Functional Evaluation and Interpretation of DNA Damage Repair Variants in Prostate Cancer

项目摘要/摘要 DNA损伤修复(DDR)途径基因的致病变异在部分男性患者中普遍存在 转移性去势抵抗前列腺癌(MCRPC)和发生在躯体和生殖系 基因组。这些异常，主要是导致蛋白质截断的插入和缺失，发生在10- 20%的mCRPC患者。这些变异也发生在较低级别的局限性前列腺癌中，尽管 DDR突变在这些肿瘤中的流行情况及其对临床结果、生存率和药物的影响 敏感性在很大程度上是未知的。在拟议拨款的其他项目中，DDR基因的特定突变将 被指认出来。项目3的重点将是确定这些突变的优先顺序并系统地评估它们，使用 结合功能分析、最先进的有机化合物技术、CRISPR介导的基因编辑和 全基因组测序分析。在项目3的目标1中，我们将使用多个前列腺癌细胞系来 确定特定的DDR基因突变是有害的还是良性的变异。我们将检验这一假设 真正的致病DDR等位基因与更具侵袭性的肿瘤表型相关，并将 与特定的功能性DNA修复缺陷和药物反应相关。在目标2中，我们将使用基因编辑来 将DDR突变引入多个前列腺癌细胞系的内源性遗传位点，以提供 对特定突变进行更多的生理学评估。在目标3中，我们将分析全基因组 来自前列腺癌的序列以识别可能由-1基因缺失引起的精确突变特征 特定DDR基因的功能突变。项目3将与调查人员在#年进行广泛合作 项目1和2以及所附研究计划中概述的基因组学核心。

PROJECT SUMMARY/ABSTRACT Pathogenic variants in DNA damage repair (DDR) pathway genes are prevalent in a subset of men with metastatic castration-resistant prostate cancer (mCRPC) and occur in both the somatic and germline genomes. These abnormalities, primarily insertions and deletions resulting in protein truncations, occur in 10– 20% of patients with mCRPC. These variants also occur in lower-grade localized prostate cancers, although the prevalence of DDR mutations in these tumors and their impact on clinical outcome, survival, and drug sensitivity are largely unknown. In other projects of the proposed grant, specific mutations in DDR genes will be identified. The focus of Project 3 will be to prioritize and systematically evaluate these mutations, using a combination of functional assays, state-of-the-art organoid technology, CRISPR-mediated gene editing, and analysis of whole genome sequencing. In Aim 1 of Project 3, we will use multiple prostate cancer cell lines to determine whether specific DDR gene mutations are deleterious or benign variants. We will test the hypothesis that bona fide pathogenic DDR alleles are associated with a more aggressive tumor phenotype and will correlate with specific functional DNA repair defects and drug responses. In Aim 2, we will use gene editing to introduce DDR mutations into the endogenous genetic locus of multiple prostate cancer cell lines to provide a more physiologic assessment of the specific mutations. In Aim 3, we will analyze the whole-genome sequences from prostate cancers to identify precise mutational signatures which may result from loss-of- function mutations in specific DDR genes. Project 3 will have extensive collaborations with the investigators in Projects 1 and 2 and the Genomics Core as outlined in the attached research plan.