(PQ5) Imaging mitochondrial heterogeneity in LKB1 mutant lung cancer

摘要 这项研究的目的是在GEMM中利用PET成像来执行线粒体的机制研究 肺癌发生过程中LKB1/AMPK信号通路失活导致的异质性。 LKB1是一种主激酶，通过以下途径调节细胞能量和线粒体功能 激活在癌症中经常发生突变的腺苷一磷酸活化激酶(AMPK)。 LKB1突变导致AMPK信号通路失活，导致细胞严重缺陷 能量学和线粒体动态平衡。这导致了人类体内高度可变的线粒体池和 由许多大小、形态和功能不同的非典型线粒体组成的小鼠肿瘤 我们将其定义为线粒体异质性。然而，在生理或机械水平上对此了解甚少。 LKB1失活导致的线粒体异质性如何影响肺肿瘤的发生或治疗。我们 用电子耦合技术检测体内肺肿瘤线粒体结构和功能的异质性 Lkb1基因工程小鼠的显微(EM)和正电子发射断层扫描(PET)成像 模型(GEM)。使用电压敏感的线粒体特异性放射性示踪剂[18F]-氟苯基- 三苯基膦(FTp)我们能够测量肺肿瘤的线粒体膜电位(∆Ψ)。 宠物成像。FTPET成像发现肺肿瘤细胞线粒体活性不均一 活着。此外，线粒体缺陷使LKB1-/-肿瘤细胞敏感地经历线粒体外部 膜通透性(MOMP)和细胞凋亡，我们发现LKB1/AMPK通路是一个潜在的 MOMP的调节和通过电压依赖性阴离子1(VDAC1)诱导的细胞凋亡。最后，由于 合成致死化学筛选，我们鉴定了蛋白质酪氨酸磷酸酶线粒体1(PTPMT1)的关键基因 心磷脂生物合成和线粒体完整性调节作为LKB1-/-肺治疗的新靶点 癌症。我们假设LKB1抑癌基因的失活导致线粒体的异质性 驱动肺癌发展的结构和功能。为了检验这一假设，我们将把PET和 肺癌Lkb1-/-GEMM的EM成像纵向研究不同部位线粒体的异质性 肺肿瘤的发展阶段。在Aim1中，我们将使用FTPPET成像来定位线粒体的异质性 以及LKB1缺失后肺肿瘤发生过程中的体内动力学。在目标2中，我们将识别分子 LKB1/AMPK途径调节线粒体外膜的机制。在AIM 3中 对LKB1-/-肺肿瘤的PTPMT1-心磷脂通路进行体内解剖。我们建议先入行- 实地研究将促进我们对线粒体生物学的基本理解，以及 线粒体的异质性对肺癌的发生有促进作用。拟议的工作与以下方面有关 人类健康在基于PET成像的肺癌检测诊断领域以及 开发新的治疗方法以改善肺癌患者的预后。

ABSTRACT The goal of this study is to utilize PET imaging in GEMMs to perform a mechanistic study of mitochondrial heterogeneity following inactivation of the LKB1/AMPK signaling pathway during lung tumor development. LKB1 functions as a master kinase that regulates cellular energetics and mitochondrial function through activation of the adenosine monophosphate activated kinase (AMPK) that is frequently mutated in cancer. LKB1 mutations lead to inactivation of the AMPK signaling pathway resulting in severe defects in cellular energetics and mitochondrial homeostasis. This results in highly variable mitochondrial pools within human and mouse tumors that consist of numerous atypical mitochondria of differing size, morphology and function that we define as mitochondrial heterogeneity,. However, little is understood at a physiological or mechanistic level how mitochondrial heterogeneity resulting from LKB1 inactivation impact lung tumorigenesis or therapy. We examined mitochondrial structural and functional heterogeneity in lung tumors in vivo by coupling electron microscopy (EM) and positron emission tomography (PET) imaging of Lkb1-/- genetically engineered mouse models (GEMMs). Using a voltage sensitive mitochondrial specific radiotracer [18F]-Fluorobenzyl- triphenylphosphonium (FTP) we are able to measure mitochondrial membrane potential (∆Ψ) in lung tumors by PET imaging. FTP PET imaging identified lung tumor populations with heterogeneous mitochondrial activity in vivo. Additionally, mitochondrial defects sensitize LKB1-/- tumor cells to undergo mitochondrial outer membrane permeabilization (MOMP) and apoptosis and we discovered the LKB1/AMPK pathway is a potential regulator of MOMP and apoptosis through voltage dependent anion 1 (VDAC1). Lastly, as a result of a synthetic lethal chemical screen, we identified protein tyrosine phosphatase mitochondria 1 (PTPMT1), a key regulator of cardiolipin biosynthesis and mitochondrial integrity as a novel therapeutic target in LKB1-/- lung cancer. We hypothesize that inactivation of the LKB1 tumor suppressor induces heterogeneity in mitochondrial structure and function that drives lung tumor development. To test this hypothesis we will integrate PET and EM imaging of Lkb1-/- GEMMs of lung cancer to longitudinally study mitochondrial heterogeneity at distinct stages of lung tumor development. In Aim1 we will use FTP PET imaging to map mitochondrial heterogeneity and dynamics in vivo during lung tumorigenesis following LKB1 loss. In Aim 2 we will identify the molecular mechanisms by which the LKB1/AMPK pathway regulates the mitochondrial outer membrane. In Aim 3 perform an in vivo dissection of the PTPMT1-cardiolipin pathway in LKB1-/- lung tumors. We propose first-in- field studies that will advance our fundamental understanding of mitochondrial biology and the impact of mitochondrial heterogeneity has on promoting lung tumorigenesis. The proposed work has relevance to human health in the areas of PET imaging based detection diagnosis of lung cancer as well as the development of new therapies to improve outcomes for lung cancer patients.