Evaluating the mechanism by which the DYNLL1-MRE11 complex regulates DNA end resection and genome stability.

评估 DYNLL1-MRE11 复合物调节 DNA 末端切除和基因组稳定性的机制。

• 批准号: 10606076
• 负责人: Michelle Swift
• 金额: $ 6.91万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-01 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10606076
• 关键词: Attenuated; BARD1 gene; BRCA deficient; BRCA1 Protein; BRCA1 gene; Binding; Blood Cells; CRISPR screen; Cancer Biology; Cancer Patient; Carcinoma; Cell Cycle; Cells; Clinical; Complex; DNA; DNA Damage; DNA Double Strand Break; DNA Repair; DNA Repair Gene; DNA-PKcs; Development; Equilibrium; Excision; Frequencies; G1 Phase; G2 Phase; Genes; Genome Stability; Genomic DNA; Genomic Instability; Germ-Line Mutation; Impairment; Malignant Neoplasms; Malignant neoplasm of ovary; Malignant neoplasm of pancreas; Malignant neoplasm of prostate; Mediating; Methodology; Modeling; Modification; Molecular; Mutation; Nonhomologous DNA End Joining; Ovarian Serous Adenocarcinoma; Pathway interactions; Patients; Phase; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphotransferases; Poly(ADP-ribose) Polymerase Inhibitor; Poly(ADP-ribose) Polymerases; Primary Neoplasm; Proteins; Regulation; Resected; Resistance; Resistance development; Role; S phase; Sampling; Serine; Serous; Sister Chromatid; Therapeutic; Ubiquitination; brca gene; burden of illness; cancer cell; clinically relevant; combat; genome-wide; genomic profiles; homologous recombination; in vivo; inhibitor; inhibitor therapy; malignant breast neoplasm; member; mutant; next generation sequencing; novel; nuclease; p53-binding protein 1; patient derived xenograft model; prevent; recruit; refractory cancer; repaired; response; restoration; synthetic lethal interaction; therapeutic target; tumor; tumorigenesis

PROJECT SUMMARY The synthetic lethal interaction of BRCA deficiency with PARPi is being exploited therapeutically in diverse clinical contexts, including ovarian, pancreatic, and prostate cancers). However, PARPi resistance has emerged as a vexing clinical problem for the treatment of BRCA1/2-deficient ovarian cancer carcinomas. Analyses of PARPi resistance in BRCA1-mutant tumors across independent studies suggest that loss of genes involved in either promoting NHEJ or suppressing HR leads to the partial restoration of HR in the absence of BRCA1 activity. 53BP1 emerged as the master regulator that brings multiple complexes to the DSB. Loss of any of these factors causes PARPi resistance in BRCA1 mutant cells. Therefore, further understanding of factors that regulate the processing of DNA ends that are crucial for pathway choice have tremendous relevance in cancer biology. Our lab has previously identified that in ovarian cancer cells, loss of DYNLL1, a factor that is constitutively bound to 53BP1, also results in enhanced HR and PARPi resistance. We found that DYNLL1pS88 directly binds to and inhibits MRE11, thereby blocking the initiation of DNA end resection. However, how the interaction of DYNLL1 with MRE11 impairs its nuclease activity or its recruitment to foci remains unexplored. Furthermore, how this interaction is regulated in the context of the DNA damage response, for example, which kinase/phosphatase(s) regulates the phosphorylation of S88 residue of DYNLL1 to modulate its interaction with MRE11 needs to be investigated. Our overall objectives in this application are to identify the molecular mechanism(s) regulating repair pathway choice by further understanding how end resection proteins are regulated. Our central hypothesis is that cell cycle-specific regulation of DYNLL1 promotes BRCA1/BARD1 mediated ubiquitination of MRE11, thereby facilitating end resection at DSBs in S phase. We aim to understand the mechanism by which DYNLL1 mediates end resection and HR. We will first investigate the dynamics between BRCA1 and DNA-PKcs and how it regulates the DYNLL1 and MRE11 activity in the context of the cell cycle. Investigating the dynamics of these end resecting factors and their regulation in DSB repair bear significant clinical relevance in combating PARPi resistance in BRCA1-mutant tumors. Therefore, we propose to determine whether alterations in mechanisms involved in attenuated DNA end resection may drive development of PARPi resistance in the high grade serous ovarian cancer (HGSOC) using patient derived xenografts (PDXs) and primary tumors. We expect that these studies will further our understanding of how end resection machinery is regulated to ensure that the appropriate repair pathway is activated. Further understanding of these end resection factors and how they are dysregulated in cancer may lead us to a relevant therapeutic target to combat PARPi resistant cancers.

项目摘要 BRCA缺陷与PARPi的合成致死相互作用正在各种治疗中得到利用。 临床背景，包括卵巢癌、胰腺癌和前列腺癌）。然而，PARPi的耐药性已经出现 作为治疗BRCA 1/2缺陷型卵巢癌的一个棘手的临床问题。分析 BRCA 1突变肿瘤中PARPi耐药的独立研究表明，参与BRCA 1突变肿瘤的基因缺失， 促进NHEJ或抑制HR导致在缺乏BRCA 1活性的情况下HR的部分恢复。 53 BP 1作为主调节剂出现，为DSB带来多个复合物。任何这些因素的损失 在BRCA 1突变细胞中引起PARPi抗性。因此，进一步了解调节因素， 对途径选择至关重要的DNA末端的加工在癌症生物学中具有巨大的相关性。 我们的实验室先前已经确定，在卵巢癌细胞中，DYNLL 1的缺失，DYNLL 1是一种组成性调节卵巢癌细胞增殖的因子， 与53 BP 1结合，也导致增强的HR和PARPi抗性。我们发现DYNLL 1 pS 88直接结合 并抑制MRE 11，从而阻断DNA末端切除的起始。然而， DYNLL 1与MRE 11削弱其核酸酶活性或其募集到病灶仍然未探索。此外，委员会认为， 这种相互作用是如何在DNA损伤反应的背景下进行调节的，例如， 激酶/磷酸酶调节DYNLL 1的S88残基的磷酸化，以调节其与DYNLL 1的相互作用。 MRE 11需要进行研究。 本申请的总体目标是确定调节修复的分子机制 通过进一步了解末端切除蛋白是如何调节的途径选择。我们的核心假设是 DYNLL 1的细胞周期特异性调节促进BRCA 1/BARD 1介导的MRE 11的泛素化， 从而便于在S期的DSB处进行末端切除。 我们的目标是了解DYNLL 1介导末端切除和HR的机制。 研究BRCA 1和DNA-PKcs之间的动力学以及它如何调节DYNLL 1和MRE 11活性 在细胞周期中。研究这些终末切除因素的动态变化及其在肿瘤发生中的调节作用， DSB修复在对抗BRCA 1突变型肿瘤中的PARPi耐药性方面具有显著的临床相关性。 因此，我们建议，以确定是否在机制的改变参与衰减的DNA末端， 切除术可能会导致高级别浆液性卵巢癌（HGSOC）中PARPi耐药的发展， 患者来源的异种移植物（PDX）和原发性肿瘤。我们希望这些研究将进一步促进我们的 了解如何调节末端切除机制，以确保适当的修复途径， 激活进一步了解这些末端切除因子以及它们在癌症中是如何失调的， 引导我们找到一个相关的治疗靶点来对抗PARPi抗性癌症。