Examining the Role of RNF168 Activity in BRCA1 Mutant Cancers

检查 RNF168 活性在 BRCA1 突变癌症中的作用

基本信息

批准号：
10572529
负责人：
John Krais
金额：
$ 15.22万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-09-01 至 2026-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10572529
关键词：
Address Alleles Automobile Driving BARD1 gene BRCA1 Mutation BRCA1 Protein BRCA1 gene BRCT Domain Back Biological Biological Assay Biological Models C-terminal CRISPR/Cas technology Cancer Biology Cancer Patient Cancer cell line Cell Line Cell Proliferation Cell Survival Cells Clinical Coiled-Coil Domain DNA Damage DNA Double Strand Break DNA Repair DNA Repair Gene Data Defect Dependence Development Embryo Embryonic Development Exons Foundations Funding Future Genetic Goals Growth Immunofluorescence Immunologic Impairment Knock-out Knowledge Malignant Neoplasms Malignant neoplasm of ovary Mediating Modeling Mutant Strains Mice Mutation N-terminal Nonhomologous DNA End Joining PARP inhibition Pathogenicity Pathway interactions Phenotype Poly(ADP-ribose) Polymerase Inhibitor Positioning Attribute Process Proteins Reporter Research Research Personnel Resistance Resistance development Role Secure Signal Transduction System Testing Transgenic Mice Transgenic Organisms Work Xenograft procedure addiction cancer cell cancer risk cancer therapy career drug development effective therapy homologous recombination inhibitor therapy insight mouse model mutant patient derived xenograft model programs recruit refractory cancer repaired response restoration success targeted treatment therapeutic target treatment response treatment strategy tumor tumor growth tumor progression tumorigenesis ubiquitin-protein ligase

项目摘要

PROJECT SUMMARY/ABSTRACT My career goal is to develop an independent research program focused on DNA damage signaling and how it can be exploited for the treatment of cancer. The work that I propose here explores DNA damage signaling mechanisms in BRCA1 mutant cancers and the implications for PARP inhibitor response. Pathogenic BRCA1 mutations increase the risk of cancer development and impair the homologous recombination (HR) DNA damage response pathway. This HR defect confers vulnerability to DNA damaging agents, including PARP inhibitors. PARP inhibitor efficacy is limited by the development of resistance, often caused by the restoration of HR. In BRCA1 mutant cancers, PARP inhibitor resistance and HR activity can be promoted by expression of mutant BRCA1 proteins. Several types of truncated hypomorphic BRCA1 proteins have been detected, though the mechanism of localization to DNA damage and HR activity has not been established. Recently, we determined that wild-type BRCA1 recruitment to DNA damage can proceed through RNF168-dependent and -independent pathways involving the BRCA1 RING and BRCT domains. In preliminary data, we found that RNF168 depletion impairs tumor growth for some, but not all, BRCA1 mutant cancers. Additionally, we show that some cancer cells are entirely reliant on the RNF168 pathway for HR, whereas others rely on the reciprocal pathway. These findings are in line with transgenic mouse studies where different Brca1 mutant alleles produced contrasting phenotypes upon RNF168 knockout. Here, we explore the possibility that BRCA1 mutation-specific differences arise from the capacity to recruit various hypomorphic BRCA1 proteins to DNA damage. We will use cell line, patient- derived xenograft, and transgenic mouse model systems to address the fundamental question of how the RNF168 pathway contributes to tumor progression, DNA repair, and PARP inhibitor response in different BRCA1 mutant backgrounds. Specifically, we hypothesize that depletion of RNF168 in cancers expressing BRCA1 RING domain-containing hypomorphic proteins will impair tumor progression, eliminate HR activity, and restore sensitivity to PARP inhibitors. To test this hypothesis, we developed PARP inhibitor sensitive and resistant cell line and patient-derived xenograft models that express hypomorphic BRCA1 proteins. Additionally, we generated isogenic expression systems as well as transgenic mice with Rnf168 and various Brca1 mutations. Using these models, we will examine the effects of genetic depletion of RNF168 on tumorigenesis and growth, localization of BRCA1 hypomorphs and other DNA repair proteins, and response to PARP inhibition. These studies will provide mechanistic insight into the repair processes occurring in cancers and assess the potential efficacy of RNF168 pathway-targeted therapeutics. Moreover, funding of this proposal will help me to secure an independent investigator position and lay the foundation for future R01 applications to build a sustainable research program focused on DNA repair and cancer biology.

项目摘要/摘要我的职业目标是制定一项针对DNA损伤信号及其如何的独立研究计划可以利用用于治疗癌症。我在这里提出的工作探讨了DNA损伤信号传导 BRCA1突变癌的机制以及对PARP抑制剂反应的影响。致病性BRCA1 突变增加了癌症发展的风险并损害同源重组（HR）DNA损伤响应途径。该人力资源缺陷使DNA破坏药物（包括PARP抑制剂）赋予了脆弱性。 PARP抑制剂疗效受到耐药性的发展的限制，通常是由人力资源的恢复引起的。在 BRCA1突变体癌，PARP抑制剂耐药性和HR活性可以通过表达突变促进 BRCA1蛋白。已经检测到了几种类型的截短的肌电BRCA1蛋白，尽管尚未建立针对DNA损伤和HR活性的定位机制。最近，我们确定野生型BRCA1募集到DNA损伤可以通过RNF168依赖性和无关涉及BRCA1环和BRCT域的途径。在初步数据中，我们发现RNF168耗竭损害某些但不是全部BRCA1突变体癌症的肿瘤生长。此外，我们表明一些癌细胞完全依赖于HR的RNF168途径，而其他人则依靠相互途径。这些发现与转基因小鼠研究一致，其中不同的BRCA1突变等位基因产生了对比表型在RNF168淘汰赛上。在这里，我们探讨了BRCA1突变特异性差异的可能性募集各种肌差BRCA1蛋白造成DNA损伤的能力。我们将使用细胞系，患者 - 派生的异种移植物和转基因小鼠模型系统，以解决如何解决的基本问题 RNF168途径在不同的BRCA1中有助于肿瘤进展，DNA修复和PARP抑制剂反应突变背景。具体而言，我们假设表达BRCA1环的癌症中RNF168的耗竭含结构域的肌蛋白会损害肿瘤进展，消除HR活性并恢复对PARP抑制剂的敏感性。为了检验该假设，我们开发了PARP抑制剂敏感和抗性细胞线和患者衍生的异种移植模型，表达肌肉brca1蛋白。此外，我们生成了同源表达系统以及具有RNF168和各种BRCA1突变的转基因小鼠。使用这些模型，我们将研究RNF168遗传耗竭对肿瘤发生和生长的影响， BRCA1肌电和其他DNA修复蛋白，以及对PARP抑制作用的反应。这些研究将提供机械洞察癌症中发生的修复过程并评估RNF168的潜在功效针对途径的治疗学。此外，该提案的资金将有助于我确保独立研究者的职位并为未来R01应用程序奠定基础，以建立可持续研究计划专注于DNA修复和癌症生物学。