Metabolic Recycling and Compartmentalization in Tumor Progression

肿瘤进展中的代谢循环和区室化

基本信息

批准号：
10356031
负责人：
Jessica Brooke Spinelli
金额：
$ 10.52万
依托单位：
WHITEHEAD INSTITUTE FOR BIOMEDICAL RES
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-03-01 至 2022-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10356031
关键词：
Address Amino Acids Ammonia Aspartate Assimilations Award Biochemical Bioinformatics Biomass Breast Cancer Cell Breast Cancer Patient Budgets CRISPR screen Cancer Model Carbon Catabolism Clinical Collaborations Consumption Core Facility Data Dependence Drops Energy Supply Environment Equilibrium Equipment Family Fellowship Genetic Genetic Models Glucose Glutamate Dehydrogenase Glutamate Metabolism Pathway Glutamates Glutaminase Glutamine Growth Growth and Development function Homeostasis Immunoprecipitation In Vitro Inner mitochondrial membrane Institutes KRAS2 gene Lead Life Expectancy Malate-Aspartate Shuttle Pathway Malignant Neoplasms Mass Spectrum Analysis Mediating Mentorship Messenger RNA Metabolic Metabolic Pathway Metabolism Mitochondria Modeling Mus Mutation NADH Nitrogen Normal tissue morphology Nucleotide Biosynthesis Nutrient Oncogenic Outcome Pathway interactions Patients Phase Play Production Proteins Protocols documentation Recycling Research Resistance Resources Role Sampling Science Source Technology Testing Therapeutic Tissues Training Translations Tumor Subtype Work Xenograft Model antiporter cancer cell cancer subtypes career development experience follow-up improved in vivo inhibitor macromolecule malignant breast neoplasm medical schools meetings member metabolic fitness mitochondrial metabolism neoplastic cell novel pyruvate carrier rapid growth research and development responsible research conduct skills solute stable isotope symposium targeted cancer therapy therapy development three dimensional cell culture treatment response tumor tumor growth tumor initiation tumor metabolism tumor microenvironment tumor progression tumorigenesis tumorigenic uptake validation studies wasting

项目摘要

PROJECT SUMMARY Cancer cells elevate nutrient catabolism, causing excess production and accumulation of metabolic waste, especially lactate and ammonia. Using stable isotope tracing and mass spectrometry, we discovered that breast cancer cells scavenged ammonia generated by amino acid catabolism as a re-purposed nitrogen source for biomass (Spinelli et al. Science, 2017). Ammonia recycling accelerated proliferation in 3D cell culture and in vivo mouse xenograft models. These studies lead to two follow-up questions that will be addressed in Specific Aim 1 (F99 phase) of this proposal. First (Aim 1A), what is the mechanism by which ammonia stimulates breast cancer proliferation? Preliminary data supports the hypothesis that subcellular compartmentalization of ammonia metabolism is required for its effect on proliferation rate. Using metabolic tracing and rapid immunoprecipiation, we tracked the localization of ammonia assimilation to the mitochondria and the subsequent efflux of metabolites to the cytosolic fraction, showing a role in ammonia-stimulated proliferation. Second (Aim 1B), what is the effect of this novel ammonia-recycling pathway on response to therapy? Preliminary data shows that ammonia assimilation circumvents the effect of glutaminase (GLS) inhibitors through replenishing glutamate levels via glutamate dehydrogenase (GDH). Our proposed studies will determine if GDH and GLS inhibition are synergistic in vivo and in primary breast cancer tumors to elucidate a mechanism by which breast cancer cells are resistant to GLS inhibition. Beyond ammonia, mitochondria promote tumor growth and proliferation through numerous metabolic pathways. However, the mechanisms by which nutrients traverse the inner mitochondrial membrane are little studied, particularly because 45% of the mitochondria nutrient transporter family is uncharacterized. Therefore, using my expertise in metabolite tracing and mass spectrometry in addition to new skills gained in genetic (CRISPR) screening and bioinformatics, the proposed studies (Aim 2, K00 phase) will systematically assess the role of mitochondrial nutrient transporters in tumor growth and survival. These data will be the first to globally evaluate the essentiality of metabolic compartmentalization in cancer cells harboring oncogenic drivers such as KRAS, IDH and PI3K mutations. Furthermore, this study will elucidate the function of uncharacterized mitochondrial nutrient transporters. In addition to the proposed studies, the fellowship training plan includes gaining experience with mentorship, taking courses on responsible conduct of research, team management, and budgeting, and attending scientific conferences such as the Tumor Metabolism Keystone meeting to develop a network of scientific collaborators. The proposed studies will occur at Harvard Medical School and The Broad Institute, which have environments with the equipment, technology, core facilities, potential for collaboration, and the resources for research and career development needed to complete the proposed training plan.

项目概要癌细胞提高营养物质的分解代谢，导致代谢物的过量产生和积累废物，特别是乳酸和氨。使用稳定同位素示踪和质谱分析，我们发现乳腺癌细胞清除氨基酸分解代谢产生的氨作为重新利用的氮生物质来源（Spinelli 等人 Science，2017）。氨回收加速 3D 细胞增殖培养和体内小鼠异种移植模型。这些研究引出了两个后续问题本提案的具体目标 1（F99 阶段）中已解决。首先（目标 1A），其机制是什么？氨会刺激乳腺癌增殖吗？初步数据支持以下假设：亚细胞氨代谢的区室化对于其对增殖率的影响是必需的。利用代谢通过追踪和快速免疫沉淀，我们追踪了氨同化在线粒体中的定位以及随后代谢物流出至胞质部分，显示出在氨刺激中的作用增殖。其次（目标 1B），这种新型氨循环途径对响应的影响是什么？治疗？初步数据表明氨同化规避了谷氨酰胺酶 (GLS) 的作用通过谷氨酸脱氢酶（GDH）补充谷氨酸水平来抑制抑制剂。我们提出的研究将确定 GDH 和 GLS 抑制在体内和原发性乳腺癌肿瘤中是否具有协同作用阐明乳腺癌细胞抵抗 GLS 抑制的机制。除了氨之外，线粒体还通过多种代谢促进肿瘤生长和增殖途径。然而，营养物质穿过线粒体内膜的机制却很少。研究，特别是因为 45% 的线粒体营养转运蛋白家族是未知的。所以，除了在遗传方面获得的新技能之外，还利用我在代谢物追踪和质谱方面的专业知识（CRISPR）筛选和生物信息学，拟议的研究（Aim 2，K00阶段）将系统评估线粒体营养转运蛋白在肿瘤生长和存活中的作用。这些数据将首先全面评估含有致癌驱动因素的癌细胞代谢区室化的必要性例如 KRAS、IDH 和 PI3K 突变。此外，本研究将阐明未表征的功能线粒体营养转运蛋白。除了拟议的研究之外，奖学金培训计划还包括获得以下方面的经验：指导，参加有关负责任的研究行为、团队管理和预算的课程，以及参加肿瘤代谢基石会议等科学会议，以建立一个网络科学合作者。拟议的研究将在哈佛医学院和布罗德研究所进行，具备设备、技术、核心设施、合作潜力、完成拟议的培训计划所需的研究和职业发展资源。