Investigating DNA polymerase O in replication stress and cancer therapy - Principal Investigator: Wu, Xiaohua Project Summary Replication stress often occurs when DNA replication is disrupted, leading to DNA double-strand break (DSB) formation. Replication stress can also be triggered by oncogene expression and is associated with tumor development. Notably, replication stress creates vulnerabilities in cancer cells that can be exploited for cancer treatment. Nevertheless, our understanding of the detailed DNA repair mechanisms upon fork breakage due to replication stress, as well as our capacity to effectively target the associated vulnerabilities for cancer treatment, remains limited. DNA polymerase θ (POLQ) is a specialized DNA polymerase critical for DSB repair, and compromised POLQ function leads to genomic instability and radiation sensitivity. Substantial evidence suggests that POLQ plays an important role in microhomology-mediated end joining (MMEJ). Given the prevalence of microhomologies at cancer breakpoints, POLQ-mediated MMEJ is thought to be involved in promoting genome instability associated with cancer, but the precise underlying mechanism remains unclear. In this study, we discovered a unique role of POLQ in repair of DSBs linked to fork breakage, revealing a new mechanism that requires POLQ to cope with replication stress. Based on the role of POLQ in repair of damaged forks, we found that inhibiting POLQ leads to cell death under replication stress, and this effect is significantly intensified when ATR is inhibited. We propose that combined therapeutic strategy utilizing POLQ inhibitors and ATR inhibitors would synergistically eradicate cancer cells experiencing high replication stress. Given the current proposed use of POLQ inhibitors is primarily for treating BRCA-deficient tumors, this study will substantially broaden the application of POLQ inhibitors. We hypothesize that the role of POLQ in repairing broken forks forms the basis for treating replication-stressed cancer cells with POLQ inhibitors and ATR inhibitors, and propose to investigate the mechanism behind POLQ-mediated MMEJ in repair of broken forks. We will examine how POLQ is engaged to repair replication-associated DSBs and study the collaborative functions of POLQ-mediated MMEJ and break-induced replication (BIR) in repairing broken forks. We will use cell-based assay and xenograft mouse model to explore cancer therapeutic strategy of using POLQ inhibitors and ATR inhibitors. This study will not only shed light on the role of POLQ-mediated MMEJ in coping with replication stress, but will also lay the groundwork for potential cancer treatments by targeting cancer vulnerabilities associated with replication stress. Since replication stress is highly associated with cancer, this study will additionally bring insights into the DNA repair mechanisms that modulate genome integrity during tumor development.
