Comparative Activity and Off-Target Effects in Cells of the CHK1 Inhibitors MK-8776, SRA737, and LY2606368.

DNA damage activates the checkpoint protein CHK1 to arrest cell cycle progression, providing time for repair and recovery. Consequently, inhibitors of CHK1 (CHK1i) enhance damage-induced cell death. Additionally, CHK1i elicits single agent cytotoxicity in some cell lines. We compared three CHK1i that have undergone clinical trials and exhibited different toxicities. Each CHK1i inhibits other targets at higher concentrations, and whether these contribute to the toxicity is unknown. We compared their sensitivity in a panel of cell lines, their efficacy at inhibiting CHK1 and CHK2, and their ability to induce DNA damage and abrogate damage-induced S phase arrest. Published in vitro kinase analyses were a poor predictor of selectivity and potency in cells. LY2606368 was far more potent at inhibiting CHK1 and inducing growth arrest, while all three CHK1i inhibited CHK2 at concentrations 10- (MK-8776 and SRA737) to 100- (LY2606368) fold higher. MK-8776 and SRA737 exhibited similar off-target effects: higher concentrations demonstrated transient protection from growth inhibition, circumvented DNA damage, and prevented checkpoint abrogation, possibly due to inhibition of CDK2. Acquired resistance to LY2606368 resulted in limited cross-resistance to other CHK1i. LY2606368-resistant cells still abrogated DNA damage-induced S phase arrest, which requires low CDK2 activity, whereas inappropriately high CDK2 activity is responsible for sensitivity to CHK1i alone. All three CHK1i inhibited protein synthesis in a sensitive cell line correlating with cell death, whereas resistant cells failed to inhibit protein synthesis and underwent transient cytostasis. LY2606368 appears to be the most selective CHK1i, suggesting that further clinical development of this drug is warranted.