Defining the role of chromosomal break end synapsis factors for DNA repair

定义染色体断裂末端突触因子在 DNA 修复中的作用

• 批准号: 10743770
• 负责人: Metztli Cisneros
• 金额: $ 3.32万
• 依托单位: BECKMAN RESEARCH INSTITUTE/CITY OF HOPE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10743770
• 关键词: Address; Affect; Binding; Biological Assay; Catalytic Domain; Cell Line; Chromosomal Breaks; Chromosome Pairing; Clinical Trials; Complex; DNA; DNA Binding; DNA Double Strand Break; DNA Repair; DNA-dependent protein kinase; Defect; Development; Distal; Double Effect; Double Strand Break Repair; Event; Excision; Frequencies; G22P1 gene; Ionizing radiation; Knock-out; Knowledge; Learning; Link; Liquid substance; Malignant Neoplasms; Measures; Mediating; Mission; Molecular Genetics; Mutation; Nonhomologous DNA End Joining; Outcome; Pathway interactions; Patient-Focused Outcomes; Phase; Phosphotransferases; Poison; Postdoctoral Fellow; Publications; Publishing; Radiation therapy; Regulation; Research; Research Personnel; Research Project Grants; Research Proposals; Role; Single-Stranded DNA; Site; Synapses; Techniques; Testing; Therapeutic; Topoisomerase; Training; Treatment outcome; United States National Institutes of Health; Work; XRCC4 gene; cancer cell; cancer clinical trial; cancer therapy; chromatin immunoprecipitation; clastogen; genetic approach; improved; inhibitor; insertion/deletion mutation; insight; mutant; nuclease; p53-binding protein 1; protein kinase inhibitor; repaired; response; skills; small molecule inhibitor; targeted cancer therapy; therapeutic target; treatment response

SUMMARY. Clastogenic cancer therapeutics, i.e., agents that induce chromosomal DNA double-strand breaks (DSBs), such as radiotherapy, remain a cornerstone of cancer treatment. A central aspect of DSB repair is synapsis of the two DSB ends to mediate repair, such that DSB end synapsis is a potential therapeutic target. Indeed, one factor important for DSB end synapsis, DNA-dependent protein kinase (DNA-PKcs), is the target of several inhibitors. This includes M3814/Peposertib (EMD Sereno), which is being tested in clinical trials. However, at least two other factors, XLF and 53BP1, have also been implicated in DNA end synapsis during repair. This suggests there may be some redundancy between these factors that could influence response to inhibitors of DNA-PKcs. In this research proposal, I will investigate the role of end synapsis factors on end joining (EJ) repair, and the regulation of DNA end resection. Aim 1: Define the role of end synapsis factors on EJ. Aim 1a (Completed): EJ without insertions or deletions (No Indel EJ) is a repair outcome that represents high-fidelity EJ, which requires factors involved in canonical non-homologous end joining (C-NHEJ), including KU70 and XRCC4. I sought to define the relative influence of two end synapsis factors (DNA-PKcs and XLF) on No Indel EJ. I found that DNA-PKcs and XLF promote No Indel EJ, however loss of XLF caused a much greater decrease in No Indel EJ compared to DNA-PKcs. Importantly, I found that disrupting DNA-PKcs (knockout or treatment with inhibitor) when combined with XLF mutations that weaken interaction interfaces, caused a synergistic loss of No Indel EJ. Thus, the conclusion of my published study is that the role of DNA-PKcs is magnified to promote No Indel EJ when XLF is weakened. Aim 1b (F99 Phase). Building on these findings, I will investigate another end synapsis factor, 53BP1. Specifically, I will address the hypothesis that 53BP1 has a partially redundant role with DNA-PKcs and XLF to promote chromosomal EJ. In performing this research, I will expand my skill set in several ways, including learning to assess EJ outcomes using different computational approaches from the completed research. Aim 2 (K00 Phase). In the postdoctoral phase of the research proposal, I will be assessing the role of synapsis factors on cleaving of DNA ends bound to DNA-PKcs to initiate end resection. End resection is the first step of homology-directed repair, and is mediated by the MRN/CtIP nuclease complex. Recently, MRN/CtIP was shown to cleave DNA ends bound to DNA-PKcs, which may be the key initiating step of end resection. I propose to test the hypothesis that disruption of end synapsis causes an increase in this initiation step of end resection, which I will test with several techniques that will expand my research skill set. Altogether, these studies will fill a major gap in our understanding of the role of end synapsis factors on the regulation of DNA DSB repair mechanisms. Such research is significant for developing end synapsis as a target for cancer therapy, including identifying the ideal circumstances to apply inhibitors of DNA-PKcs (e.g. M3814/Peposertib) during cancer treatment.

总结。致裂性癌症治疗剂，即诱导染色体DNA双链断裂的药物