Extended Data Fig. 2: In vivo screens identify genes that modify CAR-T cell abundance and transcriptional phenotype.
From: In vivo CRISPR screens identify modifiers of CAR T cell function in myeloma
a. Genes ranked by Log2(fold change) during in vitro manufacturing in IL-7/IL-15 (end-of-production vs. baseline, left panel) and early in vivo (day 7 vs. end-of-production, middle panel) or late in vivo (day 21 vs. end-of-production, right panel). Enriched genes are shown in red and depleted genes are shown in blue with circle size corresponding to –log10 (FDR). n = 2 healthy donor T cells (ND106, ND216). b. Frequency histograms of enrichment or depletion of sgRNAs for known T cell regulatory genes, grouped by respective period. In vitro manufacturing was performed in IL-2. n = 3 healthy donor T cells (ND216, ND99, ND106) c. CD4 and CD8 T cell composition (left panel) and phenotype (middle and right panels) at the beginning of CAR-T cell production (day 0) or at the EOP (day 13) from three healthy donors. Data are presented as mean +/– SEM. Naïve (CD45RA + CCR7 + CD95-), T memory stem cells (TSCM: CD45RA + CCR7 + CD95+), central memory (CM: CD45RA-CCR7+), effector memory (EM: CD45RA-CCR7-), terminally differentiated effector memory (TEMRA: CD45RA + CCR7-) T cells. One-way ANOVA, with Sidak’s multiple comparison test (statistical analysis is calculated between IL-2 and IL-7/IL-15 conditions for each genetic perturbation). CM: intergenic IL2 vs IL7/15 **p = 0.0063, PTPN2 KO IL2 vs IL7/15 **p = 0.0067, CDKN1B KO IL2 vs IL7/15 **p = 0.0072; TEMRA: intergenic IL2 vs IL7/15 ***p = 0.0002, PTPN2 KO IL2 vs IL7/15 ***p = 0.0001, CDKN1B KO IL2 vs IL7/15 ***p = 0.0001. d. Abundance of sgRNAs targeting individual genes across the entire screen workflow for Mario-CAR-T cells produced in IL-7/IL-15.
