Extended Data Fig. 7: Combined blockade increases the ratio of cell-associated SIV-RNA relative to SIV-DNA content in PBMCs and limit intact provirus in LN CD4+ T cells. | Nature Medicine

Extended Data Fig. 7: Combined blockade increases the ratio of cell-associated SIV-RNA relative to SIV-DNA content in PBMCs and limit intact provirus in LN CD4+ T cells.

From: CTLA-4 and PD-1 dual blockade induces SIV reactivation without control of rebound after antiretroviral therapy interruption

Extended Data Fig. 7

(a) The number of log-transformed cell-associated SIV-DNA copies per 106 PBMCs were quantified by 12 replicate reaction RT-qPCR prior to the first infusion (d0 p.t.), and 1 week following the first (d7 p.t.) and fourth infusions (d28 p.t.); from which (b) the ratio of SIV-RNA to -DNA copies was computed. All RMs are color-coded and grouped based on ICB therapy as follows with population sizes as indicated for all RT-qPCR analyses: controls (n=6), black; αCTLA-4 (n=6), blue; αPD-1 (n=6), pink; combined blockade (n=7), red; and BsAb (n=8), purple. By IPDA with ddPCR, (c) the DNA shearing index pre- and post-ICB, and (d) the fold change in intact provirus per 106 cells relative to pre-treatment baseline were quantified in LN CD4+ cells. For all IPDA analyses, population sizes are as follows: control (n=3), αCTLA-4 (n=6), αPD-1 (n=5), combined blockade (n=7), and BsAb (n=7). (e) An example IPDA plot is given for the amplification of the env relative response element (RRE) on VIC dye against pol on FAM dye in relative fluorescence units (RFU). A positivity cutoff of 1250 is used for env and 2000 for pol. Sample plots shown (as indicated in red) are as follows: negative control, merged analysis from no-template control and PBMCs at pre-infection from RLt16; env positive control, env gBlock (IDT); pol positive control, pol gBlock (IDT); and IPDA (shown are LN CD4+ cells at d-29 p.t. from RWs16; 1 of 58 image sets of 8 replicate reactions). Averaged data are presented as the mean ± s.e.m. Each data point represents an individual animal, as indicated by shape, and those with ICB-related viral reactivation in plasma are represented as closed data points. Data were analyzed with a two-sided (a,b) Wilcoxon matched-pairs signed rank test, (d) a Mann-Whitney U test, or (c) a two-way ANOVA with Bonferroni’s correction for multiple comparisons relative to baseline and controls.

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