Deciphering the mechanism underlying BRCA1 breast cancer development

项目总结/摘要 遗传性BRCA 1突变通常会导致受影响家庭中的乳腺癌和卵巢癌的高频率。 这项提议旨在阐明无肿瘤、BRCA 1突变的乳腺癌患者月经初潮后发生的关键步骤。 乳腺上皮并在很大程度上促进未来BRCA 1乳腺癌（BrCa）的发展。 我们已发表的数据和正在进行的工作表明存在一系列相互关联的生化步骤 当中断时，导致表面正常的BRCA 1突变乳腺上皮细胞的转化， 进入一种异常的、去分化的、异常原始的、可能是癌前状态。这一变化涉及到 一组6个特定基因中任何一个成员的功能丧失，转化为DNA的破坏 损伤控制，其反过来促进乳腺上皮分化的破坏。乳腺细胞中 这些基因通常在功能上相互交流，就好像它们是生物学上的一个组成部分。 这是一个重要的途径，部分致力于BrCa抑制。事实上，它们的蛋白质产物形成了一种独特的 一种分子复合物，也含有BRCA 1肿瘤抑制蛋白p220，这种复合物的作用是 在这一过程中它扮演着重要的角色。 在其他6个复合体成员（BRG 1，FancD 2，CtIP，NUMB，HES-1和BARD 1）中，一些（例如FancD 2， BRG 1）在散发性BrCa中发生体细胞突变。另一种（Numb）是动物中已知的BrCa抑制剂， 正常干细胞增殖的支持者，并且6个中的至少5个必须存在并与p220结合， 整个复合体形成和乳腺上皮分化正常进行。我们正在进行的工作也 这表明这种复合物的完整性是防止BRCA 1 BrCa发育所必需的。 拟议的研究将测试这一假设的有效性，沿着这样的生理重要性， 路径，使用尖端方法的组合。包括将努力a）测试血统 乳腺上皮细胞（MEC）DNA损伤修复失败与MEC去分化的关系 MEC去分化和BRCA 1肿瘤发展之间的关系，以及B）解释MEC去分化和BRCA 1肿瘤发展之间的调控机制， 这些事件的相互关系。 这些努力的一个主要组成部分将是应用单细胞RNAseq来评估上述基因。 谱系关系和以乳腺上皮为中心的基因组CRISPR和蛋白质过表达筛选 检测参与BRCA 1 BrCa发育中这些重要步骤的单个蛋白质。我们将使用 一个新开发的小鼠模型在这些实验中。

Project Summary/Abstract Inherited BRCA1 mutations very often result in high frequency breast and ovarian cancer in affected families. This proposal is aimed at illuminating key steps that occur after menarche in tumor-free, BRCA1-mutated mammary epithelium and contribute in a major way to future BRCA1 breast cancer (BrCa) development. Our published data and work in progress suggest the existence of a series of interrelated biochemical steps that, when interrupted, result in the conversion of ostensibly normal, BRCA1- mutated breast epithelial cells into an abnormal, dedifferentiated, unusually primitive, and likely pre-malignant state. This change involves the loss of function of any member of a group of 6 particular genes which translates into a breakdown in DNA damage control which, in turn, promotes a breakdown in mammary epithelial differentiation. In breast cells these genes normally communicate with one another, functionally, as if they were components of a biologically important pathway dedicated, in part, to BrCa suppression. Indeed, their protein products form a distinct molecular complex which also contains the BRCA1 tumor suppressor protein, p220, and this complex operates in ways that suggest a major role for it in such a pathway. Among the 6 other complex members (BRG1, FancD2, CtIP,NUMB, HES-1, and BARD1), some (e.g. FancD2, BRG1) are somatically mutated in sporadic BrCa. Another (Numb) is a known BrCa suppressor in animals and a supporter of normal stem cell proliferation, and at least 5 of the 6 must be present and bind to p220 for the entire complex to form and mammary epithelial differentiation to proceed normally. Our work in progress also suggests that the integrity of this complex is required to prevent BRCA1 BrCa from developing. The proposed research will test the validity of this hypothesis along with the physiological importance of such a pathway, using a combination of cutting edge methods. Included will be an effort to a) test for lineage relationships between failed mammary epithelial cell (MEC) DNA damage repair and MEC dedifferentiation and between MEC dedifferentiation and BRCA1 tumor development and b) to decipher the mechanisms governing the interrelationships of these events. A major component of these efforts will be the application of single cell RNAseq to assess the above-noted lineage relationships and mammary epithelium- focused genomic CRISPR and protein overexpression screens to detect individual proteins that participate in these important steps in BRCA1 BrCa development. We will use a newly developed mouse model in these experiments.!