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Posted: 16 Jul 2011
30 years in 30 weeks, 1991

Recognition of the challenges in the development of an HIV vaccine led to studies that aimed to characterize and understand human immune responses during natural infection, as well as viral ability to evade them. In their commentary, Drs. Burton, Barbas and Lerner describe one of the first large-scale approaches to isolate anti-HIV antibodies. This work led to the appreciation of viral ability to misdirect immune responses away from the critical parts of its envelope. In parallel, Dr. McMichael describes the discovery that HIV is able to escape from cell-based immunity.

A large array of human monoclonal antibodies to type 1 human immunodeficiency virus from combinatorial libraries of asymptomatic seropositive individuals.
Proc Natl Acad Sci U S A.
1991 Nov 15;88(22):10134-7.
Burton DR, Barbas CF 3rd, Persson MA, Koenig S, Chanock RM, Lerner RA.

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Human immunodeficiency virus genetic variation that can escape cytotoxic T cell recognition.
Nature. 1991 Dec 12;354(6353):453-9.
Phillips RE, Rowland-Jones S, Nixon DF, Gotch FM, Edwards JP, Ogunlesi AO, Elvin JG, Rothbard JA, Bangham CR, Rizza CR, & Andrew J. McMichael

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Read the commentary by Drs. Burton, Barbas and Lerner

Read the commentary by Dr. McMichael

View the rest of the series:
1990 < All years > 1992

Commentaries by Dr. Burton and Dr. McMichael

Drs. Burton, Barbas and Lerner

At the end of the eighties, mouse monoclonal antibodies were one of the most valuable tools in biology thanks to the breakthrough of Köhler and Milstein in 1975 (1) but generating human monoclonal antibodies was proving far more problematic. Then came antibody libraries and the possibility of generating antibodies from any species using molecular biological approaches (2, 3). Just two months before publishing this study, we had published the first report describing the creation and selection of a human antibody library using a molecular biological approach called phage display (4). With our successful development of antibody phage display we were anxious to attempt to tackle the important problem of HIV using this new technology. Our hope at the time was to assess the utility of protective epitopes in the context of a natural infection. In order to do this we needed to be able to clone and study human anti-HIV antibodies produced in infected patients. We imagined the returns would be two-fold. Firstly, a collection of potent neutralizing monoclonal antibodies might be used to treat infected individuals. Secondly, if we could work out the epitopes on HIV that these antibodies bound to, we might be able to create a vaccine that could elicit antibodies of this type and prevent infection or act therapeutically.

We set about applying phage technology to HIV and it was an immediate success. Just two weeks after starting the project we had amassed the largest collection of human anti-HIV antibodies known at the time. Twenty years later and the memories of this exciting result are still vivid. We thought we were on the doorstep to an HIV therapy and vaccine.  Like everyone in HIV research, we quickly learned how difficult these goals would be to achieve.  We had at hand a very large array of anti-HIV antibodies. From these many antibodies we identified only one broadly neutralizing antibody known as b12 (5, 6). We had expected to find many more very quickly. What we learned is that the natural immune response to HIV-1 infection elicits many antibodies but only a small portion of these antibodies actually impact the virus through neutralization. Most elicited antibodies recognize epitopes of the envelope not found on the viral particle, i.e. HIV virus could outsmart natural immunity by allowing the generation of a robust response against viral debris. These antibodies do not neutralize virus. Following the description of b12, Hermann Katinger in Vienna described the isolation of two more broadly neutralizing antibodies and these formed the backbone of much HIV antibody research for many years. In fact, it would be more than 15 years before a new generation of broadly neutralizing antibodies could be isolated particularly using single B cell approaches (7-11). These antibodies, isolated from several different individuals and against a number of epitopes, offer new promise to the HIV vaccine field, especially with new insights in to immunogen design and immunization strategies on the horizon.

