The induction of neutralizing antibody (NAb) and cytotoxic T-lymphocyte (CTL) responses are key goals for HIV-1 vaccine development. Recently, the phase III efficacy trial of the prime-boost combination of vaccines containing ALVAC-HIV and AIDSVAX B/E has offered the first evidence of vaccine-induced partial protection in humans (39
). The vaccine appeared to induce NAb responses with a narrow specificity profile and minimal CD8+
CTL responses (39
), suggesting that nonneutralizing Ab and cellular responses other than those of CD8+
CTL have played a role in conferring protection.
A number of studies have suggested that ADCC play an important role in the control of SIV and HIV-1 infection. Several studies have shown that the magnitude of ADCC Ab responses correlates inversely with virus set point in acute SIV infection in both unvaccinated macaques (45
) and in vaccinated animals after challenge (2
). In humans, ADCC-mediating Abs have been shown to protect against HIV-1 infection in mother-to-infant transmission (26
) and to correlate with both control of virus replication (22
) and lack of progression to overt disease (3
). In contrast, weakly neutralizing and nonneutralizing antibodies were shown to not protect against vaginal simian-human immunodeficiency virus (SHIV) challenge in macaques (8
ADCC is one of the mechanisms that might have conferred protection from infection in RV144 (17
). For this reason, we sought to isolate MAbs that can mediate ADCC from ALVAC-HIV/AIDSVAX B/E vaccine recipients and determine their specificity, clonality, and maturation. In this study, we have demonstrated that the ALVAC-HIV/AIDSVAX B/E vaccine elicited antibodies that mediate ADCC in the majority of the vaccinated subjects, which is in line with previous observations (17
), and that gp120 C1 region-specific A32-like antibodies significantly contributed to the overall ADCC responses. By isolating 23 ADCC-mediating MAbs from multiple vaccine recipients, we also demonstrated the presence of ADCC-mediating MAbs of additional specificities. In addition, we determined that the ADCC-mediating MAbs underwent limited affinity maturation and preferentially used VH1 gene segments.
Antibody responses that mediate ADCC were directed toward A32-blockable conformational epitopes (n
= 19), a non-A32-blockable conformational epitope (n
= 1), the gp120 Env V2 region (n
= 2) (23
), and a linear epitope in the gp120 V3 region (n
= 1). The conformational epitope recognized by the A32 MAb is a dominant target of HIV-1-positive plasma ADCC antibodies (13
), and A32-like MAbs are among the anti-HIV-1 CD4i Ab responses that are detected following HIV-1 transmission (38
). The identification of A32-like MAbs in vaccine recipients suggests that the gp120 epitope recognized by the A32 MAb is an immunodominant region not just in response to natural infection but also upon vaccination. Our data suggest that this A32-binding region reacts with antibodies that have a diverse binding profile, suggesting that the RV144 vaccine targeted multiple related but distinct conformational epitopes on gp120. These epitopes have been shown to be upregulated on the RV144 immunogen and to be efficiently presented by novel Env designs (S. M. Alam, unpublished data), thus it will be possible to test this vaccine strategy in future vaccine trials targeted to different HIV-1 subtypes.
In contrast to ADCC-mediating antibodies, HIV-1 bNAb responses have been reported to appear an average of 2 to 4 years after HIV-1 transmission (16
), suggesting that different levels of Ab maturation are required to mediate ADCC and neutralizing activities. Indeed, the mutation frequencies observed in the MAbs isolated from the ALVAC-HIV/AIDSVAX B/E vaccine recipients in our study were low (0.5 to 5.1%) and well below the ~6% changes in variable-domain amino acid sequences commonly seen as greater affinity for the cognate antigen is acquired (30
). We did, however, find that higher degrees of VH somatic mutation correlated with greater maximal percent GzB activity (), consistent with vaccine-driven affinity maturation. Whether repeated boosting of vaccine recipients would result in on-going maturation of these antibodies to further increase ADCC activity, CD4 blocking, or addition of neutralizing activity remains to be determined.
Finally, while ADCC-mediating MAbs were isolated that used diverse VH genes, we observed a clear preferential usage of the VH1 heavy-chain gene (74%) similar to that of potent bNAbs directed against the CD4bs (41
). Therefore, while these findings prove that the ADCC-mediating response in these subjects was not restricted to a specific VH gene family and are consistent with there being no obvious strong regulatory mechanisms that would inherently limit the generation of antibodies with ADCC activity, the preferential use of the VH1 gene raises the possibility that the Env proteins used in RV144 or Env gp120 proteins in general preferentially induce the use of the VH1 gene family. Whether a vaccine regimen can be developed that will harness the observed Ig VH1 gene-using B cells to also induce CD4bs antibodies with a high degree of mutation is currently unknown. It is interesting that we were able to recover ADCC antibodies with a degree of CD4 blocking activity that had low levels of mutation, suggesting that B cells expressing those antibodies can be harnessed to produce the desired potent CD4-blocking antibody response under the right conditions.
In summary, the ALVAC-HIV/AIDSVAX B/E vaccine induced potent ADCC responses mediated by modestly mutated and predominantly A32-blockable MAbs that have overlapping but distinct binding profiles. This response is qualitatively similar to anti-HIV-1 responses observed during chronic HIV-1 infections and may have been partly responsible for the modest degree of protection observed. ADCC-mediating MAbs predominantly utilized the VH1 Ig heavy-chain family, which has been previously reported for CD4bs-directed broadly neutralizing antibodies. This observation raises the possibility that continued boosting with this vaccine formulation leads to further somatic mutations of VH1 gp120-specific antibodies and, perhaps, to an enhanced ability to augment any protective effect they might have had to limit HIV-1 acquisition.