The core spliceosomal Sm proteins were recently proposed as cancer-selective lethal targets in non-small cell lung cancer (NSCLC). In contrast, the loss of the commonly mutated cancer target SF3B1... Show moreThe core spliceosomal Sm proteins were recently proposed as cancer-selective lethal targets in non-small cell lung cancer (NSCLC). In contrast, the loss of the commonly mutated cancer target SF3B1 appeared to be toxic to non-malignant cells as well. In the current study, the transcriptomes of A549 NSCLC cells, in which SF3B1 or SNRPD3 was silenced, were compared using RNA sequencing. The skipping of exon 4 of the proteasomal subunit beta type-3 (PSMB3) mRNA, resulting in a shorter PSMB3-S variant, occurred only after silencing SNRPD3. This observation was extended to the other six Sm genes. Remarkably, the alternative splicing of PSMB3 mRNA upon Sm gene silencing was not observed in non-malignant IMR-90 lung fibroblasts. Furthermore, PSMB3 was found to be overexpressed in NSCLC clinical samples and PSMB3 expression correlated with Sm gene expression. Moreover, a high PSMB3 expression corresponds to worse survival in patients with lung adenocarcinomas. Finally, silencing the canonical full-length PSMB3-L, but not the shorter PSMB3-S variant, was cytotoxic and was accompanied by a decrease in proteasomal activity. Together, silencing Sm genes, but not SF3B1, causes a cytotoxic alternative splicing switch in the PSMB3 mRNA in NSCLC cells only. Show less
Okonska, A.; Buhler, S.; Rao, V.; Ronner, M.; Blijlevens, M.; Meulen-Muileman, I.H. van der; ... ; Felley-Bosco, E. 2020
Loss of function of BRCA1-associated protein 1 (BAP1) is observed in about 50% of malignant pleural mesothelioma (MPM) cases. The aim of this study was to investigate whether this aspect could be... Show moreLoss of function of BRCA1-associated protein 1 (BAP1) is observed in about 50% of malignant pleural mesothelioma (MPM) cases. The aim of this study was to investigate whether this aspect could be exploited for targeted therapy. A genetically engineered model was established expressing either functional or nonfunctional BAP1, and whole-genome siRNA synthetic lethality screens were performed assessing differentially impaired survival between the two cell lines. The whole-genome siRNA screen unexpectedly revealed 11 hits (FDR < 0.05) that were more cytotoxic to BAP1-proficient cells. Two actionable targets, ribonucleotide reductase (RNR) catalytic subunit M1 (RRM1) and RNR regulatory subunit M2 (RRM2), were validated. In line with the screen results, primary mesothelioma (BAP1(+/-))overexpressing BAP1 C91A (catalytically dead mutant) was more resistant to RNR inhibition, while BAP1 knockdown in the BAP1-proficient cell lines rescued the cells from their vulnerability to RNR depletion. Gemcitabine and hydroxyurea were more cytotoxic in BAP1-proficient cell line-derived spheroids compared with BAP1 deficient. Upregulation of RRM2 upon gemcitabine and hydroxyurea treatment was more profound in BAP1 mut/del cell lines. Increased lethality mediated by RNR inhibition was observed in NCI-H2452 cells reconstituted with BAP1-WT but not with BAP1 C91A. Upregulation of RRM2 in NCI-H2452-BAP1 WT spheroids was modest compared with control or C91A mutant. Together, we found that BAP1 is involved in the regulation of RNR levels during replication stress. Our observations reveal a potential clinical application where BAP1 status could serve as predictive or stratification biomarker for RNR inhibition-based therapy in MPM. Show less
