Synthetic therapeutic antibodies

Abstract

Advances in selection technologies have sped up the process of generating antibodies with exquisitely tailored characteristics. In particular, synthetic antibody libraries, in which the antigen-binding sites are entirely man-made, have come of age and now rival or even exceed the potential of natural immune repertoires. Control over both library design and selection conditions enables unprecedented precision in antibody engineering. Synthetic libraries have been used to gain insights into the mechanisms of antibody structure and function, to tackle particularly difficult therapeutic challenges and to expand the utility of antibodies to novel areas of research.

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Figure 1: Construction of antibody libraries from natural or synthetic diversity.
Figure 2: Sites of diversification in synthetic antibody libraries.
Figure 3: The structural epitopes for VEGFR1 and antibodies binding to human VEGF.
Figure 4: Minimalist antigen-binding sites are dominated by tyrosine.

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References

  1. 1

    Hudson, P.J. Recombinant antibody constructs in cancer therapy. Curr. Opin. Immunol. 11, 548–557 (1999).

    CAS  Article  Google Scholar 

  2. 2

    Schaedel, O. & Reiter, Y. Antibodies and their fragments as anti-cancer agents. Curr. Pharm. Des. 12, 363–378 (2006).

    CAS  Article  Google Scholar 

  3. 3

    Trikha, M., Yan, L. & Nakada, M.T. Monoclonal antibodies as therapeutics in oncology. Curr. Opin. Biotechnol. 13, 609–614 (2002).

    CAS  Article  Google Scholar 

  4. 4

    Bradbury, A. Antibodies in proteomics I: generating antibodies. Trends Biotechnol. 21, 275–281 (2003).

    CAS  Article  Google Scholar 

  5. 5

    Bradbury, A.R.M. & Marks, J.D. Antibodies from phage antibody libraries. J. Immunol. Methods 290, 29–49 (2004).

    CAS  Article  Google Scholar 

  6. 6

    Hoogenboom, H.R. Selecting and screening recombinant antibody libraries. Nat. Biotechnol. 23, 1105–1116 (2005).

    CAS  Article  Google Scholar 

  7. 7

    Brekke, O.H. & Loset, G.A. New technologies in therapeutic antibody development. Curr. Opin. Pharmacol. 3, 544–550 (2003).

    CAS  Article  Google Scholar 

  8. 8

    Fellouse, F.A. & Sidhu, S.S. in Phage Display in Biotechnology and Drug Discovery 1st edn., Vol. 3 (ed. Sidhu, S.S.) 709–740 (Taylor and Francis Group, Boca Raton, Florida, 2005).

    Google Scholar 

  9. 9

    Johnson, G. & Wu, T.T. Kabat database and its applications: 30 years after the first variability plot. Nucleic Acids Res. 28, 214–218 (2000).

    CAS  Article  Google Scholar 

  10. 10

    Padlan, E.A. X-ray crystallography of antibodies. Advan. Protein Chem. 49, 57–133 (1996).

    CAS  Article  Google Scholar 

  11. 11

    Smith, G.P. Filamentous fusion phage: novel expression vectors that display cloned antigens on the virion surface. Science 228, 1315–1317 (1985).

    CAS  Article  Google Scholar 

  12. 12

    Dobson, C.L., Minter, R.R. & Hart-Shorrock, C.P. in Phage Display in Biotechnology and Drug Discovery 1st edn., Vol. 3 (ed. Sidhu, S.S.) 659–708 (Taylor and Francis Group, Boca Raton, Florida, 2005).

    Google Scholar 

  13. 13

    Hoogenboom, H.R. Overview of antibody phage-display technology and its applications. Methods Mol. Biol. 178, 1–37 (2002).

    CAS  PubMed  Google Scholar 

  14. 14

    Rader, C. & Barbas, C.F. Phage display of combinatorial antibody libraries. Curr. Opin. Biotechnol. 8, 503–508 (1997).

