Research ArticleHIV

A live-attenuated RhCMV/SIV vaccine shows long-term efficacy against heterologous SIV challenge

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Science Translational Medicine  17 Jul 2019:
Vol. 11, Issue 501, eaaw2607
DOI: 10.1126/scitranslmed.aaw2607
  • Fig. 1 Immunogenicity of ΔRh110 RhCMV/SIV vectors.

    (A) Schematic of the RM groups analyzed in this study. (B) Longitudinal and plateau-phase analysis of the vaccine-elicited, SIV Gag, Rev/Tat/Nef (RTN), Pol, and Env insert–specific CD4+ and CD8+ T cell responses in peripheral blood. In the top panel, the background-subtracted frequencies of cells producing TNF and/or IFN-γ by flow cytometric ICS assay to peptide mixes comprising each of the SIV inserts (SIVmac239 sequence) within the memory CD4+ or CD8+ T cell subsets were summed for overall responses, with the figure showing the mean (+ SEM) of these overall responses at each time point. In the bottom panel, boxplots compare the overall and individual SIV insert–specific CD4+ and CD8+ T cell response frequencies between the vaccine groups at the end of the vaccine phase (each data point is the mean of response frequencies in all samples from weeks 30 to 58 after first vaccination). Two-sided Wilcoxon rank-sum tests were used to compare the significance of differences in plateau-phase response frequencies between group 1 and group 2 (SIVmac239 versus SIVsmE660 inserts in ΔRh110 68-1 vectors) and between group 1 and group 4 (SIVmac239 inserts in WT 68-1 versus ΔRh110 68-1 vectors). (C) Boxplots compare the memory differentiation of the vaccine-elicited CD4+ and CD8+ memory T cells in peripheral blood responding to SIV Gag peptide mix (SIVmac239 sequence) with TNF and/or IFN-γ production at the end of vaccine phase (week 54 for groups 1 and 2; week 60 for group 4). Memory differentiation state was based on CD28 and CCR7 expression, delineating central memory (TCM), transitional effector-memory (TTrEM), and effector-memory (TEM), as designated. Two-sided Wilcoxon rank-sum tests were used to compare the significance of differences in the fraction of responding cells with a TCM phenotype (reciprocal of fraction with effector differentiation − TTrEM + TEM). (D) Same analysis as in (B), but for responses in lung airspace (BAL). Each data point for the boxplots is the mean of response frequencies in all samples from weeks 30 to 54 after first vaccination. (E) Boxplots show plateau-phase analysis (each point is the average of all samples between weeks 24 and 30 after first vaccination) of the vaccine-elicited CD8+ T cell responses to SIV Gag supertopes (SIVmac239 sequence; fig. S1B) in peripheral blood of group 1, group 2, and group 4 RMs by the same ICS assay described above. Gag276–284 (69) and Gag482–490 (120) are MHC-E–restricted supertopes; Gag211–222 (53) and Gag290–301 (73) are MHC-II–restricted supertopes (9, 10). Statistical testing was performed as described in (B). In all panels, n = 14, 14, and 16, respectively, for groups 1, 2, and 4, except group 4 in (E), where n = 10. Analyses were adjusted for multiple comparisons across inserts (B and D), epitopes (C), and supertopes (E) using the Holm method, and P values of ≤0.05 were considered significant. Analyses of total responses (B and D) were not adjusted.

  • Fig. 2 Cross-recognition by ΔRh110 RhCMV/SIVmac239 and RhCMV/SIVsmE660 vector–elicited T cells.

    (A and B) Flow cytometric ICS analysis of SIV-specific CD4+ and CD8+ T cell response frequencies (using TNF and/or IFN-γ readout in memory subset) in the blood of group 1 (n = 14; SIVmac239 inserts) and group 2 (n = 14; SIVsmE660 inserts) RMs in plateau phase (week 44 after first vaccination) comparing recognition of matched versus mismatched peptide mixes (SIVmac239 versus SIVsmE543; see fig. S2), including overall (summed) responses and responses to each SIV insert. Two-sided paired Wilcoxon rank-sum tests were used to compare the significance of differences in matched versus mismatched peptide mix recognition. Unadjusted (total responses) or Holm-adjusted (each insert-specific response) P values of ≤0.05 were considered significant. When significant differences were observed (reduction in response frequencies with mismatched peptide mixes), the median effect size (% reduction with mismatch) is shown. (C) ICS analysis of CD8+ T cell recognition of autologous CD4+ T cells infected with the SIVmac239 versus SIVsmE543 viruses (after background subtraction of the response to mock-infected autologous CD4+ T cells) in plateau phase (between weeks 49 and 57 after first vaccination). Statistical analysis was performed as described above, with n = 12 and 13 for groups 1 and 2, respectively.

