Figure 5.

PD1⁺CD4⁺ memory TILs show features of T_FH-like cells and are poised to help B cells. Heatmap with unsupervised clustering in (a), and ssGSEA scores in (b), showing enrichment of a 4PD-1^hi TIL immunosuppressive signature based on a published dataset by Zappasodi et al.³⁰ and T_FH gene signature by Gu-Trantien et al.³¹ in CD4 TILs separated based on genotype (tumor versus normal) and PD1 status (PD1⁺ and PD1⁻). Each column in a represents a patient. (c) Representative panels illustrating localization of CD4/PD-1 in relation to CD19⁺ B cells (green) in tumor via confocal imaging. (d) Histograms and proliferation index of carboxyfluorescein succinimidyl ester (CFSE)-labeled CD19⁺ B cells stimulated (Stim) with staphylococcal enterotoxin B (SEB, 500 ng/ml) alone or SEB cocultured with PD1⁺CD4 TILs (n = 4–6, biological replicates). (e) Histograms and frequency of CD38/CD27 subsets in CD19⁺ B cells after stimulation with SEB (500 ng/ml) alone or SEB in presence of PD1+ CD4 TILs (n = 13, biological replicates). (f) Histograms and frequency of CXCR5⁺CCR7⁺ expression in stimulated CD19⁺ B cells. (g) Frequencies of CD38/CD27 subsets for expression of CD24/CD25 stimulated with SEB in the presence of PD1⁺CD4 TILs. (h) Histograms and change in PD1 expression in CD4 TILs following stimulation with SEB alone or SEB in presence of CD19⁺ B cells. (i and j) Histograms (i) and frequencies (j) of PD1⁺CD200⁺ cells as a % of CD4 TILs stimulated with SEB alone or SEB in the presence of CD19⁺ B cells (n = 6, biological replicates). (k) gMFI of degranulation marker CD107a in PD1⁺CD200⁺ CD4 TILs co-expressing CD25 and CTLA4 (n = 6, biological replicates). (l) Histogram and frequencies of CXCR5⁺CCR7⁺ CD4 TILs stimulated with SEB alone or SEB in the presence of CD19 B cells. Data are displayed as mean ± SD. Data in d, e, f, and k by paired Students t-test, two-tailed. Data in h, j and l by one-way ANOVA followed by post-hoc Newman-Keuls. ns, not significant.