Abstract B20: Schedule-dependent activation of DNA helicases by Checkpoint Kinase 1 inhibition following dNTP depletion causes CDK2-independent replication catastrophe

DNA damaging agents are standard of care therapies for many cancers, but frequently produce poor outcomes in many patients. Using DNA damage checkpoint inhibitors to sensitize cancer cells to standard therapies is a rapidly emerging strategy. Understanding the mechanism behind sensitization will help clinicians develop rational combinations and schedules to improve cancer care. Additionally, understanding how defects in the DNA damage response affect treatment outcomes may help identify which patients will respond to treatment. We have previously demonstrated that inhibition of Checkpoint Kinase 1 (Chk1) with MK8776 sensitizes cells to the DNA damaging agent, gemcitabine, in a schedule-dependent manner. However, the mechanism of sensitization as well as the reason why sensitization increases with time following gemcitabine are not well understood. Gemcitabine inhibits ribonucleotide reductase and rapidly depletes deoxyribonucleotides (dNTPs) leading to S phase arrest. We found that delayed (18-24 h), but not concurrent (0-6 h), Chk1 inhibition following gemcitabine activated the replication machinery. Using chromatin fractionation and western blotting, we found that the helicase co-factor, CDC45, is recruited to DNA during delayed Chk1i after gemcitabine. However, the continued lack of dNTPs prevented functional DNA replication during activation of replicative helicases. This resulted in formation of excessive single-stranded DNA as measured by native BrdU fluorescent microsc...
Source: Molecular Cancer Research - Category: Cancer & Oncology Authors: Tags: Therapies Targeting Checkpoints and Mismatch Repair: Poster Presentations - Proffered Abstracts Source Type: research