TLR agonists and/or IL-15 adjuvanted mucosal SIV vaccine reduced gut CD4+ memory T cell loss in SIVmac251-challenged rhesus macaques
Introduction
Dramatic loss of resident memory CD4+ T cells in the intestine occurs within 2–3 weeks post-SIV-infection irrespective of the route of initial viral entry [1], [2], [3], [4], [5]. This rapid and profound depletion of CD4+ T cells is more severe in the gut mucosa than the other compartments [6], and hard to be reconstituted in patients even on highly active antiretroviral therapy (HAART) [2], [7]. Recently, a long-term benefit of protecting against gut mucosal [8], [9], and systemic CD4+ T cell loss [10] has been demonstrated. For example, long-term non-progressors (LTNPs) had a higher frequency of mucosal CD4+ T cells as compared to progressors [8], and early restoration of mucosal CD4+ memory CCR5+ T cells in the gut of SIV-infected rhesus predicted LTNP [9]. In a vaccine trial, Letvin et al. showed that preserved CD4+ central memory T cells several months after infection correlated with long-term protection and predicted the efficacy of an HIV-vaccine better than set-point viral load (VL) [10]. If CD4+ T cells were depleted during immunization as demonstrated by Vaccari et al., a decreased protection against SIVmac251 challenge was observed, which further confirmed the important role of CD4+ T cells in the course of HIV infection and AIDS development [11]. These studies suggested the significance of protecting mucosal/systemic CD4+ T cells from infection and destruction during HIV infection; however, current vaccine strategies hardly achieved this goal. Even if plasma and tissue viral loads were reduced in some of the macaque studies, significant mucosal CD4+ T cell preservation was not observed [17], [57]. Here we surprisingly observed the opposite discordance, namely that CD4+ T cells were preserved even when VL was not reduced. Although vaccine protection of CD4+ T cells may be expected to occur if VL is reduced, protection of CD4+ T cells without VL reduction is novel and unexpected, and requires further examination, as we have attempted here. Strategies that could protect against both systemic and gut mucosal CD4+ T cell loss during HIV/SIV infection would be desirable.
One way to achieve this, which we demonstrated in this study, was to use molecular adjuvants, such as Toll-like receptor (TLR) agonists and/or IL-15, during SIV vaccine immunization. It has been known that adjuvant plays an important role to increase the magnitude, breadth, and the quality of the immune responses, and vaccines with certain adjuvants confer better protection against the subsequent SIV/SHIV challenge. TLR agonists activate and mature dendritic cells to enhance immune responses [12], [13], [14], [15], [16], while IL-15 promotes the homeostatic expansion of CD8+ memory T cells [17], [18], [19], [20], and the induction of higher avidity, longer-lived T cells [21], [22], [23], [24]. Both have been shown to be good adjuvants in mouse and macaque models [12], [13], [15], [16], [23], [24]. In our current macaque study, we found that the combination of TLR agonists and IL-15 as an adjuvant in a mucosal SIV vaccine regimen enhanced the quality of vaccine-induced responses, which included long-lived APOBEC3G (A3G) and polyfunctional CD8+ T cell responses, and partially protected the SIVmac251-challenged macaques [25]. Though we did not observe any reduction of VLs in the animals vaccinated with the same mucosal SIV vaccine regimen with only TLR agonists or IL-15 alone as an adjuvant [25], we have now surprisingly found a significant preservation of CD4+ T cell numbers in the colon mucosa (and to a lesser extent in the ileum and peripheral blood) of these animals 6 months post-infection.
In an attempt to explore the possible mechanisms of this CD4+ T cell preservation in the SIVmac251 infected macaques, we first examined whether the pre-challenge A3G expression levels and antigen-specific polyfunctional CD8+ T cell responses, which we have explored in our previous study, were correlated with CD4+ T cell preservation. Interestingly, A3G, which was previously found inversely correlated with plasma VL reduction, demonstrated a positive correlation with CD4+ T cell preservation in the SIV-infected colons, indicating the profound effects of APOBEC3G. This was in contrast to pre-challenge polyfunctional CD8+ T cell responses, which were inversely correlated with postchallenge plasma viral loads, but surprisingly did not correlate with post-infection CD4+ T cell preservation in the colon at all. We further explored cytokine/chemokine expression levels, gp120 binding and neutralizing antibodies, and post-challenge mucosal viral-specific tetramer+ CD8+ T cell responses, and found that a significantly higher level of SIV-specific tetramer responses in the colons of the adjuvanted animals was induced/maintained even 6 months after SIVmac251 challenge compared to those without adjuvant. These SIV-specific tetramer responses were also positively correlated with the CD4+ T cell preservation. It is unusual here to see tetramer responses after 6 months of infection correlating with vaccine efficacy, as such responses are heavily influenced by viral load, which did not correlate.
Taken together, we found that the usage of certain molecular adjuvants in HIV mucosal vaccines was beneficial for improving the gut CD4+ memory T cell numbers upon infection, unexpectedly independent of its effect on VL, and thus might be useful for future HIV vaccine design.
Section snippets
Rhesus macaques, immunization and challenge
25 Indian rhesus macaques (Macaca mulatta) were immunized and challenged as previously described [25]. Briefly, the macaques were maintained in accordance with guidelines of the Association for Assessment and Accreditation of Laboratory Animal Care International and with approval of the NCI Animal Care and Use Committee. They were all seronegative for SIV, simian retroviruses 1, 2 and 5, and simian T-cell leukemia/lymphotropic virus type 1 prior to the study. All macaques were Mamu-A*01+,
CD4+ memory T cell numbers were preserved after SIV infection in the colons of the macaques immunized with vaccines containing adjuvants
In our recent macaque study, we have intrarectally immunized four groups of macaques with a peptide-prime, MVA-boost SIV vaccine regimen with or without adjuvant: group 1 adjuvanted with a combination of TLR 2, 3, and 9 agonists, group 2 with IL-15, group 3 with both TLR agonists and IL-15, and group 4 with vaccine but without any adjuvant [25]. A group 5 was added with TLR agonists and IL-15 adjuvant only but without vaccine as an adjuvant control group [25]. Upon SIVmac251 challenge, only
Discussion
In our recent SIV-macaque study, we have evaluated the protective efficacy of using TLR agonists, IL-15 and the combination of these two as adjuvants in a peptide-primed/MVA-boosted mucosal SIV vaccine against intrarectal SIVmac251 challenge [25]. As shown before, mucosal administration of the combination of both adjuvants in the macaques induced high levels of antigen-specific CD4+ and polyfunctional CD8+ T cell responses after immunization, and conferred partial protection against the
Acknowledgments
This work was supported in part by the Intramural Program of the National Institutes of Health, National Cancer Institute, Center for Cancer Research, and the NIH Intramural AIDS Targeted Antiretroviral Program. We thank the NIAID tetramer core facility for providing the tetramers.
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Present address: University of Michigan, School of Medicine, Ann Arbor, MI, United States.