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

项目总结 基因干扰DNA聚合酶Theta(POLθ)活性已被证明有效地靶向突变的BRCA1 细胞，同时保持BRCA1野生型(WT)细胞完整。POLθ促进theta介导的dna末端连接 在含有3‘单链DNA的断裂部位促进DNA突触修复和修复合成 悬垂的。尽管目前正在开发POLθ活性的小分子抑制剂用于治疗 在BRCA1突变的癌症中，关于激活TMEJ并导致Polθ的机制知之甚少。 BRCA1突变细胞的依赖性。此外，目前的范式假设同源重组 (HR)-缺乏PolθI敏感性，因此PARPI反应性有望成为Polθ的生物标志物 抑制剂(POLθI)敏感性。在我们的初步数据中，我们出人意料地发现了常用的BRCA1突变 生长相对不受基因Polq(POLθ)基因敲除(KO)干扰的细胞，表明POLθI和PARPI 敏感度可能不一定相关。在这项提案中，我们将阐明TMEJ的分子要求 并确定区分PARPI和POLθI敏感性的生物学因素。我们将解决以下问题 具体目的：1)揭示BRCA1突变细胞中TMEJ激活的分子基础；2)揭示遗传Polθ- 在BRCA1突变细胞中的依赖性；以及3)在BRCA1突变细胞中检测药理上的POLθ抑制。 总而言之，这些研究将为未来使用POLθI的临床研究提供参考。

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.