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

项目摘要 破坏DNA修复因子的突变增加了基因组的不稳定性和癌症易感性。预测 突变的表型效应被普遍的多态性和难以捉摸的环境修饰剂所混淆。 这项提案将通过利用一种基本的表观遗传机制来解决这个问题， 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功能;这是 这是在一种叫做蛋白质折叠伴侣的特殊蛋白质的帮助下实现的。帮助蛋白质折叠， 功能，丰富的伴侣HSP 90“缓冲”（即减轻）有害突变的影响。 环境应激因素对细胞内HSP 90的利用率征税，从而揭示了在一种特定的细胞内的神秘表型。 路上了然而，HSP 90对癌症突变的选择性仍然不清楚。处理这个问题 将能够抢先识别人口中对环境敏感的基因型， 将受益于使用HSP 90抑制剂的癌症患者分层。事实上，HSP 90介导 通过缓冲乳腺癌相关1（BRCA 1）蛋白折叠突变体在乳腺癌细胞中的耐药性。 HSP 90的靶向抑制恢复癌细胞对遗传毒性剂的敏感性并诱导基因组不稳定性。 我假设HSP 90缓冲了BRCA 1中广泛的突变，这样做使得诱导 基因组的不稳定性取决于环境应激因素，这些因素会损害HSP 90的功能。我来测试一下 通过使用基于下一代功能基因组学和经典的 细胞遗传学方法：1）评估伴侣结合作为BRCA 1错义突变体的分类器 癌症患者的严重程度，2）确定BRCA 1突变的HSP 90缓冲区，3）确定如何 HSP 90影响BRCA 1杂合子的条件性单倍不足每个目标都将利用高度具体的 以非毒性剂量施用的HSP 90抑制剂特异性靶向HSP 90缓冲突变体而非野生型。 类型变量控件。我将设计非恶性和恶性细胞来表达HSP 90缓冲的BRCA 1 突变体，并测量HSP 90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BRCA 1突变表现的基本机制，但也将建立一个 使用HSP 90抑制剂改善BRCA 1靶向治疗的有效性的实用方法 HSP 90缓冲基因组不稳定性突变的影响。这个建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细胞的分子机制。 DNA修复复合物在单分子分辨率下的功能。我成立了一个职业委员会 他将在高通量系统生物学、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 致力于博士后培训人员和教授和临床医生的支持网络，致力于 精准抗癌药物的进步。

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.