Endogenous replication stress creates a unique therapeutic window for ALC1 targeting.

Poly (ADP-ribose) polymerase inhibitors (PARPi) are approved for homologous recombination-deficient (HRD) high-grade serous ovarian cancers. However, their durable benefit is severely limited by resistance and hematologic toxicity. PARPi are also largely ineffective in homologous recombination-proficient tumors, especially CCNE1-amplified cancers, which remain therapeutically intractable. We show that loss of the chromatin remodeler Amplified in Liver Cancer 1 (ALC1) markedly sensitizes serous ovarian cancer cells, xenografts, and patient-derived models to PARPi across both HRD and CCNE1-amplified contexts. Unlike other PARPi sensitizers, ALC1 depletion circumvents clinically relevant PARPi resistance while sparing BRCA wild-type and heterozygous nonmalignant cells, revealing an unusually wide therapeutic index. Mechanistically, this selectivity is dictated by intrinsic replication-stress burden, which renders cancer cells uniquely dependent on ALC1-mediated stress tolerance. Accordingly, collective data from cell line and patient samples reveal endogenous phospho-T21 RPA2 as a predictive biomarker of PARPi responsiveness following ALC1 targeting. These findings establish ALC1 targeting as a mechanistically grounded, biomarker-guided strategy to safely expand PARPi efficacy and support the clinical advancement of ALC1 inhibitors currently in Phase I trials.