  1. Kohler, G., and C. Milstein. 1975. Continuous cultures of fused cells secreting antibody of predefined specificity. Nature 256:495-7.
  2. Huse, W. D., L. Sastry, S. A. Iverson, A. S. Kang, M. Alting-Mees, D. R. Burton, S. J. Benkovic, and R. A. Lerner. 1989. Generation of a large combinatorial library of the immunoglobulin repertoire in phage lambda. Science 246:1275-1281.
  3. McCafferty, J., A. D. Griffiths, G. Winter, and D. J. Chiswell. 1990. Phage antibodies: filamentous phage displaying antibody variable domains. Nature 348:552-4.
  4. Barbas, C. F., 3rd, A. S. Kang, R. A. Lerner, and S. J. Benkovic. 1991. Assembly of combinatorial antibody libraries on phage surfaces: the gene III site. Proc. Natl. Acad. Sci. U S A 88:7978-7982.
  5. Barbas, C. F., 3rd, E. Bjorling, F. Chiodi, N. Dunlop, D. Cababa, T. M. Jones, S. L. Zebedee, M. A. Persson, P. L. Nara, E. Norrby, and D. R. Burton. 1992. Recombinant human Fab fragments neutralize human type 1 immunodeficiency virus in vitro. Proc. Natl. Acad. Sci. U S A 89:9339-9343.
  6. Burton, D. R., J. Pyati, R. Koduri, S. J. Sharp, G. B. Thornton, P. W. Parren, L. S. Sawyer, R. M. Hendry, N. Dunlop, P. L. Nara, and et al. 1994. Efficient neutralization of primary isolates of HIV-1 by a recombinant human monoclonal antibody. Science 266:1024-1027.
  7. Walker, L. M., S. K. Phogat, P. Y. Chan-Hui, D. Wagner, P. Phung, J. L. Goss, T. Wrin, M. D. Simek, S. Fling, J. L. Mitcham, J. K. Lehrman, F. H. Priddy, O. A. Olsen, S. M. Frey, P. W. Hammond, G. P. I. Protocol, G. Miiro, J. Serwanga, A. Pozniak, D. McPhee, O. Manigart, L. Mwananyanda, E. Karita, A. Inwoley, W. Jaoko, J. Dehovitz, L. G. Bekker, P. Pitisuttithum, R. Paris, S. Allen, S. Kaminsky, T. Zamb, M. Moyle, W. C. Koff, P. Poignard, and D. R. Burton. 2009. Broad and potent neutralizing antibodies from an African donor reveal a new HIV-1 vaccine target. Science 326:285-9.
  8. Corti, D., J. P. Langedijk, A. Hinz, M. S. Seaman, F. Vanzetta, B. M. Fernandez-Rodriguez, C. Silacci, D. Pinna, D. Jarrossay, S. Balla-Jhagjhoorsingh, B. Willems, M. J. Zekveld, H. Dreja, E. O'Sullivan, C. Pade, C. Orkin, S. A. Jeffs, D. C. Montefiori, D. Davis, W. Weissenhorn, A. McKnight, J. L. Heeney, F. Sallusto, Q. J. Sattentau, R. A. Weiss, and A. Lanzavecchia. 2010. Analysis of memory B cell responses and isolation of novel monoclonal antibodies with neutralizing breadth from HIV-1-infected individuals. PLoS One 5:e8805.
  9. Wu, X., Z. Y. Yang, Y. Li, C. M. Hogerkorp, W. R. Schief, M. S. Seaman, T. Zhou, S. D. Schmidt, L. Wu, L. Xu, N. S. Longo, K. McKee, S. O’Dell, M. K. Louder, D. L. Wycuff, Y. Feng, M. Nason, N. Doria-Rose, M. Connors, P. D. Kwong, M. Roederer, R. T. Wyatt, G. J. Nabel, and J. R. Mascola. 2010. Rational design of envelope surface identifies broadly neutralizing human monoclonal antibodies to HIV-1. Science 329:856-61.
  10. Scheid, J. F., H. Mouquet, B. Ueberheide, R. Diskin, F. Klein, T. Y. K. Olivera, J. Pietzsch, D. Fenyo, A. Abadir, K. Velinzon, A. Hurley, S. Myung, F. Boulad, P. Poignard, D. R. Burton, F. Pereyra, D. D. Ho, B. D. Walker, M. S. Seaman, P. J. Bjorkman, B. T. Chait, and M. C. Nussenzweig. 2011. Sequence and structural convergence of broad and potent HIV antibodies that mimic CD4 binding. Science:In Press.
  11. Walker, L. M., M. Huber, K. J. Doores, E. Falkowska, R. Pejchal, J. P. Julien, S. K. Wang, A. Ramos, P. Y. C. Hui, J. L. Mitcham, P. H. Hammond, O. A. Olsen, P. Phung, S. Fling, C. H. Wong, S. Phogat, T. Wrin, M. D. Simek, P. G. P. Investigators, W. C. Koff, I. A. Wilson, D. R. Burton, and P. Poignard. 2011. Redefining potency and breadth in antibody neutralization of HIV. Nature:Accepted.

About the authors: Dr. Burton is a professor of immunology and microbial science at The Scripps Research Institute (TSRI) in La Jolla. He also directs the International AIDS Vaccine Initiative Neutralizing Antibody Consortium.