    CAS  Article  Google Scholar 

  15. 15

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

    CAS  Article  Google Scholar 

  16. 16

    Georgiou, G. et al. Display of heterologous proteins on the surface of microorganisms: from the screening of combinatorial libraries to live recombinant vaccines. Nat. Biotechnol. 15, 29–34 (1997).

    CAS  Article  Google Scholar 

  17. 17

    Wittrup, K.D. Protein engineering by cell-surface display. Curr. Opin. Biotechnol. 12, 395–399 (2001).

    CAS  Article  Google Scholar 

  18. 18

    Harvey, B.R. et al. Anchored periplasmic expression, a versatile technology for the isolation of high-affinity antibodies from Escherichia coli-expressed libraries. Proc. Natl. Acad. Sci. USA 101, 9193–9198 (2004).

    CAS  Article  Google Scholar 

  19. 19

    Urban, J.H. et al. Selection of functional human antibodies from retroviral display libraries. Nucleic Acids Res. 33, e35 (2005).

    Article  Google Scholar 

  20. 20

    Lipovsek, D. & Pluckthun, A. In vitro protein evolution by ribosome display and mRNA display. J. Immunol. Methods 290, 51–67 (2004).

    CAS  Article  Google Scholar 

  21. 21

    Reiersen, H. et al. Covalent antibody display - an in vitro antibody-DNA library selection system. Nucleic Acids Res. 33, e10 (2005).

    Article  Google Scholar 

  22. 22

    Koch, H., Grafe, N., Schiess, R. & Pluckthun, A. Direct selection of antibodies from complex libraries with the protein fragment complementation assay. J. Mol. Biol. 357, 427–441 (2006).

    CAS  Article  Google Scholar 

  23. 23

    der Maur, A.A. et al. Direct in vivo screening of intrabody libraries constructed on a highly stable single-chain framework. J. Biol. Chem. 277, 45075–45085 (2002).

    Article  Google Scholar 

  24. 24

    Urech, D.M., Lichtlen, P. & Barberis, A. Cell growth selection system to detect extracellular and transmembrane protein interactions. Biochim. Biophys. Acta 1622, 117–127 (2003).

    CAS  Article  Google Scholar 

  25. 25

    Visintin, M., Meli, G.A., Cannistraci, I. & Cattaneo, A. Intracellular antibodies for proteomics. J. Immunol. Methods 290, 135–153 (2004).

    CAS  Article  Google Scholar 

  26. 26

    Sidhu, S.S. et al. Phage-displayed antibody libraries of synthetic heavy chain complementarity determining regions. J. Mol. Biol. 338, 299–310 (2004).

    CAS  Article  Google Scholar 

  27. 27

    Feldhaus, M.J. et al. Flow-cytometric isolation of human antibodies from a nonimmune Saccharomyces cerevisiae surface display library. Nat. Biotechnol. 21, 163–170 (2003).

    CAS  Article  Google Scholar 

  28. 28

    Barbas, C.F. III, Amberg, W., Simoncsits, A., Jones, T.M. & Lerner, R.A. Selection of human anti-hapten antibodies from semisynthetic libraries. Gene 137, 57–62 (1993).

    CAS  Article  Google Scholar 

  29. 29

    Fellouse, F.A. et al. Molecular recognition by a binary code. J. Mol. Biol. 348, 1153–1162 (2005).

    CAS  Article  Google Scholar 

  30. 30

    Fellouse, F.A., Wiesmann, C. & Sidhu, S.S. Synthetic antibodies from a four-amino-acid code: a dominant role for tyrosine in antigen recognition. Proc. Natl. Acad. Sci. USA 101, 12467–12472 (2004).

    CAS  Article  Google Scholar 

  31. 31

    Griffiths, A.D. et al. Isolation of high affinity human antibodies directly from large synthetic repertoires. EMBO J. 13, 3245–3260 (1994).

    CAS  Article  Google Scholar 

  32. 32

    Hoet, R.M. et al. Generation of high-affinity human antibodies by combining donor-derived and synthetic complementarity-determining-region diversity. Nat. Biotechnol. 23, 344–348 (2005).

