Developing an immunogen that elicits broadly neutralizing antibodies (bNAbs) is an elusive but important goal of HIV vaccine research, especially after the recent failure of the leading T cell based ...HIV vaccine in human efficacy trials. Even if such an immunogen can be developed, most animal model studies indicate that high serum neutralizing concentrations of bNAbs are required to provide significant benefit in typical protection experiments. One possible exception is provided by the anti-glycan bNAb 2G12, which has been reported to protect macaques against CXCR4-using SHIV challenge at relatively low serum neutralizing titers. Here, we investigated the ability of 2G12 administered intravenously (i.v.) to protect against vaginal challenge of rhesus macaques with the CCR5-using SHIV(SF162P3). The results show that, at 2G12 serum neutralizing titers of the order of 1:1 (IC(90)), 3/5 antibody-treated animals were protected with sterilizing immunity, i.e. no detectable virus replication following challenge; one animal showed a delayed and lowered primary viremia and the other animal showed a course of infection similar to 4 control animals. This result contrasts strongly with the typically high titers observed for protection by other neutralizing antibodies, including the bNAb b12. We compared b12 and 2G12 for characteristics that might explain the differences in protective ability relative to neutralizing activity. We found no evidence to suggest that 2G12 transudation to the vaginal surface was significantly superior to b12. We also observed that the ability of 2G12 to inhibit virus replication in target cells through antibody-mediated effector cell activity in vitro was equivalent or inferior to b12. The results raise the possibility that some epitopes on HIV may be better vaccine targets than others and support targeting the glycan shield of the envelope.
A comprehensive understanding of the development and evolution of human B cell responses induced by pathogen exposure will facilitate the design of next-generation vaccines. Here, we utilized a ...high-throughput single B cell cloning technology to longitudinally track the human B cell response to the yellow fever virus 17D (YFV-17D) vaccine. The early memory B cell (MBC) response was mediated by both classical immunoglobulin M (IgM) (IgM⁺CD27⁺) and switched immunoglobulin (swIg⁺) MBC populations; however, classical IgM MBCs waned rapidly, whereas swIg⁺ and atypical IgM⁺ and IgD⁺ MBCs were stable over time. Affinity maturation continued for 6 to 9 mo following vaccination, providing evidence for the persistence of germinal center activity long after the period of active viral replication in peripheral blood. Finally, a substantial fraction of the neutralizing antibody response was mediated by public clones that recognize a fusion loop-proximal antigenic site within domain II of the viral envelope glycoprotein. Overall, our findings provide a framework for understanding the dynamics and complexity of human B cell responses elicited by infection and vaccination.
Most animal studies using passive administration of HIV broadly neutralizing monoclonal antibodies (bnMAbs) have associated protection against high-dose mucosal viral challenge with relatively high ...serum concentrations of antibody. We recently identified several bnMAbs remarkable for their in vitro potency against HIV. Of these bnMAbs, PGT121 is one of the most broad and potent antibodies isolated to date and shows 10- to 100-fold higher neutralizing activity than previously characterized bnMAbs. To evaluate the protective potency of PGT121 in vivo, we performed a protection study in rhesus macaques. Animals were i.v. administered 5 mg/kg, 1 mg/kg, or 0.2 mg/kg PGT121 24 h before being vaginally challenged with a single high dose of chimeric simian-human immunodeficiency virus (SHIV) SF₁₆₂P₃. Sterilizing immunity was achieved in all animals administered 5 mg/kg and 1 mg/kg and three of five animals administered 0.2 mg/kg PGT121, with corresponding average antibody serum concentrations of 95 µg/mL, 15 µg/mL, and 1.8 µg/mL, respectively. The results suggest that a protective serum concentration for PGT121 is in the single-digit µg/mL for SHIV SF₁₆₂P₃, showing that PGT121 can mediate sterilizing immunity at serum concentrations that are significantly lower than those observed in previous studies and that may be achievable through vaccination with the development of a suitable immunogen.
