Dual inhibition of PARP and ATR induces synthetic lethal vulnerability in prostate cancer with CDK12 deficiency.

Prostate cancers (PCa) harboring CDK12 alterations exhibit aggressive clinical behavior and poor responses to current therapeutic strategies. Although CDK12 is categorized as a homologous recombination repair (HRR)-related gene, clinical evidence indicates limited benefit of poly(ADP-ribose) polymerase (PARP) inhibitor monotherapy in CDK12-defective tumors. To clarify the functional consequences of CDK12 loss and identify a mechanistically rational therapeutic approach, we generated CDK12-knockout (KO) PCa cell lines using CRISPR/Cas9 and assessed cell-cycle regulation, DNA damage repair (DDR), and replication stress by flow cytometry, γH2AX immunofluorescence, and DNA fiber assays. CDK12 knockout abrogated the G0/G1 checkpoint, permitting cell cycle progression into M phase despite the persistence of unrepaired DNA double-strand breaks (DSBs), and reduced ATM transcription, resulting in a DSB response phenotype resembling ATM deficiency. CDK12 loss also increased intrinsic replication stress and sensitized PCa cells to ATR inhibition. In patient-derived xenograft models harboring CDK12 alterations, combined PARP and ATR inhibition led to enhanced DSB accumulation and selectively suppressed tumor growth in CDK12-defective models. These findings highlight the importance of interrogating individual genes within the HRR pathway to define distinct mechanistic vulnerabilities and provide a strong rationale for combined PARP and ATR inhibition as a novel therapeutic strategy for patients with CDK12-altered prostate cancer.