Extended Data Fig. 5: Transgene expression profiles of shortlisted target genes in human anti-LeY CAR T cells.
From: Rewiring endogenous genes in CAR T cells for tumour-restricted payload delivery
a-f. Human anti-LeY CAR T cells were engineered to express Gfp from the indicated gene loci and cocultured with OVCAR-3 (a-b) or MCF-7 (c-f) tumor cells for 72 h before GFP expression in CD4+ (a-b, e-f) or CD8+ (c-d) CAR T cells was assessed. a, c, e. Quantification of GFP percentage and b, d, f. fold change in GFP MFI of human CAR T cells following coculture. (a, c, e) Data represent mean ± SD of technical triplicates from n = 3 (CLU, DUSP4, RGS1, RGS2, TNFAIP3), 4 (PD-1) and 8 (NR4A2, RGS16) donors. (b, d, f) Data represent mean ± SEM pooled from n = 3 (CLU, DUSP4, RGS1, RGS2, TNFAIP3), 4 (PD-1) and 8 (NR4A2, RGS16) donors. g-h. Human anti-LeY CAR T cells engineered as per (a-f) were adoptively transferred into mice bearing subcutaneous OVCAR-3 tumors. 14 days later, tumors, spleens and livers were harvested and GFP expression by CD8+ human CAR T cells was assessed. g. Flow cytometry plots from concatenated samples of n = 4 mice/group, representative of n = 2 donors. h. Quantification of GFP percentage of CD8+ human CAR T cells in the tumor, spleen and liver. Data represent mean ± SEM from n = 8 mice/group pooled from n = 2 donors. i. Human anti-LeY CAR T cells engineered as per (a-f) were cocultured with OVCAR-3 tumor cells for 72 h. Concatenated flow cytometry plots showing CAR and GFP expression in CD8+ T cells from technical triplicates, representative of n = 8 donors. (a, c, e) Paired student’s t-test. (b, d, f) One-way ANOVA. (h) Two-way ANOVA. n.s. not significant, *p < 0.05, **p < 0.01, ****p < 0.0001.
