of a Mr 34,000 human epithelium-specific surface glycoprotein in normal and malignant tissues. Cancer Res . 47, 2883–2891 ( 1987).

    Google Scholar 

  24. Thampoe, I.J., Ng, J.S.C. & Lloyd, K.O. Biochemical analysis of a human epithelial surface antigen: differential cell expression and processing. Arch. Biochem. Biophys . 267, 342–352 ( 1988).

    Article  CAS  PubMed  Google Scholar 

  25. Simon, B. et al. Epithelial glycoprotein is a member of a family of epithelial surface antigens homologous to nidogen, a matrix adhesion protein. Proc. Natl. Acad. Sci. USA 87, 2755–2759 ( 1990).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Litvinov, S.V., Velders, M.P., Bakker, H.A.M., Fleuren, G.J. & Warnaar, S.O. Ep-CAM: a human epithelial antigen is a homophilic cell-cell adhesion molecule. J. Cell Biol. 125, 437–446 (1994).

    Article  CAS  PubMed  Google Scholar 

  27. Steplewski, Z. et al. Biological activity of human-mouse IgG1, IgG2, IgG3 and IgG4 chimeric monoclonal antibodies with antitumor specificity. Proc. Natl. Acad. Sci. USA 85, 4852–4856 ( 1988).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Pak, K.Y. et al. 1991. Evaluation of the 323/A3 monoclonal antibody and the use of technetium-99m-labeled 323/A3 Fab' for the detection of panadenocarcinoma. Int. J. Rad. Appl. Instrum. 18, 483–497 (1991).

    Article  CAS  Google Scholar 

  29. Velders, M.P. et al. The role of antibody affinity in tumor immunotherapy evaluated in in-vivo models for minimal residual disease. J. Immunother. 19, 245–256 (1996).

    Article  CAS  Google Scholar 

  30. Heijnen, I.A.F.M. & van de Winkel, J.G.J. Human IgG Fc receptors. Int. Rev. Immunol. 16, 29–56 ( 1997).

    Article  CAS  PubMed  Google Scholar 

  31. Valerius, T. et al. Involvement of the high affinity receptor for IgG (FcgammaRI:CD64) in enhanced tumor cell cytotoxicity of neutrophils during G-CSF therapy. Blood 82, 931–939 ( 1993).

    CAS  PubMed  Google Scholar 

  32. Naramura, M., Gillies, S.D., Mendelsohn, J., Reisfeld, R.A. & Mueller, B.M. Therapeutic potential of chimeric and murine anti-EGF receptor antibodies in a metastasis model for human melanoma. Cancer Immunol. Immunother. 37, 343– 349 (1993).

    Article  CAS  PubMed  Google Scholar 

  33. Nishimura, Y. et al. Recombinant human-mouse chimeric monoclonal antibody specific for common acute lymphocytic leukemia antigen. Cancer Res. 47, 999–1005 (1987).

    CAS  PubMed  Google Scholar 

  34. Liu, A.Y. et al. Chimeric mouse-human IgG1 antibody that can mediate lysis of cancer cells. Proc. Natl. Acad. Sci. USA. 84, 3439– 3443 (1987).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Takahashi, H., Nakada, T., Nakaki, M. & Wands, J.R. Inhibition of hepatic metastases of human colon cancer in nude mice by a chimeric SF-25 monoclonal antibody. Gastroenterology. 108, 172–182 (1995).

    Article  CAS  PubMed  Google Scholar 

  36. Walker, M.R., Lund, J., Thompson, K.M. & Jefferis, R. Aglycosylation of human IgG1 and IgG3 monoclonal antibodies can eliminate recognition by human cells expressing Fc gamma RI and/or Fc gamma RII receptors. Biochem. J. 259, 347–353 (1989).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. LoBuglio, A.F. et al. Mouse/human chimeric monoclonal antibody in man: kinetics and immune response. Proc. Natl. Acad. Sci. USA 86, 4220–4224 (1989).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Balzar, M. et al. Cytoplasmic tail regulates the intercellular adhesion function of the epithelial cell adhesion molecule. Mol. Cell. Biol. 18, 4833–4843 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. De Leij, L., Helfrich, W., Stein, R. & Mattes, M.J. SCLC Cluster 2 antibodies detect the pancarcinoma/epithelial glycoprotein EGP-2. Int. J. Cancer suppl. 8, 60–63 (1994).

    Article  CAS  PubMed  Google Scholar 

  40. Meerman, H.J. & Georgiou, G. Construction and characterization of a set of E.coli strains deficient in all known loci affecting the proteolytic stability of secreted recombinant proteins. Bio/Technology 12, 1107–1109 ( 1994).

    Article  CAS  Google Scholar 

  41. De Kruif, J., Storm, G., van Bloois, L. & Logtenberg, T. Biosynthetically lipid-modified human scFv fragments from phage display libraries as targeting molecules for immunoliposomes. FEBS Lett . 199, 232–236 ( 1996).

    Article  Google Scholar 

  42. Hale, J.E. Irreversible, oriented immobilization of antibodies to cobalt-iminodiacetate resin for use as immunoaffinity media. Anal. Biochem. 231, 46–49 (1995).

    Article  CAS  PubMed  Google Scholar 

  43. Lankhof, H. PhD thesis. University of Utrecht. The Netherlands ( 1996).

  44. Karlsson, R., Michaelson, A. & Mattson, L. Kinetic analysis of monoclonal antibody-antigen interactions with a new biosensor based analytical system. J. Immunol. Methods . 198, 129–137 ( 1991).

    Google Scholar 

  45. Van Strijp, J.A.G., van Kessel, K.P.M., van der Tol, M.E. & Verhoef, J. Complement-mediated phagocytosis of herpes simplex virus by human granulocytes: binding or ingestion. J. Clin. Invest. 84, 107–112 (1989).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

The authors would like to thank T. Furebring and A. Marguart for advice on affinity measurements on the BIAcore and N. Barker for critically reading the manuscript. I.A.F.M. Heijnen is supported by grant 13104-CT97-2216 from the European Community.

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Correspondence to Ton Logtenberg.

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Huls, G., Heijnen, I., Cuomo, M. et al. A recombinant, fully human monoclonal antibody with antitumor activity constructed from phage-displayed antibody fragments. Nat Biotechnol 17, 276–281 (1999). https://doi.org/10.1038/7023

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