  • Fig. 3 Efficacy of ΔRh110 RhCMV/SIV vectors.

    (A and B) Assessment of the outcome of effective challenge by longitudinal analysis of plasma viral load (A) and de novo development of SIV Vif–specific CD4+ (B, top panel) and CD8+ (B, bottom panel) T cell responses. RMs were challenged until the onset of any above-threshold SIV Vif–specific T cell response, with the SIV dose administered 2 or 3 weeks before this response detection considered the infecting challenge (week 0). RMs with sustained viremia were considered not protected (black); RMs with no or transient viremia were considered protected (red) (8). The fraction of protected RMs in the vaccinated groups (groups 1 and 2, n = 13 and 14, respectively) were compared to that of the unvaccinated group (group 3, n = 17) by Barnard’s exact test of binomial proportions, with the P values shown in (A). (C) BM cells and PBMCs were collected and cryopreserved from ΔRh110/SIVmac239/smE660 vaccine–protected RMs without any detectable viremia (RMs #1 to #3 from group 1; RMs #4 to #6 from group 2) at the indicated time points post-effective challenge (left panel; PID, post-infection day). Cells were thawed and administered intravenously (left panel) to six SIV-naïve RMs to assess the presence of replication-competent SIV, with the plasma viral dynamics in recipient RMs shown (right panel).

  • Fig. 4 Clearance of cell-associated SIV in the BM of ΔRh110 68-1 RhCMV/SIV vector–protected RMs.

    (A to D) Longitudinal analysis of PBMC-associated (A and C) and BM cell–associated (B and D) SIV RNA (left panels) and DNA (right panels) from 3 randomly selected unvaccinated RMs with progressive infection (A and B) and all 16 ΔRh110/SIVmac239/smE660 vector–protected RMs in groups 1 and 2 (C and D).

  • Fig. 5 Loss of circulating SIV infection–induced, SIV Vif–specific T cells in ΔRh110 68-1 RhCMV/SIV vector–protected RMs.

    (A) Long-term longitudinal analysis of plasma viral load in ΔRh110/SIVmac239/smE660 vector–protected (left and middle panels for groups 1 and 2, respectively) and WT 68-1/SIVmac239 vector–protected RMs [group 4, right panel; (8)]. (B) Long-term longitudinal analysis of SIV Vif–specific CD4+ (top panels) and CD8+ (bottom panels) among the same groups of ΔRh110 and WT 68-1 RhCMV/SIV vector–protected RMs, with the figure showing the mean (+SEM) of these SIV Vif–specific T cell response frequencies in the memory subset at each time point. (C) Wald tests comparing the slope (±95% confidence intervals) of decline of log-transformed SIV Vif–specific CD4+ (left panel) and CD8+ (right panel) T cell response frequencies. Calculation of slopes is described in Materials and Methods. In all analyses, n = 7, 9, and 8 for groups 1, 2, and 4, respectively.

  • Fig. 6 Necropsy analysis of ΔRh110 68-1 RhCMV/SIV vector–protected RMs.

    (A to C) Analysis of SIV Gag + Pol–specific (A) and SIV Vif–specific (B) CD4+ and CD8+ T cell response frequencies by flow cytometric ICS (using SIVmac239 peptide mixes; see Fig. 1) and tissue-associated SIV DNA and RNA by nested qPCR/RT-PCR (C) in tissues of four ΔRh110/SIVmac239/smE660 vector–protected RMs (RMs #7 and #8 from group 1; RMs #9 and #10 from group 2) taken to necropsy at 713 days (RM #7), 681 days (RM #8), 738 days (RM #9), and 745 days (RM #10) after infection. (D and E) Analysis of tissue-associated SIV DNA and RNA in tissues of two ΔRh110 68-1 RhCMV/SIVgag (SIVmac239 sequence insert) vector–vaccinated RMs that were taken to necropsy 531 and 763 days after vaccination without SIV challenge (negative controls) (D) and one SIVmac239-infected RM with progressive infection taken to necropsy 172 days after infection (positive control) (E). In (C) to (E), each data point indicates an independent tissue sample of the indicated tissue type and the dotted lines indicate the detection threshold. (F and G) Assessment of residual replication-competent SIV in cell suspensions obtained from the indicated tissue samples by in vitro coculture analysis (F) and by adoptive transfer of cells into four SIV-naïve RMs (G).

