Assessing DNA polymerase theta as a therapeutic target in BRCA1 mutant cancer

PROJECT SUMMARY Genetic disruption of DNA polymerase theta (Polθ) activity has been shown to effectively target BRCA1 mutated cells, while leaving BRCA1 wild-type (WT) cells intact. Polθ facilitates theta-mediated DNA end joining (TMEJ) repair by promoting DNA synapsis and repair synthesis at break sites containing 3' single stranded (ss)DNA overhangs. Although small molecule inhibitors of Polθ activity are currently under development for the treatment of BRCA1 mutant cancers, very little is known regarding the mechanisms that activate TMEJ and result in Polθ- dependency in BRCA1 mutant cells. Moreover, the current paradigm assumes that homologous recombination (HR)-deficiency confers Polθi sensitivity, therefore PARPi responsiveness is expected to be a biomarker for Polθ inhibitor (Polθi) sensitivity. In our preliminary data, we unexpectedly identified commonly used Brca1 mutant cells that grow relatively unperturbed with genetic Polq (Polθ) knockout (KO), indicating that Polθi and PARPi sensitivity may not necessarily correlate. In this proposal, we will elucidate the molecular requirements for TMEJ activation and identify biological factors that distinguish PARPi and Polθi sensitivity. We will address the following Specific Aims: 1) reveal the molecular basis of TMEJ activation in Brca1 mutant cells; 2) uncover genetic Polθ- dependencies in Brca1 mutant cells; and 3) examine pharmacologic Polθ inhibition in Brca1 mutant cells. Collectively, these studies will be informative for future clinical studies employing Polθi.