INVESTIGATING THE ROLE OF HSP90 IN SHAPING THE CONSEQUENCES OF BRCA1 MUTATIONS

项目总结 使DNA修复因子失活的突变增加了基因组的不稳定性和癌症的易感性。预测未来 突变的表型效应被普遍存在的多态和难以捉摸的环境修饰物混淆。 这一建议将通过利用一种基本的表观遗传机制来解决这个问题，该机制通过基因- 生物系统中的环境相互作用，蛋白质折叠。蛋白质必须正确折叠才能发挥作用；这是 这是在被称为蛋白质折叠伴侣的特殊蛋白质的帮助下实现的。在帮助蛋白质折叠和 功能，丰富的伴侣热休克蛋白90“缓冲”(即减轻)的有害突变的影响。 环境应激源使HSP90在细胞内的可获得性增加，从而揭示了突变体特有的隐蔽表型 道路。然而，HSP90对癌症突变的选择性仍然没有明确的定义。回答这个问题 将能够先发制人地识别人口中对环境敏感的基因类型以及 对将从使用HSP90抑制剂中受益的癌症患者进行分层。事实上，HSP90在 通过缓冲乳腺癌相关蛋白1(BRCA1)折叠突变体来提高乳腺癌细胞的耐药性。 靶向抑制HSP90可恢复癌细胞对遗传毒物的敏感性，并诱导基因组不稳定。 我假设HSP90缓冲了BRCA1中广泛范围的突变，并在这样做的过程中呈现了诱导 基因组的不稳定性取决于损害HSP90功能的环境应激源。我要测试一下这个 使用基于下一代功能基因组学和经典基因组学的强大方法 细胞遗传学方法学：1)评估伴侣结合作为BRCA1错义突变体分类器的作用 癌症患者的严重程度，2)确定BRCA1突变的HSP90缓冲情况，3)决定如何 HSP90影响BRCA1杂合子的条件性单倍体功能不全。每个目标都将利用高度具体的 HSP90抑制剂以无毒剂量给药，专门针对HSP90缓冲的突变体，而不是野生的 类型变量控件。我将改造非恶性和恶性细胞来表达HSP90缓冲的BRCA1 检测HSP90抑制对基因组稳定性和细胞活力的影响。这项工作不仅将 揭示形成BRCA1突变表现的基本机制，但也将建立一个 应用HSP90抑制恶化提高BRCA1靶向治疗疗效的实用途径 HSP90缓冲的基因组不稳定突变的影响。这项提议将为我的职业生涯做好准备。 独立科学家研究癌症突变扰乱人类健康的分子机制 DNA修复复合体在单分子分辨率下的功能。我已经成立了一个由专家组成的职业委员会 谁将在高通量系统生物学和功能基因组学、基因组不稳定性和癌症方面培训我 生物学、基因组工程和单分子方法。我的职业目标与MD安德森的 致力于博士后培训和教授和临床医生的支持网络致力于 精准抗癌药物研究进展。

PROJECT SUMMARY Mutations that inactivate DNA repair factors increase genome instability and cancer susceptibility. Predicting the phenotypic effects of mutations is confounded by pervasive polymorphisms and elusive environmental modifiers. This proposal will tackle this problem by utilizing a fundamental epigenetic mechanism that mediates gene-by- environment interactions in biological systems, protein folding. Proteins must properly fold to function; this is achieved with help from specialized proteins called protein-folding chaperones. In helping proteins fold and function, the abundant chaperone HSP90 “buffers” (that is – mitigates) the effects of deleterious mutations. Environmental stressors tax HSP90 availability within cells thus revealing cryptic phenotypes in a mutant-specific way. However, the selectivity of HSP90 for cancer mutations remains poorly defined. Addressing this question will enable preemptive identification of environmentally sensitive genotypes in the population as well as stratification of cancer patients who would benefit from the use of HSP90 inhibitors. Indeed, HSP90 mediates resistance in breast cancer cells by buffering a Breast Cancer Associated 1 (BRCA1) protein folding mutant. Targeted inhibition of HSP90 restores cancer cell sensitivity to genotoxic agents and induces genome instability. I hypothesize that HSP90 buffers a broad range of mutations in BRCA1, and in doing so renders the induction of genome instability conditional upon environmental stressors that impair HSP90 function. I will test this hypothesis by using a powerful approach based on next-generation functional genomics and classical cytogenetics methodologies to: 1) evaluate chaperone binding as a classifier of BRCA1 missense mutant severity in cancer patients, 2) determine the HSP90-buffered landscape of BRCA1 mutations, 3) determine how HSP90 influences the conditional haploinsufficiency of BRCA1 heterozygotes. Each aim will utilize highly specific HSP90 inhibitors administered at non-toxic doses to specifically target HSP90-buffered mutants and not wild- type variant controls. I will engineer non-malignant and malignant cells to express HSP90-buffered BRCA1 mutants and measure the effects of HSP90 inhibition on genome stability and cell viability. This work will not only reveal fundamental mechanisms shaping the manifestations of BRCA1 mutations, but will also establish a practical approach to improve the efficacy of BRCA1 targeted therapies using HSP90 inhibition to exacerbate the effects of HSP90-buffered genome instability mutations. This proposal will prepare me for a career as an independent scientist investigating the molecular mechanisms through which cancer mutations perturb the function of DNA repair complexes at single-molecule resolution. I have established a career committee of experts who will train me in high-throughput systems biology and functional genomics, genome instability and cancer biology, genome engineering, and single-molecule approaches. My career goals synergize with MD Anderson’s commitment to postdoctoral trainees and the supportive network of professors and clinicians dedicated to the advancement of precision cancer medicines.