Fig 2: SWI/SNF-deficient cancer cells are sensitive to GSH inhibition. (a). Immunoblot analysis of SMARCA4, SMARCB1, PBRM1, SLC7A11, and β-actin expression in a cancer cell line panel. The ratio of the signal intensity of SLC7A11 protein relative to that of β-actin protein was calculated. (b). IC50 values for eprenetapopt (APR-246) in cancer cell lines derived from SWI/SNF-proficient cell lines (PC9, H2122, H2228, HCC44) and SMARCA4-deficient (H1819, KP-4, H1703, H522), SMARCB1-deficient (JMU-RTK-2, G401, G402, HS-ES-1), or PBRM1-deficient (KMRC-1, RCC-MF) cell lines after treatment for 6 days. Data are presented as the mean ± SEM (standard error of the mean), n = 3 independent experiments. p values were determined by an unpaired two-tailed Student’s t-test. *p < 0.05, ***p < 0.001.

Fig 3: SMARCA4/SMARCA2 deficiency and SS18–SSX fusion lead to upregulation of KREMEN2 gene expression. A, Expression of KREMEN2 mRNA in SWI/SNF-proficient, SMARCA4/SMARCA2-deficient, and SS18–SSX fusion cell lines (relative to that in DMS114 cells). Data are presented as the mean ± SEM; n = 3 independent experiments. B, Violin plots showing clustered expression of KREMEN2 mRNA in SWI/SNF-WT, SMARCB1-deficient, ARID1A-deficient, PBRM1-deficient, SMARCA4-deficient, SMARCA4/SMARCA2-deficient, and SS18–SSX fusion cell lines based on an analysis of mutations, gene CN, and gene expression data from DepMap (data version 23Q2). SWI/SNF-proficient (n = 1,238), SMARCB1-deficient (n = 19), ARID1A-deficient (n = 52), PBRM1-deficient (n = 23), SMARCA4-deficient (n = 34), SMARCA2-deficient (n = 27), SMARCA4/SMARCA2-deficient (n = 6), and SS18–SSX (n = 5) cell lines were analyzed, and data are presented as the mean ± SEM. n indicates the number of independent samples. C, Localization of signals generated by SMARCA4 and H3K27ac ChIP-seq, ATAC-seq, and RNA-seq around the KREMEN2 locus in SMARCA4/SMARCA2-deficient BIN-67 cells transduced with an SMARCA4 expression vector (+SMARCA4) and in BIN-67 cells without the SMARCA4 expression vector (−SMARCA4). D, Localization of signals generated by SS18, SMARCA4, RNAPII, and SUZ12 ChIP-seq, ATAC-seq, and RNA-seq around the KREMEN2 locus in SS18–SSX-fusion Aska-SS cells depleted of SS18–SSX by SSX-targeting shRNA (shSSX; SS18-WT) and in SS18–SSX fusion Aska-SS cells not depleted of SS18–SSX [i.e., transduced with NT shRNA (shNT; SS18–SSX)].

Fig 4: Damaging Mutations in PBRM1 are associated with MCL1 dependency and sensitivity to MCL inhibition in ccRCC. (A) MCL-1 gene dependency analysis in ccRCC cell lines with damaging PBRM1 mutations compared to WT. Dependency scores (Chronos, 22Q2) were retrieved from DepMap (https://depmap.org) (B). PBRM1 mRNA levels in ccRCC cell lines inversely correlate with MCL1 dependency. *P<0.05, versus WT by t-test. (C) Waterfall plot showing half maximal inhibitory concentration (IC50) for PRT1419 in the ccRCC cell line panel. (D) Cell Titer-Glo (CTG) assay assessing inhibition of 3D spheroid growth in PBRM1-mut and WT ccRCC cell lines following treatment with MCL-1 inhibitor PRT1419 (72h). Values shown are cell viability calculated as percentage of vehicle DMSO control. (E) Caspase 3/7 activity assay showing induction of apoptosis following 4h treatment with PRT1419 in PBRM1-mut and WT lines. (F) CTG assay assessing inhibition of 3D cell growth by PRT1419 in 769-P (PBRM1 WT) cells following PBRM1 depletion via RNAi. (G) Caspase 3/7 activity assays assessing induction of apoptosis by PRT1419 in 769-P (PBRM1 WT) cells following PBRM1 depletion via RNAi. (H) Western blot validating PBRM1 expression following RNAi knockdown in 769-P cells (I) OSRC-2 (PBRM1 mutant ccRCC) cell line-derived xenograft (CDX) assessing anti-tumor activity of intravenously administered PRT1419 at 20 mg/kg. Animals were dosed once weekly with PRT1419. Data represented as mean ± SEM. N=8, * P<0.05 by Mann-Whitney U test. (J) Representative images of tumors from mice in both vehicle and PRT1419-treated groups at the end of the study.

Fig 5: cBAF- and PBAF-deficient cancer cells show low basal levels of GSH due to attenuated expression of SLC7A11. (a). Relative expression of SLC7A11 mRNA in SWI/SNF-proficient cell lines (H2122, H2228, PC9) and SMARCA4-deficient (H1703, H1819), SMARCB1-deficient (JMU-RTK-2, G402), or PBRM1-deficient (KMRC-1, RCC-MF) cell lines (relative to that in the H2122 cell line). Data are presented as SEM (the mean ± standard error of the mean), n = 3 independent experiments. (b). Basal levels of GSH in SWI/SNF-proficient cell lines (H2122, H2228, PC9) and SMARCA4-deficient (H1703, H1819), SMARCB1-deficient (JMU-RTK-2, G402), or PBRM1-deficient (RCC-MF, KMRC-1) cell lines (relative to that in the H2122 cell line). Data are presented as SEM, n = 3 independent experiments. For all experiments, p values were determined by an unpaired two-tailed Student’s t-test. **p < 0.01, ***p < 0.001.