  • Fig. 7 Loss of transferable SIV in long-term ΔRh110 68-1 RhCMV/SIV vector–protected RMs.

    Second assessment of replication-competent SIV by adoptive transfer of cells from four long-term ΔRh110/SIVmac239/smE660 vector–protected RMs (RMs #1 and #2 from group 1; RMs #5 and #6 from group 2) that were previously shown to harbor replication-competent SIV by the same assay.

  • Fig. 8 Resistance of ΔRh110 68-1 RhCMV/SIV vector–protected RMs to repeat SIV challenge.

    (A and B) Outcome of repeat SIVmac239 challenge of long-term 68-1 RhCMV/SIV vector–protected RMs (n = 5, 7, and 8 for groups 1, 2, and 4, respectively) by longitudinal analysis of de novo SIV Vif–specific CD4+ and CD8+ T cell responses (A) and plasma viral load (B) with protected and nonprotected RMs defined as described in Fig. 3. (C) Third assessment of replication-competent SIV by adoptive transfer of cells from RMs #1, #2, #5, and #6 after effective rechallenge (reinduction of SIV Vif–specific T cell responses) with repeated aviremic protection.

Supplementary Materials

  • stm.sciencemag.org/cgi/content/full/11/501/eaaw2607/DC1

    Materials and Methods

    Fig. S1. ΔRh110 RhCMV/SIV vector design and supertope amino acid sequences.

    Fig. S2. Comparison of the amino acid sequence of SIVmac239 versus SIVsmE660/smE543 vaccine inserts.

    Fig. S3. Analysis of SIV Env–specific Ab responses after vaccination and after acquisition of SIV infection.

    Fig. S4. Acquisition of SIV infection by groups 1, 2, and 3 RMs with repeated, limiting-dose IVag SIVmac239 challenge.

    Fig. S5. Immune correlates analysis.

    Fig. S6. Analysis of SIV Gag– and SIV Pol–specific CD4+ and CD8+ T cell responses after SIV infection.

    Fig. S7. Analysis of circulating monocyte activation after SIV infection.

    Fig. S8. Analysis of plasma viral load and SIV Vif–, SIV Gag–, and SIV Pol–specific CD4+ and CD8+ T cell responses in vaccine-protected, necropsied RMs from groups 1 and 2.

    Fig. S9. Analysis of SIV Gag– and SIV Pol–specific CD4+ and CD8+ T cell responses in vaccine-protected and subsequently rechallenged RMs from groups 1, 2, and 4.

    Data file S1. Primary data.

    References (4046)

  • The PDF file includes:

    • Materials and Methods
    • Fig. S1. ΔRh110 RhCMV/SIV vector design and supertope amino acid sequences.
    • Fig. S2. Comparison of the amino acid sequence of SIVmac239 versus SIVsmE660/smE543 vaccine inserts.
    • Fig. S3. Analysis of SIV Env–specific Ab responses after vaccination and after acquisition of SIV infection.
    • Fig. S4. Acquisition of SIV infection by groups 1, 2, and 3 RMs with repeated, limiting-dose IVag SIVmac239 challenge.
    • Fig. S5. Immune correlates analysis.
    • Fig. S6. Analysis of SIV Gag– and SIV Pol–specific CD4+ and CD8+ T cell responses after SIV infection.
    • Fig. S7. Analysis of circulating monocyte activation after SIV infection.
    • Fig. S8. Analysis of plasma viral load and SIV Vif–, SIV Gag–, and SIV Pol–specific CD4+ and CD8+ T cell responses in vaccine-protected, necropsied RMs from groups 1 and 2.
    • Fig. S9. Analysis of SIV Gag– and SIV Pol–specific CD4+ and CD8+ T cell responses in vaccine-protected and subsequently rechallenged RMs from groups 1, 2, and 4.
    • References (4046)

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    Other Supplementary Material for this manuscript includes the following:

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