Extended Data Fig. 9: PD-1⁺TCF1⁺ stem-like CD8⁺ T cells proliferate and differentiate into effector CD8⁺ T cells expressing the high affinity trimeric (CD25, CD122, CD132) IL-2 receptor after PD-1 + IL-2 combination therapy.

a, Experimental setup for panels b–d. Stem-like (PD-1⁺CXCR5⁺Tim-3⁻) and exhausted (PD-1⁺CXCR5⁻TIM3⁺) CD8⁺ T-cell subsets were sorted from the spleens of LCMV chronically infected CD45.2⁺ mice and each subset was transferred into infection-matched CD45.1⁺ recipient mice. Groups of these mice were then either left untreated, given anti-PD-L1 antibody, IL-2 therapy, or combination therapy for 2 weeks. CD25 expression on donor CD45.2⁺ CD8⁺ T cells was checked before and after the treatments. b, Representative histogram of CD25 expression on the chronic CD8⁺ T-cell subsets pre-transfer. Naive (CD44^lo) CD8⁺ T cells are also shown as a negative control. c, d, Representative FACS plots of CD25 expression and summary data of frequency of CD25⁺ cells in donor CD45.2⁺ CD8⁺ T cells originating from stem-like or exhausted CD8⁺ T cells after the indicated treatments. e, Experimental setup for panels f–o. LCMV chronically infected mice were treated with anti-PD-L1 antibody, IL-2 alone, or combination therapy. Mice were sacrificed on the indicated days and expression of CD25, CD122 and CD132 was examined on LCMV-specific CD8⁺ T cells in the spleen. f, Representative flow plots for the co-expression of CD25 and Ki-67 on D^bGP33⁺ CD8⁺ T cells at day 0 or day 6 after treatment. g, j, m, Representative histograms showing the expression of CD25 (g), CD122 (j), and CD132 (m) on stem-like and exhausted LCMV-specific DbGP33⁺ subsets CD8⁺ T cells before starting the treatment of LCMV chronically infected mice. Naive cells are CD44^lo CD8⁺ T cells present in the same host. h, k, n, Representative histograms showing the expression of CD25 (h), CD122 (k), and CD132 (n) on D^bGP33⁺ CD8⁺ T cells at days 0-6 after starting the indicated treatment. i, l, o, Summary box plots for the frequency of CD25⁺ cells (i), MFI of CD122 (l) and MFI of CD132 (o) on D^bGP33⁺ CD8⁺ T cells after the indicated treatments. Results were pooled from 2-5 experiments with at least 4 mice per group (a–o). Data are presented as mean and SD (d) or the box (25th to 75th percentiles), the whiskers (min to max), and the line (the median) (i, l, o) with p values. Statistical comparisons were performed using one-way ANOVA with Tukey’s multiple-comparison test. Untx, untreated.

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