Inhibition of Chk1 with the small molecule inhibitor V158411 induces DNA damage and cell death in an unperturbed S-phase.
Chk1 kinase is a critical component of the DNA damage response checkpoint and Chk1 inhibitors are currently under clinical investigation. Chk1 suppresses oncogene-induced replication stress with Chk1 inhibitors demonstrating activity as a monotherapy in numerous cancer types. Understanding the mechanism by which Chk1 inhibitors induce DNA damage and cancer cell death is essential for their future clinical development. Here we characterize the mechanism by which the novel Chk1 inhibitor (V158411) increased DNA damage and cell death in models of human cancer. V158411 induced a time- and concentration-dependent increase in γH2AX-positive nuclei that was restricted to cells actively undergoing DNA synthesis. γH2AX induction was an early event and correlated with activation of the ATR/ATM/DNA-PKcs DNA damage response pathways. The appearance of γH2AX positive nuclei preceded ssDNA appearance and RPA exhaustion. Complete and sustained inhibition of Chk1 kinase was necessary to activate a robust γH2AX induction and growth inhibition. Chk1 inhibitor cytotoxicity correlated with induction of DNA damage with cells undergoing apoptosis, mitotic slippage and DNA damage-induced permanent cell cycle arrest. We identified two distinct classes of Chk1 inhibitors: those that induced a strong increase in γH2AX, pChk1 (S317) and pRPA32 (S4/S8) (including V158411, LY2603618 and ARRY-1A) and those that did not (including MK-8776 and GNE-900). Tumor cell death, induced through increased DNA damage, coupled with abrogation of cell cycle checkpoints makes selective inhibitors of Chk1 a potentially useful therapeutic treatment for multiple human cancers.
Vernalis Research, Cambridge, CB21 6GB, UK.
Joanne
J
Wayne
Vernalis Research, Cambridge, CB21 6GB, UK.
Teresa
T
Brooks
Vernalis Research, Cambridge, CB21 6GB, UK.
Andrew J
AJ
Massey
D000970:Antineoplastic Agents / Q000627:therapeutic use; D016923:Cell Death / Q000187:drug effects*; D048708:Cell Growth Processes / Q000187:drug effects; D000071877:Checkpoint Kinase 1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D003110:Colonic Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism*; D004249:DNA Damage / Q000187:drug effects*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D019073:HT29 Cells; D006657:Histones / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007211:Indoles / Q000494:pharmacology* / Q000627:therapeutic use; D011728:Pyridones / Q000494:pharmacology* / Q000627:therapeutic use; D016196:S Phase
D000970:Antineoplastic Agents; C468783:H2AX protein, human; D006657:Histones; D007211:Indoles; D011728:Pyridones; C000596547:V158411; D000071877:Checkpoint Kinase 1
Chk1; DNA damage; H2AX; kinase inhibitor; replication stress