CD4-binding-competent HIV-1 Env-based immunogens clearly interact with cells expressing primate CD4 in vivo
. We recently investigated the influence of this interaction on the immune responses elicited by soluble Env trimer immunization in rhesus macaques and showed that similar magnitudes of total Env-binding Ab and T cell responses were elicited by wt and CD4-binding-defective trimers. However, only wt trimers stimulated the production of CoRbs-directed Abs (13
). In the current study, the distinct difference in the elicitation of CoRbs Abs between animals immunized with wt and CD4-binding-defective trimer variants prompted us to investigate the role of CoRbs-directed Abs for protection against SHIV-SF162P4 and to ask whether Env-CD4 in vivo
interactions during Env trimer immunization of primates affect the protective response to SHIV challenge in any other measurable way.
The potential role of CoRbs-directed Abs for virus neutralization in vivo
and protection against HIV-1 or SHIV infection remains unclear. Access of intact Ab molecules to the conserved CoRbs on the HIV-1 functional virus spike is thought to be sterically restricted (31
), while smaller Ab subunits can access this region and interfere with viral entry (7
). Small ligands targeting this conserved region may therefore have potential for prevention and therapeutic intervention (28
). However, as a target for vaccine-induced Ab responses, the CoRbs may be less desirable unless Abs from this class that are different from and/or more potent than the Abs currently identified are elicited. Abs against the CoRbs are abundantly generated during natural HIV-1 infection in humans and following experimental SHIV challenge of naïve NHPs (12
), suggesting that the virus evolves and replicates efficiently in the presence of such humoral responses. The detection of CoRbs-directed Abs prior to autologous virus neutralization in HIV-1-infected patients also suggests that the CoRbs is nonneutralizing (17
). This is supported by data from the phase III Vaxgen clinical trial (VAX04), where no protection against acquisition of infection was observed (52
) despite high titers of CoRbs-directed Abs in all sera from a randomly selected but representative subset (14
). In contrast, CoRbs-directed Abs were suggested to be associated with more rapid viral clearance in macaques immunized with covalently linked gp120-CD4 complexes and then challenged with SHIV-SF162P3 in one study (12
). In the current study, we observed a similar reduction of viremia after intravenous SHIV-SF162P4 challenge in all immunized animals, but this was independent of the presence or absence of detectable CoRbs-directed Ab responses. These results show that CoRbs-directed Abs, at the titers elicited by the soluble YU2 Env trimers used here, did not contribute to the control of heterologous SHIV-SF162P4 infection in the current study. Additional studies are needed to determine if CoRbs-directed Abs with improved neutralization capabilities can be elicited by immunization, perhaps by using Env immunogens that better mimic the native HIV-1 Env spike. There is at present limited information about different subspecificities of CoRbs-directed Abs, and we cannot exclude that more potent neutralizing CoRbs-directed Abs can be generated by other vaccination protocols or by natural HIV-1 infection.
The SHIV-SF162P4 challenge virus is genetically related to the SHIV-SF162P3 virus, but the SHIV-SF162P4 Env is more neutralization sensitive and closely resembles the parental SF162 Env clone. The SHIV-SF162P4 stock was generated by in vivo
passage at peak viremia, likely in the absence of effective host-mediated immune pressure (1
). In contrast, the SHIV-SF162P3 stock was generated by in vivo
passage after prolonged replication in rhesus macaques, forcing the virus to evolve under the pressure of the host adaptive immune response (9
). Despite the frequent use of the SHIV-SF162P4 challenge virus, little is known about its sensitivity to well-characterized neutralizing MAbs. Here, we characterized the SHIV-SF162P4 challenge virus and a clone derived thereof (clone 41.1) for their sensitivities to known MAbs and the immune sera from monkeys immunized with the YU2-based Env trimers. We show that the SHIV-SF162P4 stock is less sensitive to several MAbs, including the two CoRbs-directed MAbs 17b and 48d and the two MPER-directed MAbs 2F5 and 4E10, than the parental SF162 virus. It is, however, highly sensitive to the CD4bs Ab b12 and the V3-directed 447 and 39F MAbs. Despite its sensitivity to these two V3-directed MAbs, markedly higher concentrations of the MAbs were required to neutralize the uncloned SHIV-SF162P4 stock compared to viruses pseudotyped with the parental SF162 Env or the SHIV-SF162P4-derived 41.1 Env. However, since the SHIV-SF162P4 clone generated here was not derived from single-genome amplification (57