    CAS  Article  Google Scholar 

  33. 33

    Jespers, L., Schon, O., James, L.C., Veprintsev, D. & Winter, G. Crystal structure of HEL4, a soluble, refoldable human VH single domain with a germ-line scaffold. J. Mol. Biol. 337, 893–903 (2004).

    CAS  Article  Google Scholar 

  34. 34

    Knappik, A. et al. Fully synthetic human combinatorial antibody libraries (HuCAL) based on modular consensus frameworks and CDRs randomized with trinucleotides. J. Mol. Biol. 296, 57–86 (2000).

    CAS  Article  Google Scholar 

  35. 35

    Lee, C.V. et al. High-affinity human antibodies from phage-displayed synthetic Fab libraries with a single framework scaffold. J. Mol. Biol. 340, 1073–1093 (2004).

    CAS  Article  Google Scholar 

  36. 36

    Persson, H., Lantto, J. & Ohlin, M. A focused antibody library for improved hapten recognition. J. Mol. Biol. 357, 607–620 (2006).

    CAS  Article  Google Scholar 

  37. 37

    Reiter, Y., Schuck, P., Boyd, L.F. & Plaksin, D. An antibody single-domain phage display library of a native heavy chain variable region: isolation of functional single-domain VH molecules with a unique interface. J. Mol. Biol. 290, 685–698 (1999).

    CAS  Article  Google Scholar 

  38. 38

    Silacci, M. et al. Design, construction, and characterization of a large synthetic human antibody phage display library. Proteomics 5, 2340–2350 (2005).

    CAS  Article  Google Scholar 

  39. 39

    van Wyngaardt, W. et al. A large semi-synthetic single-chain Fv phage display library based on chicken immunoglobulin genes. BMC Biotechnol. 4, 6 (2004).

    Article  Google Scholar 

  40. 40

    Barbas, C.F. III, Bain, J.D., Hoekstra, D.M. & Lerner, R.A. Semisynthetic combinatorial antibody libraries: a chemical solution to the diversity problem. Proc. Natl. Acad. Sci. USA 89, 4457–4461 (1992).

    CAS  Article  Google Scholar 

  41. 41

    Hoogenboom, H.R. & Winter, G. By-passing immunization: human antibodies from synthetic repertoires of germline VH gene segments rearranged in vitro. J. Mol. Biol. 227, 381–388 (1992).

    CAS  Article  Google Scholar 

  42. 42

    Lee, C.V. et al. Synthetic anti-BR3 antibodies that mimic BAFF binding and target both human and murine B cells. Nat. Biotechnol. (in the press).

  43. 43

    Liang, W.-C. et al. Cross-species vascular endothelial growth factor (VEGF)-blocking antibodies completely inhibit the growth of human tumor xenografts and measure the contribution of stromal VEGF. J. Biol. Chem. 281, 951–961 (2006).

    CAS  Article  Google Scholar 

  44. 44

    Fuh, G. et al. Structure-function studies of two synthetic anti-vascular endothelial growth factor Fabs and comparison with the AvastinTM Fab. J. Biol. Chem. 281, 6625–6631 (2006).

    CAS  Article  Google Scholar 

  45. 45

    Wiesmann, C. et al. Crystal structure at 1.7 Å resolution of VEGF in complex with domain 2 of the Flt-1 receptor. Cell 91, 695–704 (1997).

    CAS  Article  Google Scholar 

  46. 46

    Muller, Y.A. et al. VEGF and the Fab fragment of a humanized neutralizing antibody: crystal structure of the complex at 2.4 Å resolution and mutational analysis of the interface. Structure 6, 1153–1167 (1998).

    CAS  Article  Google Scholar 

  47. 47

    Desiderio, A. et al. A semi-synthetic repertoire of intrinsically stable antibody fragments derived from a single-framework scaffold. J. Mol. Biol. 310, 603–615 (2001).

    CAS  Article  Google Scholar 

  48. 48

    MacCallum, R.M., Martin, A.C. & Thornton, J.M. Antibody-antigen interactions: contact analysis and binding site topography. J. Mol. Biol. 262, 732–745 (1996).