My group started work on HIV-1 in 1986 just as my colleague Alain Townsend showed that CD8+ T cells recognize peptides bound to MHC molecules (1). We identified the first HIV peptide epitope KRWIILGLNK (KK10) (2), shortly after Bruce Walker and Fernando Plata had demonstrated the presence of HIV specific CTL in chronically infected patients for the first time (3, 4). Also in 1987 the crystal structure of HLA-2 was solved by Pam Bkorkman, Jack Strominger and Don Wiley (5). They demonstrated exactly where the epitope peptides bound.  Soon after that, they solved the structure of HLA B27 (6), while Rammensee (7) showed that most epitopes were 8-11 amino acids in length and that residues at key positions  (anchors) usually position 2 in the peptide and always the carboxy terminus were specific for particular HLA types. HLA B27 showed a near absolute requirement for arginine at position 2, leading us to speculate that if the virus mutated at this position it might escape T cell recognition. We discussed this with Simon Wain-Hobson who visited us around that time and he sequenced HIV in patients around that epitope region but did not find significant variation at this position (8).

As more and more epitopes were being defined, particularly in HIV-1 infected patients, Sarah Rowland-Jones, Douglas Nixon and I, who were working on these T cell responses, discussed with Rodney Phillips the possibility that some of the HIV-1 variants being described might evade recognition of T cells. Rodney was working in the laboratory of John Bell and Charles Bangham and had set up HIV-1 sequencing using the then-new PCR technology. We decided to collaborate looking at the interplay between T cell responses and virus sequence in some of our most-studied patients. The findings were striking. Looking at different time points in chronic infection, we found three patients with HLA B8 whose CD8+ T cells responded to three epitopes, but we found that the responses changed over time, often being lost as the infection progressed. These changes matched up to sequence changes in the peptide sequence in provirus from activated T cells of patients. The data implied that the virus was actively escaping from the CTL.

Reaction to the paper in Nature was rather disbelieving and in some cases quite hostile. We had not provided enough evidence for active selection to satisfy evolutionary biologists,  and while some accepted our findings, it was thought that such escape might be quite rare. Some pointed out that in the only precedent in the literature, as study of LCMV escape in a T cell receptor transgenic mouse by Zinkernagel’s group, the T cell responses were much larger and more homogeneous that in HIV infection (9). My own rather simplistic view was that if T cells were as important in controlling HIV-1 as many thought, they must select escape mutations given the intrinsic variability in HIV-1. Why some were slow to accept this is puzzling and we were seen as pushing a somewhat heterodox view, even to the extent of being featured in a Science news article (10).

It took some years of longitudinal studies to show unequivocally that selection of escape mutations was occurring. Three papers showed unequivocal CTL escape in humans, following T cell responses and virus sequence changes longitudinally and measuring synonymous/non-synonymous change ratios (11-13). Then in 2002, a paper in the macaque SIV model by Watkins and colleagues showed that escape was normal during acute SIV infection (14) and Moore et al showed that HLA puts altered sequence footprints on the virus (15). After that the idea gained rapid acceptance culminating in studies showing very rapid virus escape in acute infection and the suggestion that escape occurs at virtually all epitopes eventually (16). Indeed 454 sequencing shows the massive struggle that occurs between the antiviral T cells and the varying virus during ongoing infection (17). Escape is part of life for HIV-1.

