CD8+ T cells are a key focus of vaccine development efforts for HIV. However, there is no clear consensus as to which of the nine HIV proteins should be used for vaccination. The early proteins Tat, Rev, and Nef may be better CD8+ T cell targets than the late-expressed structural proteins Gag, Pol, and Env. In this study, we show that Gag-specific CD8+ T cells recognize infected CD4+ T lymphocytes as early as 2 h postinfection, before proviral DNA integration, viral protein synthesis, and Nef-mediated MHC class I down-regulation. Additionally, the number of Gag epitopes recognized by CD8+ T cells was significantly associated with lower viremia (p = 0.0017) in SIV-infected rhesus macaques. These results suggest that HIV vaccines should focus CD8+ T cell responses on Gag.

HIV-1 Nef and HIV-1–specific cytotoxic T lymphocytes (CTLs) have important and opposing roles in the immunopathogenesis of HIV-1 infection. Nef-mediated down-modulation of HLA class I on infected cells can confer resistance to CTL clearance, but the factors determining the efficiency of this process are unknown. This study examines the impact of Nef on the antiviral activity of several CTL clones recognizing epitopes from early and late HIV-1 proteins, restricted by HLA-A, -B, and -C molecules. CTL-targeting epitopes in early proteins remained susceptible to the effects of Nef, although possibly to a lesser degree than CTL-targeting late protein epitopes, indicating that significant Nef-mediated HLA down-regulation can precede even the presentation of early protein-derived epitopes. However, HLA-C–restricted CTLs were unaffected by Nef, consistent with down-regulation of cell-surface HLA-A and -B but not HLA-C molecules. Thus, CTLs vary dramatically in their susceptibility to Nef interference, suggesting differences in the relative importance of HLA-A– and HLA-B– versus HLA-C–restricted CTLs in vivo. The data thus indicate that HLA-C–restricted CTLs may have an under-appreciated antiviral role in the setting of Nef in vivo and suggest a benefit of promoting HLA-C–restricted CTLs for immunotherapy or vaccine development.

The live attenuated simian immunodeficiency virus (LASIV) vaccine SIVΔnef is one of the most effective vaccines in inducing protection against wild-type lentiviral challenge, yet little is known about the mechanisms underlying its remarkable protective efficacy. Here, we exploit deep sequencing technology and comprehensive CD8 T cell epitope mapping to deconstruct the CD8 T cell response, to identify the regions of immune pressure and viral escape, and to delineate the effect of epitope escape on the evolution of the CD8 T cell response in SIVΔnef-vaccinated animals. We demonstrate that the initial CD8 T cell response in the acute phase of SIVΔnef infection is mounted predominantly against more variable epitopes, followed by widespread sequence evolution and viral escape. Furthermore, we show that epitope escape expands the CD8 T cell repertoire that targets highly conserved epitopes, defined as anentropic specificity, and generates de novo responses to the escaped epitope variants during the vaccination period. These results correlate SIVΔnef-induced protection with expanded anentropic specificity and increased response depth. Importantly, these findings render SIVΔnef, long the gold standard in HIV/SIV vaccine research, as a proof-of-concept vaccine that highlights the significance of the twin principles of anentropic specificity and repertoire depth in successful vaccine design.

Defining the correlates of immune protection conferred by SIVΔnef, the most effective vaccine against SIV challenge, could enable the design of a protective vaccine against HIV infection. Here we provide a comprehensive assessment of immune responses that protect against SIV infection through detailed analyses of cellular and humoral immune responses in the blood and tissues of rhesus macaques vaccinated with SIVΔnef and then vaginally challenged with wild-type SIV. Despite the presence of robust cellular immune responses, animals at 5 weeks after vaccination displayed only transient viral suppression of challenge virus, whereas all macaques challenged at weeks 20 and 40 post-SIVΔnef vaccination were protected, as defined by either apparent sterile protection or significant suppression of viremia in infected animals. Multiple parameters of CD8 T cell function temporally correlated with maturation of protection, including polyfunctionality, phenotypic differentiation, and redistribution to gut and lymphoid tissues. Importantly, we also demonstrate the induction of a tissue-resident memory population of SIV-specific CD8 T cells in the vaginal mucosa, which was dependent on ongoing low-level antigenic stimulation. Moreover, we show that vaginal and serum antibody titers inversely correlated with post-challenge peak viral load, and we correlate the accumulation and affinity maturation of the antibody response to the duration of the vaccination period as well as to the SIVΔnef antigenic load. In conclusion, maturation of SIVΔnef-induced CD8 T cell and antibody responses, both propelled by viral persistence in the gut mucosa and secondary lymphoid tissues, results in protective immune responses that are able to interrupt viral transmission at mucosal portals of entry as well as potential sites of viral dissemination.