    CAS  Article  Google Scholar 

  49. 49

    Almagro, J.C. Identification of differences in the specificity-determining residues of antibodies that recognize antigens of different size: implications for the rational design of antibody repertoires. J. Mol. Recognit. 17, 132–143 (2004).

    CAS  Article  Google Scholar 

  50. 50

    Collis, A.V., Brouwer, A.P. & Martin, A.C. Analysis of the antigen combining site: correlations between length and sequence composition of the hypervariable loops and the nature of the antigen. J. Mol. Biol. 325, 337–354 (2003).

    CAS  Article  Google Scholar 

  51. 51

    Hamers-Casterman, C. et al. Naturally occurring antibodies devoid of light chains. Nature 363, 446–448 (1993).

    CAS  Article  Google Scholar 

  52. 52

    Muyldermans, S., Atarhouch, T., Saldanha, J., Barbosa, J.A.R.G. & Hamers, R. Sequence and structure of VH domain from naturally occurring camel heavy chain immunoglobulins lacking light chains. Protein Eng. 7, 1129–1135 (1994).

    CAS  Article  Google Scholar 

  53. 53

    Riechmann, L. Rearrangement of the former VL interface in the solution structure of a camelised, single antibody VH domain. J. Mol. Biol. 259, 957–969 (1996).

    CAS  Article  Google Scholar 

  54. 54

    Davies, D.R. & Cohen, G.H. Interactions of protein antigens with antibodies. Proc. Natl. Acad. Sci. USA 93, 7–12 (1996).

    CAS  Article  Google Scholar 

  55. 55

    Padlan, E.A. Anatomy of the antibody molecule. Mol. Immunol. 31, 169–217 (1994).

    CAS  Article  Google Scholar 

  56. 56

    Fellouse, F.A., Barthelemy, P.A., Kelley, R.F. & Sidhu, S.S. Tyrosine plays a dominant functional role in the paratope of a synthetic antibody derived from a four amino acid code. J. Mol. Biol. 357, 100–114 (2006).

    CAS  Article  Google Scholar 

  57. 57

    Li, B. et al. Activation of the proapoptotic death receptor DR5 by oligomeric peptide and antibody agonists. J. Mol. Biol. 361, 522–536 (2006).

    CAS  Article  Google Scholar 

  58. 58

    Sanz, L., Blanco, B. & Alvarez-Vallina, L. Antibodies and gene therapy: teaching old 'magic bullets' new tricks. Trends Immunol. 25, 85–91 (2004).

    CAS  Article  Google Scholar 

  59. 59

    Bradbury, A. et al. Antibodies in proteomics II: screening, high-throughput characterization and downstream applications. Trends Biotechnol. 21, 312–317 (2003).

    CAS  Article  Google Scholar 

  60. 60

    Chambers, R.S. High-throughput antibody production. Curr. Opin. Chem. Biol. 9, 46–50 (2005).

    CAS  Article  Google Scholar 

  61. 61

    Jung, D., Giallourakis, C., Mostoslavsky, R. & Alt, F.W. Mechanism and control of V(D)J recombination at the immunoglobulin heavy chain locus. Annu. Rev. Immunol. 24, 541–570 (2006).

    CAS  Article  Google Scholar 

  62. 62

    Kabat, E.A., Wu, T.T., Redi-Miller, M., Perry, H.M. & Gottesman, K.S. Sequences of Proteins of Immunological Interest 4th edn. (US National Institutes of Health, Bethesda, Maryland, 1987).

    Google Scholar 

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Correspondence to Sachdev S Sidhu.

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The authors are employed by Genentech, Inc., which develops antibody therapeutics for the health care market.

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Supplementary Table 1

Highly functional synthetic antibody libraries. (PDF 20 kb)

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Sidhu, S., Fellouse, F. Synthetic therapeutic antibodies. Nat Chem Biol 2, 682–688 (2006). https://doi.org/10.1038/nchembio843

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