  1. Townsend AR, Rothbard J, Gotch FM, Bahadur G, Wraith D, McMichael AJ. The epitopes of influenza nucleoprotein recognized by cytotoxic T lymphocytes can be defined with short synthetic peptides. Cell. 1986;44(6):959-68.
  2. Nixon DF, Townsend AR, Elvin JG, Rizza CR, Gallwey J, McMichael AJ. HIV-1 gag-specific cytotoxic T lymphocytes defined with recombinant vaccinia virus and synthetic peptides. Nature. 1988;336(6198):484-7.
  3. Plata F, Autran B, Martins LP, Wain-Hobson S, Raphael M, Mayaud C, Denis M, Guillon JM, Debre P. AIDS virus-specific cytotoxic T lymphocytes in lung disorders. Nature. 1987;328(6128):348-51.
  4. Walker BD, Chakrabarti S, Moss B, Paradis TJ, Flynn T, Durno AG, Blumberg RS, Kaplan JC, Hirsch MS, Schooley RT. HIV-specific cytotoxic T lymphocytes in seropositive individuals. Nature. 1987;328(6128):345-8.
  5. Bjorkman PJ, Saper MA, Samraoui B, Bennett WS, Strominger JL, Wiley DC. Structure of the human class I histocompatibility antigen, HLA-A2. Nature. 1987;329(6139):506-12.
  6. Madden DR, Gorga JC, Strominger JL, Wiley DC. The structure of HLA-B27 reveals nonamer self-peptides bound in an extended conformation. Nature. 1991;353(6342):321-5.
  7. Falk K, Rotzschke O, Deres K, Metzger J, Jung G, Rammensee HG. Identification of naturally processed viral nonapeptides allows their quantification in infected cells and suggests an allele-specific T cell epitope forecast. J Exp Med. 1991;174(2):425-34. PMCID: 2118916.
  8. Meyerhans A, Dadaglio G, Vartanian JP, Langlade-Demoyen P, Frank R, Asjo B, Plata F, Wain-Hobson S. In vivo persistence of a HIV-1-encoded HLA-B27-restricted cytotoxic T lymphocyte epitope despite specific in vitro reactivity. Eur J Immunol. 1991;21(10):2637-40.
  9. Pircher H, Moskophidis D, Rohrer U, Burki K, Hengartner H, Zinkernagel RM. Viral escape by selection of cytotoxic T cell-resistant virus variants in vivo. Nature. 1990;346(6285):629-33.
  10. Balter M. Modest Briton stirs up storm with views on role of CTLs. Science. 1998;280(5371):1860-1.
  11. Borrow P, Lewicki H, Wei X, Horwitz MS, Peffer N, Meyers H, Nelson JA, Gairin JE, Hahn BH, Oldstone MB, Shaw GM. Antiviral pressure exerted by HIV-1-specific cytotoxic T lymphocytes (CTLs) during primary infection demonstrated by rapid selection of CTL escape virus. Nat Med. 1997;3(2):205-11.
  12. Goulder PJ, Phillips RE, Colbert RA, McAdam S, Ogg G, Nowak MA, Giangrande P, Luzzi G, Morgan B, Edwards A, McMichael AJ, Rowland-Jones S. Late escape from an immunodominant cytotoxic T-lymphocyte response associated with progression to AIDS. Nat Med. 1997;3(2):212-7.
  13. Price DA, Goulder PJ, Klenerman P, Sewell AK, Easterbrook PJ, Troop M, Bangham CR, Phillips RE. Positive selection of HIV-1 cytotoxic T lymphocyte escape variants during primary infection. Proc Natl Acad Sci U S A. 1997;94(5):1890-5. PMCID: 20013.
  14. O'Connor DH, Allen TM, Vogel TU, Jing P, DeSouza IP, Dodds E, Dunphy EJ, Melsaether C, Mothe B, Yamamoto H, Horton H, Wilson N, Hughes AL, Watkins DI. Acute phase cytotoxic T lymphocyte escape is a hallmark of simian immunodeficiency virus infection. Nat Med. 2002;8(5):493-9.
  15. Moore CB, John M, James IR, Christiansen FT, Witt CS, Mallal SA. Evidence of HIV-1 adaptation to HLA-restricted immune responses at a population level. Science. 2002;296(5572):1439-43.
  16. Goonetilleke N, Liu MK, Salazar-Gonzalez JF, Ferrari G, Giorgi E, Ganusov VV, Keele BF, Learn GH, Turnbull EL, Salazar MG, Weinhold KJ, Moore S, Letvin N, Haynes BF, Cohen MS, Hraber P, Bhattacharya T, Borrow P, Perelson AS, Hahn BH, Shaw GM, Korber BT, McMichael AJ. The first T cell response to transmitted/founder virus contributes to the control of acute viremia in HIV-1 infection. J Exp Med. 2009;206(6):1253-72. PMCID: 2715063.
  17. Fischer W, Ganusov VV, Giorgi EE, Hraber PT, Keele BF, Leitner T, Han CS, Gleasner CD, Green L, Lo CC, Nag A, Wallstrom TC, Wang S, McMichael AJ, Haynes BF, Hahn BH, Perelson AS, Borrow P, Shaw GM, Bhattacharya T, Korber BT. Transmission of single HIV-1 genomes and dynamics of early immune escape revealed by ultra-deep sequencing. PLoS One. 2010;5(8):e12303. PMCID: 2924888.

About the author: Dr. McMichael is the Director of Weatherall Institute of Molecular Medicine in Oxford University and founded the Medical Research Council Human Immunology Unit. He was knighted in 2008 for services to medical sciences and is a Fellow of the Royal Society.

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