HIV Neutralizing Antibodies: Isolation, characterization, and interaction with viral variants

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Neutralizing antibodies (NAb) against HIV-1 are likely to be a major component of an effective vaccine-induced immune response. Cross-reactive NAbs commonly arise during HIV-1 infection, though only a small subset of infected patients produce NAbs with high breadth and potency. In contrast, the HIV-1 envelope glycoprotein (Env) vaccine immunogens tested to date have failed to elicit cross-reactive neutralizing antibodies. Thus, studying the development of broadly neutralizing antibodies (bNAbs) in infected individuals may provide important lessons for vaccine design. In addition, the isolation of bNAbs from selected donors and vvaccinated animals has greatly aided our understanding of HIV-1 Env structure and vulnerability to neutralizing antibodies and such antibodies have potential for prevention or treatment of HIV-1 infection. For several years our lab has been a leader in the field of isolating and characterizing broadly neutralizing antibodies from HIV-infected donors. We have pioneered the development of reagents for isolating epitope-specific B cells, as well as a method for high-throughput screening of unselected B cells. After identification by one of these methods, we recover IgG from the B cells by single-cell PCR, subcloning, and expression in mammalian cells. The resulting antibodies are assayed for virus binding and neutralization, and their breadth, potency, epitopes, and modes of recognition analyzed. We also use next-generation deep sequencing to find clonal relatives of the antibodies and to understand their origins in B cell development. For the latter studies, donors for whom we have longitudinal samples from the time of HIV infection are particularly valuable. In addition, we apply these techniques to the study of animals that were immunized with candidate vaccines. In the past year, our work has included: isolation of monoclonal antibodies from three HIV-infected patients that developed broadly neutralizing serum; for one such patient, assessment of the activity of such antibodies against the patients' own viral variants; isolation and next-generation sequencing analysis of antibodies from rhesus macaques vaccinated with candidate HIV vaccines; flow cytometric analysis of binding of monoclonal antibodies to HIV envelope proteins, in support of biophysical studies with collaborators at the VRC and Yale University; and analysis of HIV envelope proteins isolated from patients treated with the antibody VRC01.