Aerosolized adenovirus afforded the same protection against viremia and pathogenicity in an IV SIVmac251 challenge model as IM delivery, despite the lack of peripheral T-cell responses and neutralizing antibodies at the time of challenge. This included reduced viremia throughout acute infection and attenuated mucosal CD4+ T-cell depletion. SIV inoculated rectally was also controlled to a similar extent by these disparate rAd5 delivery routes, although vaccination did not impact disease indicators as dramatically as in the IV model. Specifically, rAd5 immunization lowered peak viremia, but not other acute time points, while mucosal CD4+ T-cell depletion, which was much milder in this model, was not significantly ameliorated by immunization. To our knowledge, these data represent the first test of mucosal replication-deficient rAd5 vaccination against mucosal SIV challenge and demonstrate that AE immunization provides at least comparable protection to IM delivery.
The similar IV challenge outcome for the AE and IM groups was not anticipated based on their respective pre-challenge immunogenicity profiles. No T-cell responses were detected in blood at the time of challenge in the AE group; thus, circulating virus-specific T cells did not predict virus control in these animals. A similar disconnect was previously described for animals immunized with replication-competent Ad5 host range mutant vaccination, in which oral / oral delivery was much less immunogenic than combination oral / intranasal delivery, although both regimens controlled IR SIVmac251
challenge to the same extent.27
Thus, mucosally derived T cells from tissues targeted by mucosal vaccines and associated draining lymph nodes are able to contribute to systemic virus control. This suggests that lymphocytes residing or primed in classical “ effector “ sites may be an important source of immunity, and commonly used assays measuring cellular immunogenicity in the blood may not correlate with vaccine efficacy against systemic or mucosal challenges.
It was also unexpected that the AE immunization did not provide greater protection against mucosal challenge than IM delivery. We hypothesized that T- and B-cell responses mounted in the airways would enhance rectal immunity to a greater extent than systemic delivery based on studies demonstrating a benefit to airway delivery: nasal vaccination was superior to IM vaccination in protecting against IR SIV challenge with a DNA / recombinant-modified vaccinia virus Ankara platform;28
and lower respiratory tract intratracheal delivery offered superior protection against IR SIV challenge than combined oral and nasal delivery in a replication-competent Ad5 vaccination model.2
However, we did not detect significant rectal humoral or jejunal cellular responses following rAd5 administration by either route; therefore, rAd5 is likely insufficient to induce robust mucosal responses at distal sites. This is supported by our observations of (i) significantly higher rectal IgA when AE rAd5 is primed by systemic DNA compared with an unprimed response (); and (ii) greater jejunal CD8+
T- cell responses elicited by DNA-primed IM rAd5 than unprimed IM rAd5 (Supplementary Figure S1
online). The only evidence that AE vaccination impacted the mucosal challenge outcome was a significantly greater anamnestic BAL CD4+
T-cell response to SIV ().
Why peak viremia values differed with the challenge route is unclear, although our IR values are consistent with lower values observed by other repeated limiting-dose IR challenge studies.28,29
This may be due to slower initiation of infection following IR inoculation, requiring several days for a small number of viruses to cross the mucosal barrier and replicate to levels achieved within a single day following IV or high-dose mucosal transmission. Our data suggest that such IR inoculations result in a highly asynchronous acute infection process that may not lend itself to accurate peak viremia quantification by weekly samplings.
Preservation of CD4+
T cells in the small intestine, and to a lesser extent in the BAL, following the IR challenge in control and immunized animals was remarkable and unanticipated. Mucosal immunopathogenesis following IR SIV challenge has not been studied extensively, but several reports document severe CD4+
T-cell depletion in rectal mucosa within 2–3 weeks of infection.28,30,31
The reported effects on the small intestine are less consistent, ranging from no significant loss up to 2 weeks after infection to depletion commencing on day 7 or 21.31–33
This discrepancy may be related to the dose of the IR challenge virus, which may determine viral dissemination and CD4+
T-cell cytopathicity in the upper gastrointestinal tract. Although the basis for this difference remains to be explored, our findings in the BAL were consistent with previously published findings from high-dose IR infection, demonstrating lung CD4+
T - cell depletion by day 21,31
suggesting that pathogenesis of rectal SIV challenge is less variable in the lung than in the gastrointestinal tract. Ultimately, mucosal challenge may favor a more rapid induction of the mucosal immune response, curbing viral replication and CD4+
T-cell depletion at mucosal sites.
The blood T-cell response correlates observed here are consistent with the well-documented contribution of both pre- and post-challenge responses to viremia control in IV and IR SIV challenge models.3,28,34–37
Importantly, we revealed an association between mucosal T-cell responses (primarily CD8) and viremia control as well as partial preservation of mucosal CD4+
T cells. The demonstration that these mucosally derived T cells can limit viral replication and pathogenesis is consistent with evidence that effector memory T cells residing in mucosa protect against mucosal SIV replication.1
Pre-challenge antibody responses did not predict viral control to the extent observed for cellular responses. However, the association between pre-challenge respiratory tract IgA responses and retention of BAL CD4+ T cells suggests that local antibodies limited pathogenesis. While we were unable to identify cellular or humoral correlates of IR SIV acquisition rate, it will be important to include mucosal IgA responses in these analyses in future studies. Despite this being a fairly large study by non-human primate standards, it is a relatively small study for the purpose of searching for immune correlates; our findings should be considered a guide for identifying and refining correlates in future studies. Certainly, we cannot exclude the possibility of other correlates that did not rise to statistical significance in our study.
Together, these data show comparable efficacy between systemic and AE rAd5 vaccination against SIVmac251-induced disease, regardless of the challenge route. Although we did not see any clear benefit of AE over IM immunization, comparing these regimens in the less stringent SIVsmE660 mucosal challenge model may better discriminate between their protective capacities. We demonstrate that the addition of a systemic DNA prime enhanced the distal mucosal humoral responses elicited by AE rAd5. In addition, we identified discrepancies in the pathogenesis of IV and IR SIV infection in the macaque model, raising the question of which model accurately reproduces HIV disease. We conclude that the AE modality represents a useful immunization vehicle for vaccine vectors not only against respiratory pathogens such as influenza, measles, and tuberculosis, but that it may also complement vaccine strategies against HIV and other pathogens by inducing excellent cellular and humoral immunity.