Fig. 8: Virotherapy selectively induces functional antitumor T cell immunity.
a B16.F10.9CD155-OVA-implanted CD155-tg mice were treated as shown. b IFNγ ELISpot analysis of untreated splenocytes (left), or splenocytes cultured in the presence of SIINFEKL peptide (middle; untreated counts were subtracted). Right panel depicts average spot area (n = 10 for PBS/ mRIPO, n = 11 LPS pooled from 2 experiments); see Supplementary Fig. 11 for extended data. P-values are from Tukey’s post-hoc (two-tailed) test. c Splenocytes from a subset of mice in (b) were tested in 48 h culture alone or with B16.F10.9-OVA cells (right). Supernatant cytokines are plotted for baseline (left) or antigen-specific secretion (right; baseline secretion was subtracted from B16.F10.9.OVA co-culture values). Heatmaps were normalized to maximum value for each cytokine (n = 8 mock/LPS; n = 7 mRIPO); only cytokines secreted above baseline are shown. (*) Tukey’s post-hoc p < 0.05 (two-tailed), baseline mRIPO vs mock (blue asterisks): IFNγ p = 0.03, IL-2 p = 0.004, TNF p = 0.03, IL-5 p = 0.01, IL-6 p = 0.02, IL-10 p = 0.04, IL-13 p = 0.008, IL-17A p = 0.01, IL-22 p = 0.004; baseline LPS vs mock (red asterisk): IL-2 p = 0.03; baseline subtracted mRIPO vs mock (blue asterisks): IFNγ p = 0.01, IL-4 p = 0.01, IL-5 p = 0.047, IL-9 p = 0.01. d Tumor-draining lymph nodes (TDLNs, inguinal lymph node, n = 8 mice/group) from CD155-tg mice bearing wt B16 tumors treated with mock, mRIPO, Poly(I:C), or LPS as in Fig. 8a were harvested 7 days after intratumor treatment. TDLN cells (5 × 105) were co-cultured with carboxyfluorescein succinimidyl ester (CFSE)-labeled B16 or CT2A cells in vitro (3 × 103). CFSE median florescence intensity (left panels) and PD-L1 expression (right panels) were measured by flow cytometry for both cell lines after co-culture (48 h); see Supplementary Fig. 12 for gating and extended analyses; a representative series from three experiments is shown. P-values indicate Tukey’s post-hoc two-tailed test. e, f Wt B16 tumor-bearing mice were treated as in (d) (n = 5/group). Ten days after treatment splenocytes were harvested and pooled by treatment. T cells were isolated by negative selection and adoptively transferred into naïve mice (2 × 106 T cells per mouse) receiving B16 wt tumor cell implants on the same day; mean tumor volume + SEM is shown (f, n = 8 Mock/Poly(I:C), n = 7 mRIPO, n = 6 LPS, n = 4 naïve). P-value is from two-way ANOVA (two-tailed) comparing mice receiving T cells from mRIPO vs mock-treated mice; a representative series from two experiments is shown.
