Characterizing the neutralizing antibody responses of HIV-1 superinfected individuals

Abstract

Eliciting a neutralizing antibody (Nab) response that is protective against diverse HIV-1 variants presents a major challenge to vaccine development. The identification of broad, cross-reactive Nabs from HIV-infected individuals has revealed epitopes on the HIV-1 Envelope that could be exploited by immunogen design. Ideally, presentation of these epitopes would elicit similarly broad and cross-reactive antibodies in vaccinated individuals. However, it is unclear how such immunogens should be constructed since none tested to date have been able to elicit a protective Nab response. Also, broadly Nabs isolated from HIV-infected individuals exhibit unusual characteristics, such as extensive amounts of somatic hypermutation, which is not typically induced by vaccination, but is thought to require years of chronic antigenic stimulation. To improve our understanding of the development of Nabs in response to HIV-1 during natural infection, the three studies in this thesis were initiated to investigate one potential mechanism by which Nab breadth and potency develops by studying cases of HIV-1 superinfection (SI). We previously showed that Nab breadth is positively associated with viral diversity, which first led us to hypothesize that superinfected individuals would develop broader and more potent Nab responses than singly infected individuals as a result of the increased antigenic stimulation associated with increased viral diversity. In Chapter II we show that superinfected individuals develop significantly broader and more potent Nab responses compared to singly infected individuals, which was independent of Nab breadth or potency developed prior to SI, CD4+ T cell count and viral load. We also noted two individuals exhibited elite neutralizing activity following SI, with cross-subtype breadth detected within 1.5 years following initial infection, earlier than what is typically seen in singly infected populations. Next, we sought to determine whether the Nab response that develops following SI is mediated by a dominant monoclonal response to known epitopes or whether it is mediated by a polyclonal response, targeting multiple different epitopes. In Chapter III, we used standard epitope mapping techniques and computational prediction to show that the four main regions on the HIV-1 Envelope commonly targeted by singly infected populations-- the CD4-binding site, V1/V2 glycans, V3 glycans, and the membrane proximal external region of gp41-- are not the principle Nab targets of superinfected individuals. These findings suggest that a broad and potent Nab response following exposure to diverse HIV antigens does not involve a dominant response to any one of these four conserved regions, some of which are conformationally masked or occluded. This may suggest that superinfected individuals primarily generate polyclonal Nab responses and that there are other ways to elicit a broad and potent Nab response beyond what has been uncovered with these four main epitopes. Finally, Chapter IV describes initial efforts to further characterize the Nab response of one superinfected individual (QB850) who developed the broadest and most potent Nab response in the SI cohort. Individual monoclonals were isolated and characterized, with only a subset demonstrating HIV specificity. The chapter more broadly describes the B cell culture and cloning pipeline in our laboratory and the steps that will require additional optimization with the help of a high-throughput liquid handling system. Identification of additional monoclonal Nabs from this patient may ultimately help resolve the individual targets of her Nab response, which could lead to the identification of new epitopes on Envelope. In summary, these studies demonstrate that exposure to diverse antigens during HIV-1 SI augments the Nab response, without targeting conserved epitopes. Thus, future studies should evaluate the possibility of a vaccination strategy that mimics SI using diverse immunogens to elicit a broad and potent polyclonal Nab response in vaccinated individuals.