The molecular mechanism of isocitrate dehydrogenase (lDH) mutations in cancer

描述（由申请人提供）：尽管肿瘤中的代谢改变在近一个世纪前首次被描述，但直到最近十年，代谢酶活性的变化才与肿瘤发生直接相关。异柠檬酸脱氢酶1和2 (IDH1和IDH2)，负责将异柠檬酸（ICT）转化为NADP+依赖的？-酮戊二酸盐（¿KG）在超过80%的成人II级和III级胶质瘤和继发性胶质母细胞瘤以及高达20%的急性髓性白血病中发生突变。有趣的是，许多IDH突变可能同时作为肿瘤抑制因子和致癌基因；迄今为止所描述的所有突变都导致ICT周转显著不足，但令人惊讶的是，许多突变也获得了一种新的新形态活性。许多IDH突变体不是正常的氧化脱羧，而是催化nadph依赖性的¿KG还原为所提出的肿瘤代谢物2-羟基戊二酸（2HG）。本提案旨在阐明致瘤性IDH突变的催化途径和结构特征，以阐明支持肿瘤发生的功能障碍的分子机制，使用预稳态动力学、生物物理方法、x射线晶体学、显微分光光度法和细胞分析。这将阐明缺乏正常活性和新形态活性在酶功能障碍中的作用，以及结构改变如何影响这些作用。此外，将探讨IDH突变引起氧化应激的可能性，以确定重要的致瘤途径。这些研究为设计有效的idh靶向治疗提供了重要的基础，并有助于预测患者的哪些突变可能适合这种治疗。长期的职业目标是建立一个独立的学术研究项目，研究代谢酶中酶功能障碍的分子机制，以及这些机制如何促进肿瘤的发生。要实现这一目标，两个主要的短期目标包括获得完成项目目标所需的重要培训，以及接受职业发展指导。已成立了一个由许多拟议技术专家组成的咨询委员会，以协助进行实验训练和数据分析，并在职业发展方面提供指导。这将补充博士后导师提供的重要指导和培训。进一步的支持将来自合作者，他们是本提案中描述的其余技术的专家。参加耶鲁大学的职业发展讲习班和研讨会，以及耶鲁大学，布鲁克海文国家实验室和冷泉港的课程将提供所需的培训和发展。最后，手稿准备、拨款写作、指导学生、演讲和在会议上发言的机会将进一步支持领导一个成功的独立研究项目的长期目标。通过获得实验培训和寻找职业发展机会，我将获得必要的工具来开展一个独立的研究项目，探索癌症代谢缺陷的分子机制。

DESCRIPTION (provided by applicant): Although metabolic alterations in tumors were first described nearly a century ago, it has only been within the last decade that changes in the activity of metabolic enzymes have been directly linked to tumorigenesis. Isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2), the enzymes responsible for the NADP+- dependent conversion of isocitrate (ICT) to ?-ketoglutarate (¿KG), are mutated in over 80% of adult grade II and grade III gliomas and secondary glioblastomas, and up to 20% of acute myeloid leukemias. Intriguingly, many IDH mutations may serve as both tumor suppressors and oncogenes; all mutations characterized to date result in significant deficiency in ICT turnover, but surprisingly, many also gain a novel neomorphic activity. Instead of the normal oxidative decarboxylation, many IDH mutants catalyze the NADPH-dependent reduction of ¿KG to the proposed oncometabolite, 2-hydroxyglutarate (2HG). This proposal seeks to elucidate the catalytic pathway and structural features of tumorigenic IDH mutations to elucidate the molecular mechanisms of dysfunction that support oncogenesis using pre-steady-state kinetics, biophysical methods, X-ray crystallography, microspectrophotometry, and cellular assays. This will clarify the contributions of deficient normal activity and neomorphic activity in enzymatic dysfunction, and how structural alterations influence these contributions. Further, the potential for IDH mutations to cause oxidative stress will be explored to identify important tumorigenic pathways. Such studies provide a critical foundation to aid the design of effective IDH-targeted therapy, and to help predict which mutations in patients will likely be amenable to such treatment. The long-term career goal is to establish an independent academic research program examining the molecular mechanisms of enzymatic dysfunction in metabolic enzymes, and how these mechanisms contribute to oncogenesis. To achieve this, two major short-term goals include gaining the significant training required to complete the aims of the project, and receiving career development mentoring. An advisory committee of experts of many of the techniques proposed has been assembled to assist in experimental training and data analysis and to provide mentorship in career development. This will supplement the vital mentoring and training provided by the postdoctoral mentor. Further support will come from collaborators, who are experts in the remaining techniques described in this proposal. Attendance at career development workshops and seminars at Yale University, and at courses at Yale University, the Brookhaven National Laboratory, and Cold Spring Harbor will provide required training and development. Finally, opportunities for manuscript preparation, grant writing, mentoring students, giving talks, and presenting at conferences will further support the long term goal of leading a successful independent research program. By gaining experimental training and seeking out opportunities for career development, I will acquire the tools necessary to develop an independent research program exploring the molecular mechanisms of defects in cancer metabolism.