SF3B1 inhibition suppresses prostate cancer by disrupting ATRX splicing and inducing cell cycle arrest

Prostate cancer is the most common malignancy in men in the United States, and dysregulated RNA splicing has emerged as a critical driver of prostate cancer progression. Although alternative splicing pathway genes are mutated in approximately 4% of prostate cancers, mutations in the core splicing factor SF3B1 represent a disproportionately high fraction (27.5%). However, whether and how SF3B1 inhibition suppresses prostate cancer progression remains unclear. Here, we demonstrate that the SF3B1 inhibitor Pladienolide B exerts robust antitumor activity in prostate cancer cell lines, cell-derived xenografts, and patient-derived xenografts. RNA-seq analysis revealed widespread splicing alterations enriched in pathways regulating cell-cycle arrest, which we validated using flow cytometry and live-cell imaging. Following double-thymidine synchronization, Pladienolide B-treated DU145 cells exhibited a markedly prolonged S phase, indicating that SF3B1 inhibition suppresses tumor growth primarily by inducing cell-cycle arrest. To elucidate the molecular basis of this arrest, we examined individual splicing changes and identified a key event in ATRX, which retained a 74-bp cryptic exon before exon 25 upon Pladienolide B treatment. This aberrant splicing disrupted the ATRX-MeCP2 interaction required for maintaining genome stability through repression of R-loop accumulation in heterochromatin. Interestingly, we further identified CDK12 as a previously unrecognized upstream kinase of SF3B1. CDK12 physically interacted with SF3B1, and its inhibition by THZ531 reduced SF3B1 phosphorylation at T313, induced cell-cycle arrest, and suppressed tumor growth. Given the therapeutic potential of SF3B1 inhibition, we evaluated combinatorial strategies with current prostate cancer treatments. Inhibition of SF3B1 activity by THZ531 or Pladienolide B conferred resistance to docetaxel, a first-line therapy for metastatic disease. Notably, Pladienolide B exhibited synergistic antitumor effects when combined with the PARP inhibitor olaparib. Collectively, our findings reveal how SF3B1 inhibition impairs prostate cancer progression through RNA splicing dysregulation, and they provide a rationale for integrating SF3B1-targeted therapies into prostate cancer treatment.