Abstract

Inhibition of the checkpoint kinase Chk1, both as a monotherapy and in combination with DNA damaging cytotoxics, is a promising therapeutic strategy for cancer. However, much remains to be learned in regard to the patient populations that will respond best to a Chk1 inhibitor and the optimal therapeutics to combine with a Chk1 inhibitor. In an effort to discover sensitizing mutations and novel combination strategies for Chk1 inhibition, we performed a ‘synthetic lethality’ siRNA screen with the selective Chk1 inhibitor Chk1-A. This screen employed a custom made library of siRNAs against 197 genes (3 siRNAs per gene), most of which are involved in cell-cycle control or DNA damage repair. One of the most prominent and consistent hits across runs of the screen performed in PC3, LNCaP, and A549 cell lines was Wee1 kinase. MK- 1775 is a small molecule inhibitor of Wee1 that is currently in early stage clinical trials. In confirmation of the results obtained from the siRNA screen, we found that Chk1-A and MK-1775 synergistically inhibited proliferation in multiple cell types. This anti- proliferative synergy correlated with a synergistic induction of apoptosis. We explored the mechanism of the impressive synergy by examining the cellular and biochemical effects of the Chk1-A and MK-1775 combination. We found that co-treatment with the two inhibitors resulted in dramatic decreases in inhibitory phosphorylation of cyclin- dependent kinases 1 and 2, increases in DNA damage, and the collapse of DNA replication. In conclusion, the combination of a Chk1 inhibitor and a Wee1 inhibitor may be an effective treatment strategy for cancer.