Extended Data Fig. 8: Ablation of RASA2 preserves cancer cell killing capacity in T cells after repeated target cancer cell exposures.
From: RASA2 ablation in T cells boosts antigen sensitivity and long-term function
a, Cancer cell killing assay showed control-edited NY-ESO-1-specific TCR-T cells failed to control cancer cell expansion after multiple stimulations, whereas RASA2 ablation led to persistent killing capability. Cancer cell kill after 1, 3, and 6 repeated co-cultures (columns) and for a range of effector T cells to target cancer cell ratios (rows) are displayed over time (each time point in triplicates, lines are mean ± SD, lines show a fitted curve by a generalized additive model, see methods). b, Representative flow cytometry histograms showing CD19 staining in A375 cells engineered to express CD19 (A375-CD19 in cyan) compared to unperturbed A375 cells (A375-WT in red). c, Summary statistics for area under the growth curve over target A375-CD19 cells a range of effector T cells to target cell ratios after 1 and 6 repeated cancer cell exposures (n = 3 donors in duplicates, mean ± SEM, ***p < 0.001 for two-sided Wilcoxon test). d, Representative data for cancer killing assays in (c) by CD19-specific CAR-T cells after 1 and 6 repeated co-cultures for a range of effector T cells to target cancer cell ratios (rows) with target A375-CD19 melanoma cells expressing CD19. e, Representative data from one CAR-T cell donor for cancer killing assays with target Nalm6 leukemia cells (each time point in triplicates, lines are mean ± SD). f, Traces of cancer cell count over time as detected by live-cell microscopy. CD19 CAR-T cells from two donors after repetitive stimulation were co-cultured with either unperturbed target cells (A375-WT) or an isogenic cell line engineered to express CD19 (A375-CD19). No cancer cell killing is observed with either RASA2 KO or CTRL T cells in the antigen-negative condition. Lines are mean of 3 technical replicates. g, Quantification of data in (f), for the area under the growth curve (y-axis), across multiple effector T cell to target cancer cell ratios (x-axis) (n = 2 donors in triplicates, mean ± SEM, shape denotes donor).
