Abstract 4298: Targeting the DNA damage checkpoint kinase Chk1 induces multiple pathways of cytotoxicity

Abstract

Abstract DNA damaging agents are standard therapies for many cancers, but result in limited long-term clinical benefit. DNA damage checkpoints arrest DNA synthesis and provide time for repair and recovery. Using DNA checkpoint inhibitors to sensitize cancer cells to traditional chemotherapeutics is an emerging strategy. However, clinical trials involving Checkpoint Kinase 1 inhibitors (Chk1i) plus DNA-damaging chemotherapies have shown little clinical benefit. This may be due to sub-optimal scheduling of combinations. We have published that delaying Chk1i until 18 h after gemcitabine confers the greatest sensitization of cancer cells. We have now characterized the mechanisms by which the Chk1i, MK-8776, sensitizes human cancer cells to gemcitabine and SN38. SN38, the active metabolite of irinotecan, traps topoisomerase I on DNA resulting in S phase arrest. Chk1i re-activates replication and drives cells into mitosis before the damage from SN38 is repaired. Co-incubation with a CDK1/2 inhibitor surprisingly failed to prevent S phase progression, but did block mitotic entry and prevented further DNA damage. Conversely, the replication-associated kinase, CDC7, is essential for Chk1i-mediated S phase abrogation. CDC7 is presumably required to activate dormant origins of replication. Gemcitabine inhibits ribonucleotide reductase, thus preventing production of deoxyribonucleotides. Addition of Chk1i 18 h following gemcitabine causes aberrant activation of DNA helicases, but without deoxyribonucleotides, this results in excessive single-stranded DNA. Eventually, the level of single-stranded DNA exceeds cellular stores of the protective protein, RPA, which leads to nucleolytic cleavage and double-strand DNA breaks. Chk1i-mediated helicase activation is again dependent on CDC7, but insensitive to a CDK1/2 inhibitor. Concurrent incubation with gemcitabine plus MK8776 does not activate DNA helicases. These data provide a rationale for a delayed administration of Chk1i when patients are receiving gemcitabine. These investigations have detailed how Chk1i elicits distinct mechanisms of cytotoxicity depending on the context of DNA damage. Understanding of these mechanisms may help identify biomarkers to predict patient response. Furthermore, the clinical development of several Chk1i have been terminated due to unfavorable toxicity, but the cause of toxicity remains unknown. The work described here may help the clinical development by determining whether the observed toxicities are due to on- or off-targets. Citation Format: Nicholas Warren, Jennifer Ditano, Alan Eastman. Targeting the DNA damage checkpoint kinase Chk1 induces multiple pathways of cytotoxicity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4298.