Figure 6 from Targeting PRMT5 Inhibitor–Induced Adaptation in Pancreatic Cancer with the RBM39 Degrader Indisulam

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PRMT5 and RBM39 inhibition regulate DNA repair and metabolic pathways. A, Workflow of the RNA-seq approach: PSN1 cells were treated with DMSO as the vehicle control, 4 nmol/L JNJ-64619178, 200 nmol/L indisulam, or the combination of 4 nmol/L JNJ-64619178 and 200 nmol/L indisulam for 48 hours, mRNA was isolated, and mRNA sequencing with subsequent GSEA was performed. The experiment was conducted as four biological replicates. B, Venn diagram showing downregulated GO-BP gene sets in PSN1 cells; all treatments were compared with the vehicle control in individual GSEAs. Only gene sets with an FDR < 0.05 were considered significant and used for the analysis. C, 24 GO-BP gene sets downregulated exclusively in the combination treatment are indicated. The value of the normalized enrichment score (NES) is color-coded. D, Number and composition of significant changed alternative splicing (AS) events observed upon monotherapy or combination treatment of JNJ-64619178 and/or indisulam. E, Percent spliced-in (ΔPSI) distribution of different AS events identified following monotherapy or combination treatment with JNJ-64619178 and/or indisulam. P values based on the Wilcoxon test are indicated. F, PSN1 cells were either treated with DMSO control or 4 nmol/L JNJ-64619178 and 200 nmol/L indisulam for 96 hours followed by mass spectrometry determination of the proteome. G, Volcano plot of deregulated proteins in PSN1 cells treated with the combination therapy [see (F)] vs. control. Proteins with an adjusted P value < 0.05 were colored. Proteins involved in the DNA repair pathway are highlighted lilac, whereas proteins involved in the splicing pathway are highlighted pink. H, ΔPSI distribution of different AS events in the DNA damage signaling and repair pathway genes ATM, FAN1, and FANCL observed upon monotherapy or the combination treatment with JNJ-64619178 and/or indisulam. I, Results of the γH2A.X phosphorylation assay: PSN1 cells were treated with 50 nmol/L MRTX1719 and/or 240 nmol/L indisulam for 48 hours (left) or 72 hours (right). For negative control, cells were left vehicle-treated; for positive control, cells were irradiated with 2 Gy for 30 minutes. The fold increase of γH2A.X phosphorylation is shown on the y-axis. Statistical analysis was performed using a repeated-measures one-way ANOVA with correction for multiple-testing according to the Tukey test: **, P < 0.01; *, P < 0.05, n = 3. A3SS, alternative 3′ splice site; A5SS, alternative 5′ splice site; MSE, mutually exclusive exon; RI, retained intron; SI, skipped exon. [A, Created in BioRender. Schneider, G. (2026) https://BioRender.com/fwl2ir3; F, Created in BioRender. Schneider, G. (2026) https://BioRender.com/z84e701.]

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