Activated CD4+CCR5+ T cells in the rectum predict increased SIV acquisition in SIVGag/Tat-vaccinated rhesus macaques

Abstract

An effective T-cell-based AIDS vaccine should induce strong HIV-specific CD8(+) T cells in mucosal tissues without increasing the availability of target cells for the virus. Here, we evaluated five immunization strategies that include Human adenovirus-5 (AdHu5), Chimpanzee adenovirus-6 (AdC6) or -7 (AdC7), Vaccinia virus (VV), and DNA given by electroporation (DNA/EP), all expressing Simian immunodeficiency virus group specific antigen/transactivator of transcription (SIV(mac239Gag/Tat)). Five groups of six rhesus macaques (RMs) each were vaccinated with DNA/EP-AdC6-AdC7, VV-AdC6-AdC7, DNA/-EP-VV-AdC6, DNA/EP-VV-AdC7, or AdHu5-AdHu5-AdHu5 and were challenged repeatedly with low-dose intrarectal SIVmac239. Upon challenge, there were no significant differences among study groups in terms of virus acquisition or viral load after infection. When taken together, the immunization regimens did not protect against SIV acquisition compared with controls but did result in an ∼ 1.6-log decline in set-point viremia. Although all immunized RMs had detectable SIV-specific CD8(+) T cells in blood and rectal mucosa, we found no correlation between the number or function of these SIV-specific CD8(+) T cells and protection against SIV acquisition. Interestingly, RMs experiencing breakthrough infection showed significantly higher prechallenge levels of CD4(+)C-C chemokine receptor type 5 (CCR5)(+)HLA-DR(+) T cells in the rectal biopsies (RB) than animals that remained uninfected. In addition, among the infected RMs, the percentage of CD4(+)CCR5(+)Ki-67(+) T cells in RBs prechallenge correlated with higher early viremia. Overall, these data suggest that the levels of activated CD4(+)CCR5(+) target T cells in the rectal mucosa may predict the risk of SIV acquisition in RMs vaccinated with vectors that express SIVGag/Tat.

Keywords: CD4; HIV; SIV; vaccination.

Fig. 1.

Heterologous prime-boost immunization regimens. Schematic representation of the experimental design that involved five groups of animals given three immunizations 16 wk apart: (1) DNA/EP followed by AdC6 and then AdC7; (2) VV followed by AdC6 and then AdC7; (3) DNA/EP followed by VV and AdC6; (4) DNA/EP followed by VV and AdC7; or (5) three immunizations of AdHu5. Each group included six MamuA*01⁺ adult Indian RMs. Three to five months following the final immunization all RMs and six additional unvaccinated control animals were challenged with repeated low-dose intrarectal SIV_mac239 every week up to 15 times. The times of immunization, challenge, and sample collections are indicated. PB, peripheral blood; RB, rectal biopsy.

Fig. 2.

Vaccination regimens result in robust SIV-specific immune responses. Average fraction of CD8⁺ T cells binding Gag-CM9 Mamu-A*01 tetramers at various time points postimmunization in peripheral blood (A) and rectal mucosa (B). Black dashed lines at 0, 16, and 32 wk indicate immunization time points. Error bars indicate the SEM for each vaccination group. The color scheme is as follows: DNA/EP-AdC6-AdC7, green; VV-AdC6-AdC7, red; DNA/EP-VV-AdC6, blue; DNA/EP-VV-AdC7, orange; and AdHu5-AdHu5-AdHu5, pink.

Fig. 3.

Level of activated and proliferating CCR5⁺CD4⁺ T cells after immunization in whole blood. Average levels of CCR5⁺ (A), Ki-67⁺CCR5⁺ (B), and HLA-DR⁺CCR5⁺ (C) T cells measured as the percentage of total CD4⁺ T cells in peripheral blood in vaccinated RMs. Black dashed lines at 0, 16, and 32 wk indicate immunization time points. Error bars represent SEM. The color scheme is as follows: DNA/EP-AdC6-AdC7, green; VV-AdC6-AdC7, red; DNA/EP-VV-AdC6, blue; DNA/EP-VV-AdC7, orange; and AdHu5-AdHu5-AdHu5, pink.

Fig. 4.

Viral acquisition and replication after low-dose rectal SIV_mac239 challenge. (A) Number of low-dose challenges required for acquisition of SIV infection in all vaccinated RMs and in unvaccinated controls. (B) All immunized animals combined compared with the control animals. (C) SIV plasma viral load (expressed as copies per milliliter of plasma) was measured at days 7, 21, and 42 after SIV infection by real-time PCR. (D) The average viral loads for all infected, immunized RMs combined compared with infected control animals. Error bars represent SEM. The color scheme is as follows: DNA/EP-AdC6-AdC7, green; VV-AdC6-AdC7, red; DNA/EP-VV-AdC6, blue; DNA/EP-VV-AdC7, orange; AdHu5-AdHu5-AdHu5, pink; controls, black; all vaccinated RMs combined, gray. *P < 0.05.

Fig. 5.

SIV infection is associated with higher levels of SIVGag-specific CD8⁺ T-cell responses in blood and CCR5⁺DR⁺CD4⁺ in rectal mucosa, and a low day 7 viral load is correlated with activated CCR5⁺CD4⁺ T cells in rectal mucosa. (A and B) Comparison of the percent of SIVGag peptide-responsive (A) or CD107a-expressing (B) CD8⁺ T cells in PBMCs before challenge in immunized RMs that remained uninfected (circles) or that acquired SIV infection (squares). Responsive cells are considered to be cytokine producing and/or CD107a expressing after background subtraction. (C and D) Comparison of the percent of CCR5⁺Ki-67⁺ (C) and CCR5⁺HLA-DR⁺ (D) CD4⁺ T cells in rectal mucosa before challenge in immunized RMs that remained uninfected (circles) and those that acquired SIV infection (squares). (E and F) The percentage of total CCR5⁺Ki-67⁺ (E) and effector memory (EM) CCR5⁺Ki-67⁺ (F) CD4⁺ T cells in rectal mucosa before challenge is plotted against the day 7 viral load in immunized RMs experiencing infection. The Mann–Whitney u test was used to determine differences between uninfected and infected groups. *P < 0.05; **P < 0.01 (Spearman’s correlation).