Lipid metabolism regulates cancer cell adaptation to hypoxic tumor stress

项目摘要 正常哺乳动物细胞中的合成代谢活性受到生长因子信号传导的严格调节， 对血液中营养素和氧气（O2）水平的变化高度敏感。与此相反， 癌基因激活和肿瘤抑制基因丢失扰乱代谢调节， 癌细胞的生物合成活性。我们最近发表的数据表明，这种失调的亲- 生长表型证明在模拟灌注不良、缺氧的条件下是不支持的。 实体瘤的亚结构域特征。具体地说，当具有组成性mTORC 1信号传导的细胞 暴露于血清和O2限制条件下，它们表现出高度膨胀的ER，表达 ER应激的标志物，并经历未折叠蛋白反应（UPR）诱导的细胞凋亡 效应子IRE 1 α和TXNIP。抑制蛋白质合成或mTORC 1信号转导逆转了这些变化。 表型，这表明蛋白质负荷升高导致ER应激并激活这些表型中的UPR。 细胞令人惊讶的是，添加外源性不饱和脂肪酸也挽救了细胞死亡， 这表明不饱和脂质的不足使得缺氧的、缺乏血清的癌细胞 不能扩张内质网膜以适应蛋白质合成的增加。重要的是 广泛的人类癌细胞系显示出对外源性不饱和脂肪酸的类似依赖性。 血清和氧剥夺条件下的生存脂质。总的来说，这些数据表明， 新的和一般的代谢脆弱性的癌细胞，可以利用治疗。的 本提案的具体目的是（1）确定IRE 1 α和 TXNIP在肿瘤样应激条件下促进癌细胞死亡;（2）探讨脂质体的作用 储存促进癌细胞存活。简单地说，具体目标1将通过以下方式研究机制： IRE 1 α对TXNIP表达的调控，以及ROS在TXNIP介导的细胞凋亡中的作用。 具体目标2将阐明肾透明细胞中脂滴（LD）形成的细胞保护作用 癌（ccRCC）细胞。初步数据表明，ADRP蛋白，其表达是 由缺氧诱导因子-2 α调节，是LD形成所必需的。敲低HIF-2α或 ADRP表达激活IRE 1 α并诱导TXNIP表达，促进ccRCC中的凋亡。 多种实验方法将被用来解剖肿瘤之间的因果关系， 缺氧、HIF-2α表达、ADRP功能、LD形成、从头脂肪生成和谷氨酰胺 体外和体内代谢。由于在小鼠中观察到新生脂肪生成和LD形成增加， 多种癌细胞类型，这些结果应该有一般意义的理解的作用， 恶性肿瘤的脂质转运和代谢。

Project summary Anabolic metabolic activity in normal mammalian cells is tightly regulated by growth factor signaling that is highly responsive to changing levels of blood-borne nutrients and oxygen (O2). In contrast, oncogene activation and tumor suppressor loss perturbs metabolic regulation to drive constitutive biosynthetic activity in cancer cells. Our recently published data indicate that this dysregulated pro- growth phenotype proves unsupportable under conditions that mimic the poorly perfused, hypoxic subdomains characteristic of solid tumors. Specifically, when cells with constitutive mTORC1 signaling were exposed to conditions of serum and O2 limitation, they exhibited highly distended ER, expressed markers of ER stress, and underwent apoptosis induced by the unfolded protein response (UPR) effectors IRE1α and TXNIP. Inhibition of either protein synthesis or mTORC1 signaling reversed these phenotypes, suggesting that elevated protein load caused ER stress and activated the UPR in these cells. Surprisingly, the addition of exogenous unsaturated fatty acids also rescued cell death, suggesting that an insufficiency of unsaturated lipids renders hypoxic, serum-deprived cancer cells incapable of expanding ER membranes to accommodate elevated protein synthesis. Importantly, a broad array of human cancer cell lines displayed a similar dependence on exogenous unsaturated lipids for survival under conditions of serum and O2 deprivation. Collectively, these data suggest a novel and general metabolic vulnerability of cancer cells that might be exploited therapeutically. The Specific Aims of this proposal are to (1) determine the molecular mechanisms whereby IRE1α and TXNIP promote cancer cell death under tumor-like stress conditions, and (2) to explore the role of lipid storage in promoting cancer cell survival. Briefly, Specific Aim 1 will investigate the mechanisms by which TXNIP expression is regulated by IRE1α, and the role of ROS in TXNIP-mediated apoptosis. Specific Aim 2 will address the cytoprotective role of lipid droplet (LD) formation in clear cell renal cell carcinoma (ccRCC) cells. Preliminary data indicate that the ADRP protein, the expression of which is regulated by Hypoxia Inducible Factor-2α, is essential for LD formation. Knocking down HIF-2α or ADRP expression activates IRE1α and induces TXNIP expression, and promotes apoptosis in ccRCC. Multiple experimental approaches will be used to dissect the causative relationship between tumor hypoxia, HIF-2α expression, ADRP function, LD formation, de novo lipogenesis and glutamine metabolism in vitro and in vivo. As elevated de novo lipogenesis and LD formation is observed in multiple cancer cell types, these results should have general implications for understanding the role of lipid transport and metabolism in malignancy.