Fig. 1: Enhanced CD8⁺ T cell proliferation and effector function after temporal cell cycle inhibition.

a, Experimental setup. Experiments were performed with isolated CD8⁺ T cells derived from peripheral blood mononuclear cells from healthy donors or from splenocytes of naive mice. b, Schematic overview of cell cycle inhibition with different cell cycle inhibitors. c, Representative proliferation plots of unstimulated human CD8⁺ T cells, or ex vivo-stimulated human CD8⁺ T cells that were left untreated (non-arrested), arrested in the cell cycle using RO-3306 or ribociclib (arrested) or released from cell cycle arrest (released). d, Representative proliferation plots of unstimulated and ex vivo-stimulated human and mouse CD8⁺ T cells under the same conditions as in c, treated with HU. e, Percentage of proliferating cells and division index (mean ± s.e.m.) of ex vivo-stimulated human (n = 8) and mouse (n = 7) CD8⁺ T cells that were left untreated (non-arrested) or released after HU arrest. f, Expression (geometric mean fluorescence intensity (gMFI) ± s.e.m.) of ID2 (left, n = 4 mice) and granzyme B (right, n = 6 mice) in unstimulated and ex vivo-stimulated mouse CD8⁺ T cells that were left untreated (non-arrested), arrested with HU or released from HU-induced arrest. g, Representative histograms of CD69, CD62L, CD127, CXCR3, EOMES, PD-1 and LAG-3 expression in ex vivo-stimulated mouse CD8⁺ T cells treated temporally with HU or left untreated. h, Naive mice were vaccinated with E7 SLP/CpG on day 0 and treated with HU for 4 consecutive days (n = 5) or left untreated (n = 6). Left, percentages of circulating E7_49–57-specific CD8⁺ T cells over time (mean ± s.e.m.). Middle, normalized response relative to the peak. Right, percentages of KLRG1⁺CD44⁺ CD8⁺ T cells over time (mean ± s.e.m.). i, Slope of the E7_49–57-specific CD8⁺ T cell response depicted in h. Statistical comparisons were performed with two-sided paired t-test (e), repeated-measures analysis of variance (ANOVA) with Sidak’s multiple-comparison test (f) or two-sided unpaired t-test (h and i); P values are shown on the graphs.