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Combining DNA-damaging drugs with DNA checkpoint inhibitors is an emerging strategy to manage cancer. Checkpoint kinase 1 inhibitors (CHK1is) sensitize most cancer cell lines to DNA-damaging drugs and also elicit single-agent cytotoxicity in 15% of cell lines. Consequently, combination therapy may be effective in a broader patient population. Here, we characterized the molecular mechanism of sensitization to gemcitabine by the CHK1i MK8776. Brief gemcitabine incubation irreversibly inhibited ribonucleotide reductase, depleting dNTPs, resulting in durable S phase arrest. Addition of CHK1i 18 h after gemcitabine elicited cell division cycle 7 (CDC7)- and cyclin-dependent kinase 2 (CDK2)-dependent reactivation of the replicative helicase, but did not reinitiate DNA synthesis due to continued lack of dNTPs. Helicase reactivation generated extensive single-strand (ss)DNA that exceeded the protective capacity of the ssDNA-binding protein, replication protein A. The subsequent cleavage of unprotected ssDNA has been termed replication catastrophe. This mechanism did not occur with concurrent CHK1i plus gemcitabine treatment, providing support for delayed administration of CHK1i in patients. Alternative mechanisms of CHK1i-mediated sensitization to gemcitabine have been proposed, but their role was ruled out; these mechanisms include premature mitosis, inhibition of homologous recombination, and activation of double-strand break repair nuclease (MRE11). In contrast, single-agent activity of CHK1i was MRE11-dependent and was prevented by lower concentrations of a CDK2 inhibitor. Hence, both pathways require CDK2 but appear to depend on different CDK2 substrates. We conclude that a small-molecule inhibitor of CHK1 can elicit at least two distinct, context-dependent mechanisms of cytotoxicity in cancer cells.
Pubmed ID: 30573684 RIS Download
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This unknown targets
View all literature mentionsThis monoclonal targets RPA32/RPA2
View all literature mentionsThis monoclonal targets Chk1 (G-4)
View all literature mentionsThis unknown targets Goat Mouse IgG (H L)-HRP Conjugate
View all literature mentionsThis polyclonal targets CDC45L
View all literature mentionsThis monoclonal targets CDK2
View all literature mentionsThis monoclonal targets Chk1, phospho (Ser345)
View all literature mentionsThis polyclonal targets IgG (H+L)
View all literature mentionsThis monoclonal targets PCNA (PC10) Mouse mAb (Alexa Fluor 488 Conjugate)
View all literature mentionsThis unknown targets Rabbit IgG (H+L)
View all literature mentionsThis polyclonal secondary targets IgG (H+L)
View all literature mentionsThis monoclonal targets Histone H3, phospho (Ser10)
View all literature mentionsThis unknown targets mouse IgG (H+L) (DyLight 680 Conjugate)
View all literature mentionsThis polyclonal targets RPA32, phospho
View all literature mentionsThis monoclonal targets CHEK2
View all literature mentionsThis polyclonal secondary targets IgG (H+L)
View all literature mentionsThis monoclonal targets MCM2
View all literature mentionsThis polyclonal targets MEK1/2
View all literature mentionsThis unknown targets Phospho-MCM2 (Ser53)
View all literature mentionsThis monoclonal targets cyclin E (HE12)
View all literature mentionsThis unknown targets rabbit IgG (H+L) (DyLight 800 Conjugate)
View all literature mentionsThis monoclonal targets Histone H2A.X, phospho (Ser139)
View all literature mentionsThis monoclonal targets RPA32
View all literature mentionsThis monoclonal targets Actin, beta
View all literature mentionsThis polyclonal secondary targets IgG (H+L)
View all literature mentionsThis polyclonal targets Chk1, phospho, (Ser296)
View all literature mentionsThis monoclonal targets Chk1 (G-4)
View all literature mentionsThis monoclonal targets CHEK2
View all literature mentionsThis monoclonal targets MCM2
View all literature mentionsThis monoclonal targets Histone H3, phospho (Ser10)
View all literature mentionsThis unknown targets Goat Mouse IgG (H L)-HRP Conjugate
View all literature mentionsThis unknown targets Goat Mouse IgG (H L)-HRP Conjugate
View all literature mentionsThis polyclonal targets IgG (H+L)
View all literature mentionsThis monoclonal targets Chk1, phospho (Ser345)
View all literature mentionsThis unknown targets rabbit IgG (H+L) (DyLight 800 Conjugate)
View all literature mentionsThis monoclonal targets Histone H2A.X, phospho (Ser139)
View all literature mentionsThis unknown targets mouse IgG (H+L) (DyLight 680 Conjugate)
View all literature mentionsThis monoclonal targets RPA32/RPA2
View all literature mentionsThis polyclonal targets CDC45L
View all literature mentionsThis unknown targets Phospho-MCM2 (Ser53)
View all literature mentionsThis monoclonal targets Actin, beta
View all literature mentionsThis polyclonal targets RPA32, phospho
View all literature mentionsThis polyclonal targets MEK1/2
View all literature mentionsThis unknown targets
View all literature mentionsThis monoclonal targets CDK2
View all literature mentionsThis monoclonal targets Histone H3, phospho (Ser10)
View all literature mentionsThis monoclonal targets PCNA (PC10) Mouse mAb (Alexa Fluor 488 Conjugate)
View all literature mentionsThis polyclonal targets Chk1, phospho, (Ser296)
View all literature mentionsThis unknown targets Rabbit IgG (H+L)
View all literature mentionsThis polyclonal secondary targets IgG (H+L)
View all literature mentionsThis polyclonal secondary targets IgG (H+L)
View all literature mentionsThis monoclonal targets cyclin E (HE12)
View all literature mentionsThis polyclonal secondary targets IgG (H+L)
View all literature mentionsThis monoclonal targets RPA32
View all literature mentionsThe SciCrunch Infrastructure was developed as a cooperative data platform to be used by diverse communities in making data more FAIR.
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