Abstract
The DNA-damage-activated checkpoint protein CHK1 is required to prevent replication or mitosis in the presence of unrepaired DNA damage. Inhibitors of CHK1 (CHK1i) circumvent this checkpoint and enhance cell killing by DNA-damaging drugs. CHK1i also elicit single-agent cytotoxicity in a small subset of cell lines. Resolving the mechanisms underlying the single-agent activity may permit patient stratification and targeted therapy against sensitive tumors. Our recent comparison of three CHK1i demonstrated that they all inhibited protein synthesis only in sensitive cells. LY2606368, the most selective of these CHK1i, was used in the current study. Comparison across a panel of cell lines demonstrated that sensitive cells died upon incubation with LY2606368, whereas resistant cells underwent growth inhibition and/or cytostasis but failed to die. Sensitive cells exhibited inhibition of protein synthesis, elevated DNA damage, impaired DNA repair, and subsequently death. The consequence of CHK1 inhibition involved activation of cyclin A/CDK2 and MUS81, resulting in DNA damage. This damage led to activation of AMPK, dephosphorylation of 4E-BP1, and inhibition of protein synthesis. Inhibition of MUS81 prevented activation of AMPK, while inhibition of AMPK enhanced DNA repair and cell survival. The activation of AMPK may involve a combination of LKB1 and CaMKKβ. This study raises questions concerning the potential importance of the inhibition of protein synthesis in response to other drugs, alone or in combination with CHK1i. It also highlights the importance of clearly discriminating among growth inhibition, cytostasis, and cell death, as only the latter is likely to result in tumor regression.