The recent failure of the T-cell-based HIV vaccine trial led by Merck & Co., Inc. prompts the urgent need to refocus on the question of which T-cell responses are required to control HIV replication. ...The well-described association between the expression of particular MHC class I molecules and successful containment of HIV or, in the macaque model, SIV replication provide a valuable starting point from which to evaluate more precisely what might constitute effective CD8(+) T-cell responses. Here, we review recent studies of T-cell-mediated control of HIV and SIV infection, and offer insight for the design of a successful T-cell-based HIV vaccine in the future.
We previously demonstrated that HIV replication is concentrated in lymph node B cell follicles during chronic infection and that HIV-specific CTL fail to accumulate in large numbers at those sites. ...It is unknown whether these observations can be generalized to other secondary lymphoid tissues or whether virus compartmentalization occurs in the absence of CTL. We evaluated these questions in SIVmac239-infected rhesus macaques by quantifying SIV RNA(+) cells and SIV-specific CTL in situ in spleen, lymph nodes, and intestinal tissues obtained at several stages of infection. During chronic asymptomatic infection prior to simian AIDS, SIV-producing cells were more concentrated in follicular (F) compared with extrafollicular (EF) regions of secondary lymphoid tissues. At day 14 of infection, when CTL have minimal impact on virus replication, there was no compartmentalization of SIV-producing cells. Virus compartmentalization was diminished in animals with simian AIDS, which often have low-frequency CTL responses. SIV-specific CTL were consistently more concentrated within EF regions of lymph node and spleen in chronically infected animals regardless of epitope specificity. Frequencies of SIV-specific CTL within F and EF compartments predicted SIV RNA(+) cells within these compartments in a mixed model. Few SIV-specific CTL expressed the F homing molecule CXCR5 in the absence of the EF retention molecule CCR7, possibly accounting for the paucity of F CTL. These findings bolster the hypothesis that B cell follicles are immune privileged sites and suggest that strategies to augment CTL in B cell follicles could lead to improved viral control and possibly a functional cure for HIV infection.
The properties of the human immunodeficiency virus (HIV) pose serious difficulties for the development of an effective prophylactic vaccine. Here we describe the construction and characterization of ...recombinant (r), replication-competent forms of rhesus monkey rhadinovirus (RRV), a gamma-2 herpesvirus, containing a near-full-length (nfl) genome of the simian immunodeficiency virus (SIV). A 306-nucleotide deletion in the pol gene rendered this nfl genome replication-incompetent as a consequence of deletion of the active site of the essential reverse transcriptase enzyme. Three variations were constructed to drive expression of the SIV proteins: one with SIV's own promoter region, one with a cytomegalovirus (cmv) immediate-early promoter/enhancer region, and one with an RRV dual promoter (p26 plus PAN). Following infection of rhesus fibroblasts in culture with these rRRV vectors, synthesis of the early protein Nef and the late structural proteins Gag and Env could be demonstrated. Expression levels of the SIV proteins were highest with the rRRV-SIVcmv-nfl construct. Electron microscopic examination of rhesus fibroblasts infected with rRRV-SIVcmv-nfl revealed numerous budding and mature SIV particles and these infected cells released impressive levels of p27 Gag protein (>150 ng/ml) into the cell-free supernatant. The released SIV particles were shown to be incompetent for replication. Monkeys inoculated with rRRV-SIVcmv-nfl became persistently infected, made readily-detectable antibodies against SIV, and developed T-cell responses against all nine SIV gene products. Thus, rRRV expressing a near-full-length SIV genome mimics live-attenuated strains of SIV in several important respects: the infection is persistent; >95% of the SIV proteome is naturally expressed; SIV particles are formed; and CD8+ T-cell responses are maintained indefinitely in an effector-differentiated state. Although the magnitude of anti-SIV immune responses in monkeys infected with rRRV-SIVcmv-nfl falls short of what is seen with live-attenuated SIV infection, further experimentation seems warranted.
