The primary goal for a cellular immune response-based vaccine in humans is to reduce chronic-phase viremia enough to extend the time between infection and AIDS. A secondary goal should be to reduce viral transmission to uninfected contacts of the infected vaccinee. A third goal of such a cellular immune response-based vaccine would be to ameliorate the loss of memory CD4+
cells that takes place in the acute phase (35
). One hallmark of long-term nonprogression in adults and children is preservation of the CD4+
memory compartment (13
). Reduction from 30,000 copies/ml to less than 1,000 copies/ml, a 1.5 log reduction, prevents transmission of HIV (27
) and increases longevity (43
). In Indian rhesus macaques, the usual viral set point for SIVmac239 is between 105
copies/ml, so a 1.5 log reduction would result in a plasma viral concentration of approximately 3,000 to 30,000 copies/ml.
Very few vaccine regimens have influenced disease course after challenge with the highly pathogenic SIVs. By contrast, a variety of vaccines have induced immune responses that have controlled chronic-phase viral replication of SHIV89.6P (3
). Unfortunately, many of these vaccines included an envelope with strong sequence similarity to the challenge virus. It is unlikely that the envelope of the vaccine and the envelope of the challenge virus will be similar in natural exposure to HIV type 1, given the sequence diversity of the envelope glycoprotein (10
). Broadly neutralizing antibodies will be required to neutralize the natural diversity of envelope sequences. These have been difficult to generate. In the absence of neutralizing antibody vaccines, we need to determine whether disease course can be ameliorated using T-cell responses alone. Vaccines that induce cellular immune responses are unlikely to provide sterilizing immunity because viral replication has to take place before cellular immune responses can be effective. However, it is possible that vaccine-induced cellular immune responses might control replication in the chronic phase. This would benefit both the health of the vaccinee and prevent transmission of the virus. Here we provide the first evidence that a vaccine that induces solely cellular immune responses can control chronic-phase viral replication.
Design of this vaccine regimen was based on a study that showed some control of chronic-phase replication of the highly pathogenic SIVmac239 challenge (11
). In this previous study, Mamu
macaques were vaccinated with vectors encoding gag
using a DNA prime-Ad5 boost regimen, then challenged with a high dose of SIVmac239. Similarly, a DNA prime-Sendai virus boost encoding gag
resulted in control of viral replication in 5 of 8 Burmese macaques challenged with SIVmac239 (40
). Evidence suggested an MHC-based control in the 5 successful vaccinees. We decided to build on these two encouraging studies by adding the early expressed genes of SIV to our vaccine regimen. We postulated that Tat-, Rev-, and Nef-specific CD8+
effector lymphocytes would be able to kill virally infected target cells earlier in the viral life cycle than Gag-specific CD8+
effector lymphocytes would, potentially prior to release of mature virus (29
). Furthermore, since Nef downregulates MHC class I molecules (16
), it is possible that CD8+
effector lymphocytes directed against Tat, Rev, and Nef might be more effective, since these proteins may be expressed before MHC class I molecules are downregulated. In the vaccine, the nef
antigen has been inactivated by incorporating a G2A mutation at the site of myristoylation to avoid this downregulation, which could decrease the effectiveness of the vaccine. Several investigators have made these arguments (2
), but the hypothesis has not been rigorously tested with a stringent viral challenge. Recently, Hel et al. showed some efficacy using early proteins, along with Gag, Pol, and Envelope, to ameliorate the pathological effects of SIVmac251 infection (31
), observing both decreased viremia and increased protection of CD4+
responses to Gag in particular.
Peak viremia and early-acute-phase viral concentrations were comparable in vaccinees between the gag-only study and this study until about 80 days postinfection (Fig. ). After 80 days postinfection, the plasma viral concentration in vaccinees in the gag-only study began to rise, eventually becoming indistinguishable from the control group. By comparison, in the current study, the plasma viral concentrations in the vaccinees have remained low for a year postinfection.
It is unclear which cellular immune responses are actually responsible for control of viral replication in our vaccinees. We know that Env-specific antibodies played no role in the control of viral replication in the vaccinees in the initial stages of infection, since Env was not included as an immunogen and there are no neutralizing antibodies prior to challenge. Since the CD4+
memory compartment shows some sign of preservation, it could be argued that Env-specific antibodies might have played a role in control later on in infection. Analysis of neutralizing antibodies, however, suggests that this was not the case. Enzyme-linked immunosorbent assay and Western blot analyses confirmed that all animals made antibodies to structural proteins, including Env (data not shown). However, the vaccinees did not make neutralizing antibodies, with the exception of 97111, who did not control her viral loads. Precedence for this result is found in the lymphocytic choriomeningitis virus system, where the presence of CD4 T helper responses prevented or delayed development of neutralizing antibody (53
). In fact, the animals that exerted the best control of viral replication (97113, 00044, and 01080), and also exhibited preservation of the memory CD4 subset, did not have significant titers of neutralizing antibodies. Several lines of evidence implicate immunodominant CD8+
lymphocyte responses in control. In the previous study, using only Gag in a DNA prime-Ad5 boost regimen, there was no evidence for control in Mamu-A*01-negative macaques (11
). Interestingly, Mamu
-positive macaques showed some level of control, if only temporarily, and this control might be attributable to the Gag CM9181-189
responses present in these animal. It is, therefore, possible that induction of Gag CM9181-189
responses played a large role in control of viral replication in our vaccinees. This might have been aided by vaccine-induced immune responses to Tat SL828-35
, another immunodominant T-cell response in Mamu
macaques. Indeed, these two CD8+
responses dominated the acute-phase anamnestic CD8+
responses, comprising more than 50% of anamnestic responses (Fig. ). This concept could be tested by removing the immunodominant epitopes from both the vaccine and the challenge virus in subsequent experiments. Additionally, despite their immunodominance, these two responses do not show a rank correlation with either decreased peak viremia or decreased set point viremia. Finally, Tat SL828-35
escapes rapidly, and by week 16 postinfection, all infected animals showed escape in this epitope, but vaccinees maintained control of viremia long after week 16 (data not shown).
While these experiments show the best observed non-Env vaccine-mediated control of SIVmac239 replication in Indian rhesus macaques thus far, there are several caveats. First, all eight vaccinees were Mamu-A*01+, and in the gag-only study, it was observed that animals expressing this allele do better after challenge, especially if they are vaccinated with regions of the virus that contain targets of their immunodominant CD8+ responses. Thus, we need to test this vaccine regimen in Mamu-A*01− macaques. Second, the challenge virus strain was exactly matched to the viral sequences in the vaccine. We therefore need to use a heterologous challenge to determine whether this regimen might have a chance of being effective in the field. Finally, the repeated low-dose challenge was administered 6 months after the Ad5 boost. We do not know enough about the long-term evolution of vaccine-induced memory cells to predict how long this vaccine-induced memory response will remain efficacious.
Despite these significant caveats, this study indicates that vaccine-induced cellular immune responses can control replication of a highly pathogenic SIV to a significant degree. The results raise the possibility that a vaccine inducing only cellular responses has the potential to contribute to control of SIVmac239.