Defining the role of Mst1/Mst2 in regulating metabolic alterations in Ras driven NSCLC

定义 Mst1/Mst2 在调节 Ras 驱动的 NSCLC 代谢改变中的作用

• 批准号: 10323278
• 负责人: Jennifer Bailey Lundberg
• 金额: $ 17.98万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10323278
• 关键词: Address; Adenocarcinoma; Aerobic; Alveolar; Area; Biological Assay; Biology; Cancer Etiology; Cell Respiration; Cell Survival; Cells; Cessation of life; Copy Number Polymorphism; Data; Development; Diagnosis; Diagnostic tests; Dimerization; Early treatment; Environment; Enzymes; Epithelial; Epithelial Cells; Event; FGFR1 gene; Future; Genetic; Genetic Models; Genus Hippocampus; Glycolysis; Goals; Grant; Head and neck structure; Health; Histologic; Human; Immunohistochemistry; In Vitro; Incidence; Investigation; KRAS2 gene; KRASG12D; Knowledge; Laboratories; Life; Lung; Lung Adenocarcinoma; Lung Adenoma; Malignant Neoplasms; Malignant neoplasm of lung; Metabolic; Metabolic Pathway; Metabolism; Mission; Modeling; Modification; Molecular; Mus; Mutation; Neoplasms; Non-Small-Cell Lung Carcinoma; Nuclear; Nuclear Translocation; Oncogenes; Oncogenic; Oxygen Consumption; Pathway interactions; Patient-Focused Outcomes; Patients; Phosphorylation; Phosphotransferases; Post-Translational Protein Processing; Production; Proteomics; Public Health; Publications; Published Comment; Publishing; Pyruvate; Pyruvate Kinase; Research; Role; Signal Pathway; Squamous cell carcinoma; Structure of parenchyma of lung; TP53 gene; Testing; The Cancer Genome Atlas; Therapeutic; Tissues; Transcription Coactivator; Transgenic Model; Type II Epithelial Receptor Cell; United States National Institutes of Health; adenoma; alveolar epithelium; base; cancer invasiveness; cell transformation; experimental study; glucose metabolism; human data; human model; improved; in vivo; knock-down; member; metabolomics; mouse model; mutant; novel; programs; public health relevance; response; targeted treatment; tumor

PROJECT SUMMARY/ABSTRACT Non-small cell lung cancer (NSCLC) remains the most commonly diagnosed malignancy and leading cause of cancer related deaths worldwide; yet there is a fundamental gap in understanding the biology of lung cancer formation. Sequencing data of human NSCLC revealed KRAS is one of the most frequent oncogene aberrations in lung adenocarcinoma patients. We have recently generated a novel mouse model of NSCLC driven by an oncogenic mutation in KRASG12D and concurrent genetic deletion of MST1/2. When mice express mutant KRASG12D alone in alveolar epithelial type II cells (AECII), we observe focal lung adenomas and neoplasia; however, when MST1/2 are genetically deleted in combination with KRASG12D, we observe aggressive lung adenocarcinoma. To define early mechanisms by which loss of MST1/2 accelerates KRASG12D driven NSCLC, we performed a proteomics screen prior to observing NSCLC on lung tissue from our mouse models. Our proteomic analysis revealed pyruvate kinase M2 (PKM2), a rate-limiting enzyme during glycolysis that catalyzes the production of pyruvate and ATP, is highly expressed, even before the onset of tumors. As improved targeted therapies and diagnostic tests have the potential to significantly improve patient outcomes, we now want to study the mechanism by which loss of MST1/2 promotes increased PKM2. Our central hypothesis is reduced expression of MST1/2 is important in the development of NSCLC. We predict loss of MST1/2 promotes increased abundance and nuclear localization of PKM2, to promote altered glucose metabolism and survival. We plan to test our overall hypothesis by pursuing the following specific aims: Aim 1: To test the hypothesis that MST1 restrains aerobic glucose metabolism in KRAS mutant epithelial cells. Rationale: We have generated new preliminary data that inhibition of MST1 increases PKM2 and pPKM2 in cultured lung epithelial cells expressing KRASG12D. We also have new data using Seahorse assays that inhibition of MST1 significantly increases oxygen consumption rate and ATP production in cultured human NSCLC. We now want to conduct in vivo and in vitro metabolomic analysis to characterize MST1-dependent metabolic programs in human and mouse models driven by mutant KRAS. Aim 2: To test the hypothesis that YAP/TAZ are required for mutant KRAS induction of PKM2. Rationale: We have generated preliminary data using transient knockdown of PKM2 which revealed decreased metabolic activity and proliferation in human NSCLC. In this aim, we will determine what molecular events are causing posttranslational modifications on PKM2 to promote dimerization and nuclear translocation. Using genetic knockdown, we will determine if YAP or TAZ are required for increased expression of PKM2. We will use Seahorse analysis and mass spec to study PKM2 modifications and other metabolic alterations in YAP or TAZ deficient cells. In addition, in response to reviewer comments, we are crossing PKM2fl/fl mice into our genetic model to define the requirement for PKM2 in adenoma-adenocarcinoma development in vivo.

