The high prevalence of pre-existing immunity to adenovirus serotype 5 (Ad5) in human populations may substantially limit the immunogenicity and clinical utility of recombinant Ad5 vector-based vaccines for HIV-1 and other pathogens. A potential solution to this problem is to use vaccine vectors derived from adenovirus (Ad) serotypes that are rare in humans, such as Ad35. However, cross-reactive immune responses between heterologous Ad serotypes have been described and could prove a major limitation of this strategy. In particular, the extent of immunologic cross-reactivity between Ad5 and Ad35 has not previously been determined. In this study we investigate the impact of pre-existing anti-Ad5 immunity on the immunogenicity of candidate rAd5 and rAd35 vaccines expressing SIV Gag in mice. Anti-Ad5 immunity at levels typically found in humans dramatically blunted the immunogenicity of rAd5-Gag. In contrast, even high levels of anti-Ad5 immunity did not substantially suppress Gag-specific cellular immune responses elicited by rAd35-Gag. Low levels of cross-reactive Ad5/Ad35-specific CD4+ T lymphocyte responses were observed, but were insufficient to suppress vaccine immunogenicity. These data demonstrate the potential utility of Ad35 as a candidate vaccine vector that is minimally suppressed by anti-Ad5 immunity. Moreover, these studies suggest that using Ad vectors derived from immunologically distinct serotypes may be an effective and general strategy to overcome the suppressive effects of pre-existing anti-Ad immunity.

The intense selection pressure exerted by virus-specific cytotoxic T lymphocytes (CTL) on replicating human immunodeficiency virus and simian immunodeficiency virus results in the accumulation of CTL epitope mutations. It has been assumed that fitness costs can limit the evolution of CTL epitope mutations. However, only a limited number of studies have carefully examined this possibility. To explore the fitness costs associated with viral escape from p11C, C-M-specific CTL, we constructed a panel of viruses encoding point mutations at each position of the entire p11C, C-M epitope. Amino acid substitutions at positions 3, 4, 5, 6, 7, and 9 of the epitope significantly impaired virus replication by altering virus production and Gag protein expression as well as by destabilizing mature cores. Amino acid substitutions at position 2 of the epitope were tolerated but required reversion or additional compensatory mutations to generate replication-competent viruses. Finally, while amino acid substitutions at positions 1 and 8 of the p11C, C-M epitope were functionally tolerated, these substitutions were recognized by p11C, C-M-specific CTL and therefore provided no selection advantage for the virus. Together, these data suggest that limited sequence variation is tolerated by the region of the capsid encoding the p11C, C-M epitope and therefore that only a very limited number of mutations can allow successful viral escape from the p11C, C-M-specific CTL response.

Previously we have shown that CD8 ؉ T cells are critical for containment of simian immunodeficiency virus (SIV) viremia and that rapid and profound depletion of CD4؉ T cells occurs in the intestinal tract of acutely infected macaques. To determine the impact of SIV-specific CD8 ؉ T-cell responses on the magnitude of the CD4 ؉ T-cell depletion, we investigated the effect of CD8 ؉ lymphocyte depletion during primary SIV infection on CD4؉ T-cell subsets and function in peripheral blood, lymph nodes, and intestinal tissues. In peripheral blood, CD8؉ lymphocyte-depletion changed the dynamics of CD4 ؉ T-cell loss, resulting in a more pronounced loss 2 weeks after infection, followed by a temporal rebound approximately 2 months after infection, when absolute numbers of CD4 ؉ T cells were restored to baseline levels. These CD4 ؉ T cells showed a markedly skewed phenotype, however, as there were decreased levels of memory cells in CD8 ؉ lymphocyte-depleted macaques compared to controls. In intestinal tissues and lymph nodes, we observed a significantly higher loss of CCR5 ؉ CD45RA ؊ CD4 ؉ T cells in CD8 ؉ lymphocyte-depleted macaques than in controls, suggesting that these SIV-targeted CD4 ؉ T cells were eliminated more efficiently in CD8 ؉ lymphocyte-depleted animals. Also, CD8؉ lymphocyte depletion significantly affected the ability to generate SIV Gag-specific CD4 ؉ T-cell responses and neutralizing antibodies. These results reemphasize that SIV-specific CD8 ؉ T-cell responses are absolutely critical to initiate at least partial control of SIV infection.