Extended Data Fig. 10: Knockout of TRAIL in 1928ζ CAR-T cells significantly impairs antitumor cytolytic activity.
From: CAR-engineered lymphocyte persistence is governed by a FAS ligand–FAS autoregulatory circuit
(a) RNA-seq values for CD19, CD3E, TNFRSF10A (the gene encoding DR4) and TNFRSF10B (the gene encoding DR5) in Nalm6, Raji, and JVM2 B cell malignancy cell lines. (b) Scatter-plot of TNFRSF10A and TNFRSF10B RNA-seq values from n = 83 B cell lines featured in the Cancer Cell Line Encyclopedia (CCLE). Horizontal line represents the median value while the vertical bars represent the interquartile range. P value calculated using an unpaired two-tailed Student’s t-test. TPM = transcript per million. (c) Schematic for the CRISPR/Cas9-mediated knockout (KO) of TRAIL in human CD8+ T cells expressing a 1928ζ CAR. (d) Cytolytic activity of TRAIL KO versus wild-type (WT)-TRAIL 1928ζ CAR-T cells or T cells transduced with tEGFR alone against Nalm6/NLS–mCherry at indicated effector-to-target (E:T) ratios. Data shown as mean ± s.e.m. (n = 3 biologically independent samples). Statistical comparisons performed using a one-way ANOVA. ns = not significant, P > 0.05. (e) Table displaying the measured frameshift insertion-deletion (Indel) frequency in TRAIL for each T cell group used in the cytolytic assay. Data shown as mean ± s.e.m. (n = 3 technical replicates from 1 experiment, repeated with similar results using n = 2 donors).
