Chk1 inhibition emerges as the most effective partner for PARP inhibition in BRCA-proficient pancreatic cancer.

PARP inhibitors have shown limited efficacy in pancreatic cancer, partly due to the low prevalence of BRCA mutations, which restricts their clinical use. Because DNA damage checkpoint kinases play critical roles in managing replication stress and DNA damage responses, we investigated whether ATR, Chk1, and WEE1 inhibitors enhance the antitumor activity of olaparib in pancreatic cancer cells, including gemcitabine-resistant clones, and compared these effects with standard chemotherapeutic agents. Cell viability was evaluated in pancreatic cancer cell lines treated with olaparib in combination with various agents using an MTT assay. Apoptosis was assessed by annexin V and propidium iodide double staining followed by flow cytometry. The combinatorial effects were further examined using an in vivo mouse xenograft model. In addition, the effects of combination therapy were evaluated in gemcitabine-resistant cell clones (BxG140 and BxG400) overexpressing ribonucleotide reductase M1 (RRM1). Co-treatment with olaparib and DNA damage checkpoint inhibitors led to a greater reduction in cell viability than combinations with conventional chemotherapeutics. Notably, olaparib combined with the Chk1 inhibitor prexasertib elicited strong synergistic cytotoxicity in BRCA1/2 wild-type MIA PaCa-2 and BxPC-3 cells, whereas the effect was minimal in BRCA2-deficient Capan-1 cells. Compared with monotherapy, combination therapy markedly enhanced apoptosis and suppressed tumor growth in xenograft models. In gemcitabine-resistant clones, olaparib plus prexasertib downregulated RRM1 expression and restored drug sensitivity through enhanced apoptotic cell death. Combined PARP and Chk1 inhibition exerts potent antitumor activity both in vitro and in vivo in pancreatic cancer models, including BRCA-proficient tumors.