Lipid metabolism regulates cancer cell adaptation to hypoxic tumor stress

项目总结 正常哺乳动物细胞中的合成代谢活动受到生长因子信号的严格调控 对血液中营养物质和氧气(O2)水平的变化高度敏感。相比之下， 癌基因激活和抑癌基因缺失扰乱代谢调节驱动结构性 癌细胞的生物合成活性。我们最近公布的数据表明，这种失调的前- 生长表型在模拟低血流、低氧的条件下被证明是不可支持的 实体瘤的亚域特征。具体地说，当具有组成性mTORC1信号的细胞 暴露于血清和氧气限制的条件下，它们表现出高度膨胀的ER，表达 内质网应激标志物，并经历了未折叠蛋白反应(UPR)诱导的细胞凋亡 效应器IRE1α和TXNIP。对蛋白质合成或mTORC1信号的抑制逆转了这些 表型，提示蛋白质负荷增加导致内质网应激并激活UPR 细胞。令人惊讶的是，外源不饱和脂肪酸的加入也挽救了细胞死亡， 提示不饱和脂类的不足会导致缺乏血清的低氧癌细胞。 不能扩张内质网膜以适应蛋白质合成的增加。重要的是，一个 大量的人类癌细胞对外源性不饱和脂肪酸也表现出类似的依赖关系 在血清和氧气缺乏条件下生存所需的脂类。总体而言，这些数据表明 癌细胞的新的和普遍的代谢脆弱性，可能被开发用于治疗。这个 这项建议的具体目的是：(1)确定IRE1α和IRE1的分子机制 TXNIP在肿瘤样应激条件下促进癌细胞死亡；(2)探讨脂质的作用 促进癌细胞存活的储存。简而言之，特定目标1将通过以下方式研究这些机制 哪些TXNIP的表达受IRE1α调控，以及ROS在TXNIP介导的细胞凋亡中的作用。 具体目标2将阐述在透明细胞肾细胞中形成脂滴(LD)的细胞保护作用 癌(CcRCC)细胞。初步数据表明，adrp蛋白的表达是 受低氧诱导因子-2α调控，对LD的形成是必不可少的。击倒HIF-2α或 ADRP表达可激活IRE1α，诱导TXNIP表达，促进肾细胞癌细胞凋亡。 将使用多种实验方法来剖析肿瘤之间的致病关系 低氧、缺氧诱导因子-2α表达、肾上腺皮质激素释放酶功能、LD形成、新生脂肪生成和谷氨酰胺 体内和体外代谢。随着新生脂肪生成和LD形成的增加，在 多种癌细胞类型，这些结果应该对理解其作用具有普遍意义 恶性肿瘤中的脂质转运和代谢。

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.