Simian immunodeficiency virus (SIV)-specific CD8(+) T cells kill SIV-infected CD4(+) T cells in an major histocompatibility complex class I (MHC-I)-dependent manner. However, they are reportedly less ...efficient at killing SIV-infected macrophages. Since the viral accessory protein Nef has been shown to downregulate MHC-I molecules and enhance cytotoxic T lymphocyte (CTL) evasion in human immunodeficiency virus type 1 (HIV-1)-infected CD4(+) T cells, we examined whether Nef played a role in protecting SIV-infected macrophages from killing by SIV-specific CD8(+) T cells. To explore the role of Nef in CD8(+) T cell evasion, we compared the ability of freshly sorted SIV-specific CD8(+) T cells to readily suppress viral replication or eliminate CD4(+) T cells or monocyte-derived macrophages infected with SIV variants containing wild-type (WT) or mutated nef genes. As expected, SIV-specific CD8(+) T cells suppressed viral replication and eliminated the majority of SIV-infected CD4(+) T cells, and this killing was enhanced in CD4(+) T cells infected with the nef variants. However, macrophages infected with nef variants that disrupt MHC-I downregulation did not promote rapid killing by freshly isolated CD8(+) T cells. These results suggest that mechanisms other than Nef-mediated MHC-I downregulation govern the resistance of SIV-infected macrophages to CD8(+) T cell-mediated killing. This study has implications for viral persistence and suggests that macrophages may afford primate lentiviruses some degree of protection from immune surveillance.
Myeloid cells are permissive for HIV/SIV replication in vitro and may contribute to viral persistence in vivo. While many studies have been geared to understanding how CD8(+) T cells control viral replication in CD4(+) T cells, the role of these cells in controlling viral replication in macrophages is less clear. Primary, unstimulated CD8(+) T cells insignificantly suppress viral replication or eliminate SIV-infected macrophages. Since the viral Nef protein downregulates MHC-I and provides infected cells some degree of protection from CD8(+) T cell-mediated effector functions, we evaluated whether Nef may be contributing to the resistance of macrophages to CD8(+) T cell suppression. Our results suggest that Nef is not involved in protecting infected macrophages from CD8(+) T cell killing and suggest that other mechanisms are involved in macrophage evasion from CD8 surveillance.
Cytotoxic T lymphocytes (CTLs) have a central role in the successful control of immunodeficiency virus infection. Evasion of this immune response through CTL escape is therefore an important factor ...in HIV and simian immunodeficiency virus pathogenesis. During the course of an infection, the precise timing of the occurrence of escape mutations and their location in the viral genome can indicate the efficacy of certain CTL specificities and the cost to viral fitness of particular escape mutations -- factors that are highly relevant to vaccine design. Also crucial for vaccine design is the extent to which CTL escape is driving the evolution of HIV at the population level. Here, we highlight the important lessons that can be learned from immunodeficiency virus CTL escape.
The adenovirus type 5 (Ad5)-based vaccine developed by Merck failed to either prevent HIV-1 infection or suppress viral load in subsequently infected subjects in the STEP human Phase 2b efficacy ...trial. Analogous vaccines had previously also failed in the simian immunodeficiency virus (SIV) challenge-rhesus macaque model. In contrast, vaccine protection studies that used challenge with a chimeric simian-human immunodeficiency virus (SHIV89.6P) in macaques did not predict the human trial results. Ad5 vector-based vaccines did not protect macaques from infection after SHIV89.6P challenge but did cause a substantial reduction in viral load and a preservation of CD4+ T cell counts after infection, findings that were not reproduced in the human trials. Although the SIV challenge model is incompletely validated, we propose that its expanded use can help facilitate the prioritization of candidate HIV-1 vaccines, ensuring that resources are focused on the most promising candidates. Vaccine designers must now develop T cell vaccine strategies that reduce viral load after heterologous challenge.
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