RESISTANCE AND RESPONSE SIGNALING TO PARP1 INHIBITORS IN OVARIAN CANCER

卵巢癌中 PARP1 抑制剂的耐药性和反应信号

• 批准号: 10197460
• 负责人: Alvaro N Monteiro
• 金额: $ 23.1万
• 依托单位: H. LEE MOFFITT CANCER CTR & RES INST
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-01 至 2023-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10197460
• 关键词: ADP ribosylation; Affinity; Apoptosis; BRCA deficient; BRCA mutations; BRCA1 Protein; BRCA1 gene; BRCA2 Protein; BRCA2 gene; Binding; Biological; Biological Assay; Cancer Patient; Cancer cell line; Cell Cycle; Cell Line; Cells; Cellular Assay; Chemoprevention; Clinic; Clinical; Cloning; Clustered Regularly Interspaced Short Palindromic Repeats; Co-Immunoprecipitations; Combined Modality Therapy; Complex; DNA Damage; Data; Development; Dissection; Drug resistance; Ectopic Expression; Event; Foundations; Future; Gene Silencing; Goals; Immunoblotting; Individual; Lead; Link; Malignant neoplasm of ovary; Mass Spectrum Analysis; Measures; Mediating; Medical; Methods; Modeling; Multiprotein Complexes; Mutation; Operative Surgical Procedures; Organoids; Pathogenicity; Patients; Pharmaceutical Preparations; Phosphorylation; Poly(ADP-ribose) Polymerases; Proteins; Proteome; Proteomics; RNA Interference; Research; Resistance; Resources; Risk; Role; Signal Pathway; Signal Transduction; Signaling Protein; Testing; Time; Tyrosine; Variant; Woman; base; brca gene; cancer cell; cancer risk; cancer therapy; candidate marker; clinical development; design; fight against; genetic testing; improved; inhibitor/antagonist; mutant; neoplastic cell; new therapeutic target; novel; novel marker; ovarian neoplasm; p53-binding protein 1; patient stratification; patient subsets; phosphoproteomics; preclinical development; predicting response; predictive marker; prophylactic; protein complex; refractory cancer; response; response biomarker; tumor

The last twenty years have seen significant progress in ovarian cancer treatment, in particular for the women carrying pathogenic variants in BRCA1 or BRCA2. The foundation for this progress was the identification and cloning of BRCA1 and BRCA2, which allowed for the development of genetic tests to identify individuals at elevated risk who could benefit from increased surveillance, prophylactic surgery, and chemoprevention. A dissection of the biological role of BRCA1 and BRCA2 proteins led to the realization that synthetic lethal approaches could be successful in treating tumors with inactivated BRCA1/2 using PARP1 inhibitors. Despite significant response rates upon PARPi therapy in BRCA-linked advanced ovarian cancer, some challenges, most notably drug resistance, persist. A sizable percentage of patients display primary or intrinsic resistance despite predictions based on their BRCA1/2 status. Our long term goal is to extend the benefits of PARPi therapy to a larger proportion of patients. We hypothesize that BRCA-linked ovarian tumors that do not respond to PARPi display significant changes in PARP1 protein complexes and the cellular signaling network that can be detected using integrated functional proteomics. We have recently shown that mass spectrometry (MS)-based affinity proteomics with DDR proteins or with PARPis as baits is a powerful method to characterize multiprotein complexes, and that phosphoproteomics is ideally suited to capture complementary proteome- wide phosphorylation changes. We propose to define PARP1 interacting complex proteins and compensatory signaling in BRCA-linked ovarian cancers. We propose the following Specific Aims: Aim 1. To determine PARP1 multiprotein complexes in PARPi-sensitive and –intrinsically resistant BRCA-mutant ovarian cancer cells. Using quantitative PARPi affinity proteomics in tumor cells, we will identify PARP1 protein complex changes that correlate with PARPi response in BRCA-deficient cancer cells. We will utilize a unique resource of established and primary ovarian cancer cell lines and CRISPR-based isogenic cell lines. Aim 2. To determine basal and drug adaptive signaling in PARPi-sensitive and –intrinsically resistant BRCA-mutant ovarian cancer cells. We will harness the panel of tumor models introduced in Aim 1, including primary and isogenic cancer cell lines, and patient-derived organoids. Time-resolved quantitative MS-based global (pSTY) and tyrosine (pY) phosphoproteomics will measure proteome-wide signaling changes in untreated and PARPi- treated sensitive and intrinsically resistant cancer cells. We will interrogate and validate specific signals using a panel of known DDR signaling pathway inhibitors by immunoblotting and cellular assays applied in Aim 1. This project will transform our understanding of the complexity and dynamics of proximal (i.e. PARP1 complex- associated) and network-wide signaling events that, individually or in conjunction, lead to primary PARPi resistance in BRCA-linked ovarian cancer.

过去二十年来，卵巢癌的治疗取得了重大进展，特别是对于女性而言 携带 BRCA1 或 BRCA2 致病变异的基因是这一进展的基础。 克隆 BRCA1 和 BRCA2，从而可以开发基因测试来识别个体 可以从加强监测、预防性手术和化学预防中受益的高风险人群 A。 对 BRCA1 和 BRCA2 蛋白生物学作用的剖析使我们认识到合成致死蛋白 尽管如此，使用 PARP1 抑制剂可以成功治疗 BRCA1/2 失活的肿瘤。 PARPi 治疗 BRCA 相关晚期卵巢癌的显着缓解率，一些挑战， 最显着的耐药性持续存在，相当大比例的患者表现出原发性或内在耐药性。 尽管根据 BRCA1/2 状态进行预测，但我们的长期目标是扩大 PARPi 的益处。 我们勇敢地接受了与 BRCA 相关的卵巢肿瘤的治疗。 对 PARPi 做出反应后，PARP1 蛋白复合物和细胞信号网络发生显着变化 我们最近证明了质谱法可以检测到这一点。 以 DDR 蛋白或 PARPis 作为诱饵的基于 (MS) 的亲和蛋白质组学是一种强有力的表征方法 多蛋白复合物，并且磷酸化蛋白质组学非常适合捕获互补蛋白质组 我们建议定义 PARP1 复合蛋白和补偿性变化。 我们提出以下具体目标： 目标 1. 确定 BRCA 相关卵巢癌中的信号传导。 PARPi 敏感且本质耐药的 BRCA 突变卵巢癌中的 PARP1 多蛋白复合物 利用肿瘤细胞中的定量 PARPi 亲和蛋白质组学，我们将鉴定 PARP1 蛋白复合物。 我们将利用独特的资源来研究与 BRCA 缺陷癌细胞中 PARPi 反应相关的变化。 目的 2. 已建立的原代卵巢癌细胞系和基于 CRISPR 的同基因细胞系。 确定 PARPi 敏感和本质耐药 BRCA 突变体的基础和药物适应性信号传导 我们将利用目标 1 中介绍的一组肿瘤模型，包括原发性和 同基因癌细胞系和患者来源的类器官，基于 MS 的时间分辨定量全局 (pSTY)。 和酪氨酸（pY）磷酸化蛋白质组学将测量未处理和 PARPi- 中蛋白质组范围内的信号变化 我们将使用敏感且具有内在抵抗力的癌细胞来询问和验证特定信号。 通过免疫印迹和细胞分析已知的 DDR 信号通路抑制剂组应用于目标 1。该测定 该项目将改变我们对近端（即 PARP1 复杂- 相关）和网络范围的信令事件，单独或联合导致主要 PARPi BRCA 相关卵巢癌的耐药性。