Inhibition of Chk1 with the small molecule inhibitor V158411 induces DNA damage and cell death in an unperturbed S-phase

Abstract
Chk1 kinase plays a crucial role in the DNA damage response checkpoint, and inhibitors targeting Chk1 are currently being evaluated in clinical trials. Chk1 helps mitigate oncogene-induced replication stress, with Chk1 inhibitors showing efficacy as a monotherapy across various cancer types. Understanding the mechanism by which Chk1 inhibitors promote DNA damage and cancer cell death is critical for advancing their clinical use. In this study, we examine how the novel Chk1 inhibitor V158411 increases DNA damage and cell death in human cancer models. V158411 caused a concentration- and time-dependent rise in γH2AX-positive nuclei, specifically in cells actively undergoing DNA synthesis. The induction of γH2AX occurred early and was linked to the activation of the ATR/ATM/DNA-PKcs DNA damage response pathways. Notably, the appearance of γH2AX-positive nuclei preceded the detection of ssDNA and RPA depletion. Full and sustained Chk1 kinase inhibition was required for robust γH2AX induction and growth suppression. Chk1 inhibitor-induced cytotoxicity was associated with DNA damage, apoptosis, mitotic slippage, and permanent cell cycle arrest. We identified two distinct categories of Chk1 inhibitors: those that strongly increased γH2AX, pChk1 (S317), and pRPA32 (S4/S8) (including V158411, LY2603618, and ARRY-1A), and those that did not (such as MK-8776 and GNE-900). Tumor cell death, driven by heightened DNA damage and the disruption of cell cycle checkpoints, suggests that selective Chk1 inhibitors could be a promising therapeutic option for treating various human cancers.