), it is possible that there are other variants in the stock exhibiting a more resistant phenotype. The cell type from which the viruses were generated, C8166-CCR5 cells for the uncloned SHIV stock and 293 cells for the pseudotyped virus, may also affect the neutralization sensitivity (39
The plasma NAb titers after five immunizations measured against the SHIV-SF162P4 challenge virus were comparable to or slightly lower than the titers reported following immunization with homologous SF162-derived gp140 oligomers (3
). In the study by Barnett et al. (3
), macaques immunized with homologous SF162ΔV2 Env were protected from infection in response to vaginal challenge with SHIV-SF162P4. Similar results were observed in a study by Bogers et al. (4
), where three out of four SF162ΔV2 Env protein-boosted macaques were protected from rectal challenge with SHIV-SF162P4. The SF162-derived gp140 oligomers used in these studies were previously shown to induce high levels of V1-specific Abs (11
), and V1-directed Abs are known to be highly potent but also strain specific (8
). Recent attempts to compete out the neutralizing activity elicited by SF162-derived gp140 oligomers against SHIV-SF162P4 using unconstrained V1 or V3 peptides suggested that these specificities were not responsible for most of the activity (2
). Nevertheless, Abs specific for variable regions of SF162, not detected in these peptide competition assays, may be an important subset contributing to the protection from infection with homologous SHIV-SF162P4. V1-directed Abs have been implied to be important for control of SHIV-SF162P3 infection, as variants escaping from immune pressure during infection with this virus exhibit mutations in this region (9
As shown in our previous study, Env-specific CD4 and CD8 T cell responses prior to challenge were similar in the groups receiving wt, 368, or 423/425/431 trimers (13
). These responses did not correlate with the reduction in cumulative viral loads in the current SHIV challenge experiment (data not shown). The initial control of SHIV-SF162P4 replication after immunization has been shown to be independent of CD8 T cells (5
), and although infection induces a strong T cell response, this apparently does not correlate to viral clearance in SHIV-SF162P3 infection (47
). In the present study we observed enhanced control of SHIV-SF162P4 viremia in all YU2 trimer-vaccinated animals, suggesting that immune responses that are partially protective against heterologous SHIV challenge are elicited by this vaccine regimen. Furthermore, one monkey (F77) was apparently protected from infection, with no detectable viral loads. Immune parameters that contributed to protection in this animal could not be identified, as the titers of total gp120 binding Abs, NAbs, and T cell responses in this animal were of similar magnitude to those in the other animals (13
). We did not investigate other factors that may have influenced the acquisition of infection, such as early innate immune responses (33
), major histocompatibility complex (MHC) haplotypes (16
), or polymorphisms in genetic resistance factors (37
), as part of the current effort, since this was not the main focus of the study.
An interesting additional observation from the current study was that the anamnestic NAb response stimulated by the SHIV challenge was increased by 1 order of magnitude over the prechallenge NAb titers. There was a strong correlation between the titers of HIV-1 Env binding Abs and the neutralizing Abs against the challenge virus. Since the anamnestic Abs reacted equally well with YU2- and SF162-derived gp120, the increase in NAb titers stimulated by the challenge virus likely resulted from efficient boosting of vaccine-induced specificities rather than elicitation of new SHIV Env-specific B cell responses by the heterologous infection. This shows that there is room to further improve the overall magnitude of the Ab responses elicited by the protein in the adjuvant vaccine regimen used here.
In conclusion, we show that immunization with soluble Env trimers in adjuvant elicited immune responses that resulted in improved control of the heterologous SHIV-SF162P4 virus. There was no difference between the animals immunized with CD4-binding-competent or CD4-binding-defective Env timers, suggesting that Env-CD4 in vivo interactions and the elicitation of CoRbs Abs did not influence the level of protection achieved. Follow-up studies are required to dissect the immune correlates of protection in this model. Further studies will also determine whether the response elicited by the YU2 Env trimers might be more effective in blunting infection of heterologous SHIV challenge via the vaginal route of administration or if the quality of the elicited responses can be enhanced by different prime-boost regimens, improved adjuvants, or modified immunogen design.