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

描述（由申请人提供）：相对于静息淋巴细胞，活化的淋巴细胞和癌细胞都表现出细胞代谢从氧化磷酸化（有效产生能量）到有氧糖酵解（产生燃料电池分裂所需的生物前体（如脂质、氨基酸和核苷酸））的独特转变。对控制这种代谢转换（称为“瓦尔堡效应”）的因素的理解是非常重要的，因为它可能导致选择性阻断自身免疫性疾病中的淋巴细胞活化和/或抑制癌细胞存活的新策略。在本申请中，我们建议研究一种称为滤泡素相互作用蛋白-1（Fnip 1）的新型蛋白质，我们的研究表明，该蛋白质在能量和营养应激期间（如淋巴细胞活化、营养限制和癌基因活化期间）对维持“代谢平衡”至关重要。基于外周血中完全不存在B淋巴细胞，我们在化学诱变筛选中鉴定了一种缺乏Fnip 1的创新小鼠新品系。Fnip 1基因敲除小鼠在前B细胞和不变的自然杀伤T（iNKT）细胞发育阶段出现阻滞，在这些阶段，细胞通常依赖于c-Myc进行大规模分裂，c-Myc是一种在人类许多癌症中失调的癌基因。值得注意的是，在伯基特B细胞淋巴瘤的小鼠模型中，Fnip 1的缺失也保护免受c-Myc诱导的前B细胞淋巴瘤。虽然Fnip 1的功能尚不清楚，但它与卵泡素（一种功能未知的蛋白质）和主代谢调节因子AMP激酶相互作用，AMP激酶是一种能量传感分子，可刺激能量产生以响应能量应激，并抑制由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Fnip 1如何控制淋巴细胞的发育、活化、代谢和转化。本研究的具体目的是：（1）明确Fnip 1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Fnip 1的缺失是否抑制“瓦尔堡效应”;（2）确定Fnip 1在B细胞淋巴瘤存活和对代谢应激和化疗药物敏感性中的重要性。我们将有条件地删除小鼠原发性B细胞淋巴瘤中的Fnip 1，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Fnip 1在自噬和mTOR信号传导途径中的分子功能。我们将使用生物化学和遗传学方法来确定Fnip 1是否对“关闭”mTOR介导的营养消耗至关重要，并“打开”自噬（细胞器的自我消化以产生营养物质）以应对营养缺乏。这些研究将解决我们的总体创新假设，即Fnip 1的抑制通过允许活化的淋巴细胞和/或肿瘤细胞在缺乏足够的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重要联系，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.