项目总结/摘要 非小细胞肺癌（NSCLC）仍然是最常诊断的恶性肿瘤，也是肺癌的主要原因。 全世界与癌症相关的死亡;然而，在理解肺癌的生物学方面存在根本性的差距 阵人类NSCLC的测序数据显示KRAS是最常见的癌基因畸变之一 在肺腺癌患者中。我们最近建立了一种新的NSCLC小鼠模型， KRASG 12 D的致癌突变和MST 12的同时遗传缺失。当小鼠表达突变体 KRASG 12 D单独在肺泡上皮II型细胞（AECII）中，我们观察到局灶性肺腺瘤和瘤形成; 然而，当MST 12与KRASG 12 D组合遗传缺失时，我们观察到侵袭性肺 腺癌为了确定MST 12缺失加速KRASG 12 D驱动的NSCLC的早期机制， 在观察来自我们的小鼠模型的肺组织上的NSCLC之前，我们进行了蛋白质组学筛选。我们 蛋白质组学分析揭示了丙酮酸激酶M2（PKM 2），一种糖酵解过程中的限速酶， 丙酮酸和ATP的产生，甚至在肿瘤发生之前就高度表达。作为改进的目标 治疗和诊断测试有可能显着改善患者的结果，我们现在想 研究MST 1/2缺失促进PKM 2增加的机制。我们的中心假设被简化了 MST 1/2的表达在NSCLC的发生发展中起重要作用。我们预测MST 1/2启动子的丢失 增加PKM 2的丰度和核定位，以促进改变葡萄糖代谢和存活。 我们计划通过追求以下具体目标来测试我们的总体假设：目标1：测试假设 MST 1抑制KRAS突变上皮细胞中的有氧葡萄糖代谢。理由：我们有 产生了新的初步数据，抑制MST 1增加了培养的肺上皮细胞中的PKM 2和pPKM 2， 表达KRASG 12 D的细胞。我们也有新的数据使用海马测定，抑制MST 1显着 增加培养的人NSCLC的耗氧率和ATP产生。我们现在要进行 体内和体外代谢组学分析，以表征人类中MST 1依赖性代谢程序， 由突变KRAS驱动的小鼠模型。目的2：检验以下假设： PKM 2的突变KRAS诱导。基本原理：我们已经使用瞬时敲低产生了初步数据 PKM 2的表达，其揭示了人NSCLC中代谢活性和增殖的降低。为此，我们将 确定哪些分子事件导致PKM 2上的翻译后修饰以促进二聚化 和核转位。使用基因敲低，我们将确定是否需要雅普或TAZ增加 PKM 2的表达。我们将使用Seahorse分析和质谱来研究PKM 2修饰和其他 雅普或TAZ缺陷细胞中的代谢改变。此外，为了回应评论者的意见，我们 将PKM 2fl/fl小鼠与我们的遗传模型杂交以确定腺瘤-腺癌中对PKM 2的需求 体内发育