Extended Data Fig. 1: Transient cell cycle inhibition enhances CD8⁺ T cell proliferation.

a, Representative proliferation plots of unstimulated mouse CD8⁺ T cells, or ex vivo-stimulated mouse CD8⁺ T cells that were either left untreated (non-arrested), arrested in the cell cycle using RO-3306, or released from RO-3306-induced arrest. b, Percentage of viable CD8⁺ T cells treated or not with HU and RO-3306, measured as Zombie-NIR⁻ cells within the singlet gate (n=5). Statistical comparisons were performed using RM ANOVA with Sidak’s multiple comparisons; P values are shown on the graphs. c-e, Mouse CD8⁺ T cells were left unstimulated or ex vivo-stimulated and either non-arrested, cell cycle-arrested with HU or RO-3306, or released from arrest (n=5). c, Contour plots showing the distribution of cells from each condition. d, Expression levels of the indicated markers overlaid on the opt-SNE map. e, CFSE intensity overlaid on the opt-SNE embedding, where black indicates CFSE^low (highly proliferating cells), and copper indicated CFSE^high (undivided/first cycle). f, Representative FSC/SSC plots of human and mouse CD8⁺ T cells that were unstimulated, cell cycle arrested with HU, non-arrested or released from arrest. Percentage of blasted cells (large, activated cells with increased FSC/SSC) is indicated. g, Representative microscopy images of ex vivo-stimulated mouse CD8⁺ T cells treated transiently with HU or left untreated (3 independent experiments with similar results). Representative FSC/SSC plots of human CD8⁺ T cells that were unstimulated, cell cycle arrested with RO-3306, topotecan, palbociclib or ribociclib, non-arrested or released from arrest. Percentage of blasted cells is indicated.