Stalled replication fork repair in cancer predisposition and cancertherapy

癌症易感性和癌症治疗中停滞的复制叉修复

基本信息

批准号：
10681456
负责人：
Ralph Scully
金额：
$ 99.59万
依托单位：
BETH ISRAEL DEACONESS MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-08-10 至 2029-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10681456
关键词：
ATP phosphohydrolase Ablation BRCA1 gene BRCA2 gene Breast Cancer therapy Cell Cycle Cells Chromatin Clinical Code DNA DNA Damage DNA Repair DNA analysis DNA biosynthesis DNA replication fork Defect Disease Fanconi&apos s Anemia Funding Genes Genetic Genomic Instability Goals Hereditary Breast and Ovarian Cancer Syndrome Human Light Link Malignant Neoplasms Malignant neoplasm of ovary Motor Mutation Oncogenes Outcome Prevention Proteins Regulation Replication-Associated Process Reporter Rest Role Site Techniques Therapeutic Time Variant Work Yeasts autosome cancer genome cancer predisposition druggable target homologous recombination human disease innovation insight loss of function mutation malignant breast neoplasm molecular targeted therapies mutant novel therapeutics prevent repaired response success synthetic lethal interaction targeted cancer therapy targeted treatment tool

项目摘要

PROJECT SUMMARY Error-free DNA repair initiated at the sites of replication fork stalling is critical for the prevention of genomic instability in cycling cells. Defects in stalled fork repair have been directly implicated in cancer predisposition and other human diseases. The clinical burden associated with failed stalled fork repair may include hereditary breast and ovarian cancer (HBOC) predisposition, in light of the involvement of BRCA1 and BRCA2 in repair of stalled replication forks, and Fanconi Anemia (FA)—a rare, autosomal recessive (or X-linked) disease caused by inactivation of any one of several FA genes. Our work previously established roles for BRCA1 and BRCA2 in regulating HR at both double strand breaks (DSBs) and in stalled fork repair. We developed innovative tools for quantifying homologous recombination (HR) and other repair outcomes at stalled mammalian replication forks and, more recently, at broken replication forks. A major goal of this proposal is to define the fundamental mechanisms of repair of stalled forks. We have developed an array of cutting-edge tools to support this study, including unique, sophisticated HR reporters that can distinguish between error-free “short tract” HR and error- prone “long tract” HR—a replicative response analogous to break-induced replication in yeast. One unusual aberrant replicative response that we observe at stalled forks specifically in BRCA1 mutant cells is the formation of <10 kb non-homologous tandem duplications (TDs). In a paradigm-shifting discovery, we found that these highly specific forms of structural variation are also abundant in the human BRCA1-linked breast and ovarian cancer genome. A major goal of this proposal is to define the genetic regulation and full mechanism of TD formation at stalled forks in BRCA1 mutant cells. Success in this project will reveal in unprecedented detail the mechanisms that regulate mammalian stalled (or broken) fork repair and their relationship to cancer predisposition. In support of this, we will develop new techniques for analyzing DNA structural intermediates, chromatin responses to fork stalling and protein composition of the stalled mammalian replication fork. These analytical studies may also identify new molecular targets for therapy of breast and ovarian cancer. Indeed, our recent work on the mechanisms underlying formation of BRCA1-linked TDs led us to discover a synthetic lethal interaction between BRCA1 and FANCM loss-of-function mutations. FANCM is a motor protein and, hence, an ATPase. We find that ablation of FANCM ATPase activity alone (leaving the rest of the protein intact and stable within the cell) is sufficient to confer lethality on BRCA1 mutant cells. Thus, FANCM may be a “druggable” target for therapy in BRCA1-linked cancer. In work proposed herein, we will define the therapeutic potential of this discovery. During the funding period, we expect to make important discoveries in this field and to open the door to new therapies in HBOC and perhaps other forms of cancer.

项目概要在复制叉停滞位点启动的无错误 DNA 修复对于预防基因组修复至关重要循环细胞的不稳定性与癌症易感性直接相关。和其他人类疾病与失速叉修复失败相关的临床负担可能包括遗传性。鉴于 BRCA1 和 BRCA2 参与修复乳腺癌和卵巢癌 (HBOC) 复制叉停滞和范可尼贫血 (FA)——一种罕见的常染色体隐性遗传（或 X 连锁）疾病我们之前的工作确定了 BRCA1 和 BRCA2 的作用。我们开发了创新工具，以调节双链断裂 (DSB) 和失速叉修复的 HR。用于量化哺乳动物复制停滞时的同源重组 (HR) 和其他修复结果该提案的一个主要目标是定义基本的分叉。我们开发了一系列尖端工具来支持这项研究，包括独特、成熟的人力资源生产者，他们可以区分无错误的“短程”人力资源和错误的人力资源倾向于“长链”HR——一种类似于酵母中断裂诱导复制的复制反应。我们在 BRCA1 突变细胞中特别是在停滞叉处观察到的异常复制反应是在一项范式转变的发现中，我们发现了 <10 kb 非同源串联重复 (TD) 的形成。这些高度特异性的结构变异形式在人类 BRCA1 相关乳房中也很丰富该提案的一个主要目标是定义遗传调控和完整的卵巢癌基因组。 BRCA1 突变细胞中停滞叉处 TD 形成的机制将在该项目中揭示。前所未有地详细描述了调节哺乳动物停滞（或损坏）叉修复的机制及其为支持这一点，我们将开发分析 DNA 的新技术。结构中间体、染色质对叉失速的反应以及失速哺乳动物的蛋白质组成这些分析研究还可能确定乳腺癌和乳腺癌治疗的新分子靶点。事实上，我们最近对 BRCA1 相关 TD 形成机制的研究引领了我们。发现 BRCA1 和 FANCM 功能丧失突变之间的合成致死相互作用是一项研究。我们发现 FANCM ATP 酶活性被单独消除（留下其余部分）。细胞内完整且稳定的蛋白质）足以致死 BRCA1 突变细胞。 FANCM 可能是治疗 BRCA1 相关癌症的“可药物”靶标。定义这一发现的治疗潜力，在资助期间，我们期望做出重要的贡献。该领域的发现为 HBOC 以及其他形式的癌症的新疗法打开了大门。