Mitochondrial Metabolism and ROS Regulate Lung Cancer

描述(申请人提供)：肺癌是美国男性和女性癌症相关死亡的最常见原因。80%的肺癌是非小细胞肺癌(NSCLC)，30%的NSCLC是腺癌，其中发生在非吸烟者的癌症病例数量不断增加。在20-30%的非小细胞肺癌中发现了Kras癌基因的突变，Kras癌基因通过一系列蛋白激酶发出促进细胞增殖的信号。针对Kras驱动的肿瘤的治疗靶向需要确定致癌Kras驱动的增殖所需的信号通路。KRAS驱动的肺癌细胞表现出比非癌症肺细胞更高的活性氧物种(ROS)水平。ROS可作为信号分子激活多种信号通路，包括PI3K、ERK1/2、MAPK和转录因子低氧诱导因子(HIF)，促进肿瘤细胞增殖、血管生成和转移。胞质中参与信号转导的ROS的主要形式是过氧化氢(H_2O_2)，它是由胞质中的铜锌超氧化物歧化酶(SOD_1)从超氧化物(O_2~-)转化而来的。我们已经报道，线粒体复合体III中的O2-及其在胞浆中转化为H2O2是启动Kras诱导的细胞增殖和肿瘤细胞中HIFs低氧激活所必需的。目前，尚不清楚在体内致癌的Kras驱动的肺肿瘤发生中，胞浆中是否需要复合体III产生的O2或H2O2依赖的信号。最近的研究表明，谷氨酰胺是TCA循环产生NADH和FADH2的主要燃料。这些还原等价物向电子传输链提供电子，导致络合物III生成超氧化物。我们最近报道，Kras驱动的肿瘤细胞也利用谷氨酰胺为TCA循环提供燃料。谷氨酰胺可以被谷氨酰胺酶(GLS)转化为谷氨酸，谷氨酸在氨基转移酶(GPT2或GOT2)或谷氨酸脱氢酶(GDH)的作用下转化为α-酮戊二酸，进入TCA循环。初步数据表明，使用GPT2的氨基转移酶或RNAi抑制剂阻止谷氨酰胺进入TCA循环可减少肿瘤细胞Kras驱动的肿瘤细胞的锚定非依赖性生长。然而，目前尚不清楚抑制GPT2是否会减轻体内的肺腺癌，或者GPT2对于成年小鼠的正常组织是否是必不可少的。这笔赠款的主要目标是从基因上确定减少TCA循环产生的超氧化物、胞浆过氧化氢的产生和谷氨酰胺的利用是否会减少致癌Kras驱动的小鼠肺腺癌模型和使用携带Kras突变的人A549肺腺癌细胞的原位小鼠模型的肿瘤形成。 公共卫生相关性：肺癌是美国男性和女性癌症相关死亡的最常见原因。主要目标是确定阻止线粒体新陈代谢或过氧化氢的产生是否对减少肺癌有效。积极的结果将为针对调节线粒体新陈代谢的代谢酶的治疗提供理论基础。

DESCRIPTION (provided by applicant): Lung cancer is the most common cause of cancer-related death in both men and women in the United States. Eighty percent of lung cancers are non-small cell lung cancers (NSCLCs) and 30% of NSCLCs are adenocarcinomas, which include a rising population of cancer cases that occur in nonsmokers. Mutations in the Kras oncogene, which signals through a cascade of kinases to promote cellular proliferation, have been identified in 20-30% of NSCLCs. Therapeutic targeting of Kras-driven tumors necessitates the identification of signaling pathways required for oncogenic Kras-driven proliferation. Kras-driven lung cancer cells display higher levels of reactive oxygen species (ROS) than noncancerous lung cells. ROS have been proposed to serve as signaling molecules to activate numerous signaling pathways, including PI3K, ERK1/2 MAPK, and the transcription factors hypoxia inducible factors (HIFs), which promote tumor cell proliferation, angiogenesis, and metastasis. The major form of ROS that participates in signaling in the cytosol is hydrogen peroxide (H2O2), which is generated by its conversion from superoxide (O2-) by copper-zinc superoxide dismutase (SOD1) in the cytosol. We have reported that O2- from mitochondrial complex III and its conversion to H2O2 in the cytosol are required to initiate Kras-induced cellular proliferation and hypoxic activation of HIFs in tumor cells. Presently, it is not known whether complex III-generated O2- or H2O2-dependent signaling in the cytosol is required for oncogenic Kras-driven lung tumorigenicity in vivo. Recent studies indicate that glutamine is a major fuel for the TCA cycle to generate NADH and FADH2. These reducing equivalents donate electrons to the electron transport chain resulting in complex III generated superoxide. We recently reported that Kras-driven tumor cells also utilize glutamine to fuel the TCA cycle. Glutamine can be converted by glutaminase (GLS) to glutamate, which enters the TCA cycle through conversion into alpha-ketoglutarate by aminotransferases (GPT2 or GOT2) or glutamate dehydrogenase (GDH). Preliminary data indicate that preventing glutamine entry into the TCA cycle using inhibitors of aminotransferases or RNAi of GPT2 reduces anchorage-independent growth of oncogenic Kras-driven tumor cells. However, it is not known if inhibition of GPT2 would attenuate lung adenocarcinoma in vivo or if GPT2 is dispensable for normal tissues in the adult mouse. The major goal of this grant is to genetically determine whether diminishing complex III generated superoxide, production of cytosolic hydrogen peroxide, and glutamine utilization by the TCA cycle will attenuate tumorigenesis in the oncogenic Kras-driven mouse model of lung adenocarcinoma and in an orthotopic mouse model using human A549 lung adenocarcinoma cells harboring a Kras mutation. PUBLIC HEALTH RELEVANCE: Lung cancer is the most common cause of cancer-related death in both men and women in the United States. The major goal is to determine whether preventing mitochondrial metabolism or production of hydrogen peroxide would be effective in diminishing lung cancer. Positive results would provide a rationale for therapies targeting metabolic enzymes that regulate mitochondrial metabolism.