Targeting Fnip1 to disrupt B cell development, metabolism, and transformation

描述（由申请人提供）：相对于静止的淋巴细胞，活化的淋巴细胞和癌细胞在细胞代谢方面都表现出独特的转变，从氧化磷酸化（有效地产生能量）到有氧糖酵解(产生燃料细胞分裂所需的生物前体（如脂质、氨基酸和核苷酸）。了解控制这种代谢开关的因素（称为“Warburg效应”）是非常重要的，因为它可能导致在自身免疫性疾病中选择性阻断淋巴细胞激活和/或抑制癌细胞存活的新策略。在本申请中，我们建议研究一种名为卵泡蛋白相互作用蛋白-1 （Fnip1）的新蛋白，我们的研究表明，在能量和营养应激（如淋巴细胞激活、营养限制和癌基因激活）期间，该蛋白对于维持“代谢平衡”至关重要。基于外周血中B淋巴细胞的完全缺失，我们在化学诱变筛选中发现了一种缺乏Fnip1的创新小鼠新品系。Fnip1缺失小鼠在b前细胞和不变自然杀伤T （iNKT）细胞发育阶段阻滞，细胞通常依赖于c-Myc进行大量分裂，c-Myc是人类许多癌症中不受调控的癌基因。值得注意的是，在伯基特B细胞淋巴瘤小鼠模型中，Fnip1的缺失也可以防止c-Myc诱导的前B细胞淋巴瘤。虽然Fnip1的功能尚不清楚，但它与卵泡蛋白（一种功能未知的蛋白质）和主要代谢调节剂AMP激酶相互作用，AMP激酶是一种能量感应分子，在能量应激下刺激能量产生，并抑制哺乳动物雷帕霉素靶蛋白（mTOR）调节的能量消耗合成代谢过程。我们的长期目标是确定Fnip1如何控制淋巴细胞的发育、激活、代谢和转化。我们的具体目标是：(1)明确Fnip1在前b细胞发育和代谢中的作用。我们将利用代谢组学、代谢通量分析和转录组学方法来确定Fnip1的缺失是否会抑制“Warburg效应”；(2)探讨Fnip1在B细胞淋巴瘤存活及对代谢应激和化疗药物敏感性中的重要性。我们将在原发性小鼠B细胞淋巴瘤中有条件地删除Fnip1，并将确定营养剥夺和化疗对肿瘤细胞存活和信号传导的影响；(3)描述Fnip1在自噬和mTOR信号通路中的分子功能。我们将使用生化和遗传方法来确定Fnip1是否对“关闭”mTOR介导的营养消耗和“打开”自噬（细胞器自我消化以产生营养物质）以应对营养不足至关重要。这些研究将解决我们的整体创新假设，即抑制Fnip1通过允许活化淋巴细胞和/或肿瘤细胞在缺乏足够能量和生物底物的情况下生长，从而“断开”合成代谢细胞生长和有氧糖酵解之间的基本联系，导致“营养耗尽”和细胞死亡。

DESCRIPTION (provided by applicant): Relative to resting lymphocytes, both activated lymphocytes and cancer cells exhibit a unique shift in cell metabolism from oxidative phosphorylation, which efficiently produces energy, to aerobic glycolysis, which generates bio-precursors (such as lipids, amino acids, and nucleotides) required to fuel cell division. An understanding of the factors that control this metabolic switch (termed "Warburg effect") is highly significant because it could lead to novel strategies to selectively block lymphocyte activation in autoimmune disease, and/or inhibit cancer cell survival. In this application, we propose to investigate a novel protein called Folliculin Interacting protein-1 (Fnip1) which our studies suggest is essential for maintaining "metabolic balance" during energy and nutrient stress such as during lymphocyte activation, nutrient restriction, and oncogene activation. We identified an innovative new strain of mice lacking Fnip1 in a chemical mutagenesis screen, based on the complete absence of B lymphocytes in peripheral blood. Fnip1 null mice have blocks in pre-B cell and invariant natural killer T (iNKT) cell development at stages where the cells normally undergo massive division dependent on c-Myc, an oncogene deregulated in many cancers in humans. Remarkably, loss of Fnip1 also protects against pre-B cell lymphoma induced by c-Myc in a mouse model of Burkitt's B cell lymphoma. Although the functions of Fnip1 are unknown, it interacts with Folliculin (a protein of unknown function) and the master metabolic regulator AMP kinase, an energy sensing molecule that stimulates energy production in response to energy stress and inhibits energy-consuming anabolic processes regulated by mammalian target of rapamycin (mTOR). Our long-term goals are to determine how Fnip1 functions to control the development, activation, metabolism, and transformation of lymphocytes. Our Specific Aims are: (1) To define the roles of Fnip1 in pre-B cell development and metabolism. We will utilize metabolomic, metabolic flux analysis, and transcriptomic approaches to determine whether loss of Fnip1 inhibits the "Warburg effect"; (2) To determine the importance of Fnip1 in B cell lymphoma survival and sensitivity to metabolic stress and chemotherapeutic agents. We will conditionally delete Fnip1 in primary murine B cell lymphomas and will determine consequences on tumor cell survival and signaling in response to nutrient deprivation and chemotherapy; and (3) To delineate the molecular functions of Fnip1 in autophagy and mTOR signaling pathways. We will use biochemical and genetic approaches to determine whether Fnip1 is essential to "turn off" mTOR mediated nutrient consumption, and "turn on" autophagy (self-digestion of organelles to generate nutrients) in response to nutrient deficit. These studies will address our overall innovative hypothesis that inhibition of Fnip1 "disconnects" the essential link between anabolic cell growth and aerobic glycolysis, by permitting activated lymphocytes and/or tumor cells to grow in the absence of sufficient energy and bio- substrates, resulting in "nutrient exhaustion" and cell death.