Figure 2 from Targeting TREX1 Induces Innate Immune Response in Drug-Resistant Small-Cell Lung Cancer

TREX1 depletion suppresses SCLC growth and induces IFN response. A, Expression levels of TREX1, IRF3, and p-IRF3 in H69AR and H196 were compared by immunoblotting between cells transduced with Scr and sgTREX1 (#1 and #2). B, Growth curves of H69AR and H196 cells were compared between siCtrl and siTREX1 (#1 and #2; mean ± SEM; n = 3). C, GSEA analysis with H (hallmark) gene sets, based on RNA-seq results of siTREX1 vs. siCtrl H69AR cells. D, Expression levels of IFNB and IFIT1 genes in cells transfected with siCtrl and siTREX1 (#1) were compared by qPCR (mean ± SEM; n = 3). E and F, HLA-A, HLA-B, and HLA-C (E) and PDL1 (F) expressions in H69AR and H196 cells transfected with siCtrl or siTREX1 were compared by flow cytometry analysis. Data are representative of three independent experiments (Left). Mean fluorescence intensity was quantified by FlowJo (Right; n = 3). Cells were analyzed 3 days after siRNA transfection. G, (Left) Schematic of cancer cell–T-cell coculture, performed at a stimulator/responder (S/R) ratio of 1:1. (Right) IL2 secretion from 72 hours cocultures of RPP-A-EtopR (Scr or sgTrex1) or parental RPP-A cells ± 24-hour IFNγ (100 ng/mL) stimulation and no (nil), parental untransduced BW5147.3 (P), or Trav14D-3-DV8; Trbv29 transgenic BW5147.3 (TCR) T cells. BW (−/+) and TCR (−/+) bars represent P or TCR T cells alone without (−) or with (+) PMA (50 nmol/L) and ionomycin (1 μg/mL). ^, above assay threshold of detection (mean ± SEM; n = 4). Data represent mean ± SEM. ns, not significant; *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001 by unpaired Student t test (D, E and F), one-way ANOVA followed by Dunnett’s multiple comparisons test (B) and two-way ANOVA followed by Tukey’s multiple comparisons test (G).