Metabolic Barriers to T Cell Activation in Clear Cell Renal Cell Carcinoma

摘要 多年来，利用免疫系统来消灭癌细胞一直是一个目标，但它已经成为 显然，肿瘤会主动抑制免疫细胞功能。同时抑制免疫调节作用 受体，如通过PD-1检查点阻断治疗，具有巨大的前景，这种治疗是 仅对部分患者有效。决定对肿瘤免疫应答的因素仍然存在 很大程度上是不确定的。然而，我们的数据表明，T细胞的代谢需求可能是导致 免疫治疗的成功。我们已经证明，效应T细胞(T细胞)的激活需要高水平的葡萄糖 而合成代谢方面，癌细胞和肿瘤微环境可以抑制TEF代谢途径。 这可能代表了肿瘤免疫抑制的一个基本机制。为了更好地理解 肿瘤微环境对T细胞代谢的影响及免疫治疗的改进 手术切除的人肾透明细胞癌肿瘤浸润性淋巴细胞(TIL)的检测 (CcRCC)肿瘤样本，一种对PD-1阻断有反应并具有预后免疫标志的癌症。 CcRCC与冯·希佩尔·林道(Von Hippel Lindau，VHL)抑癌基因的突变和缺失高度相关，其中 导致HIF1α和HIF2α的稳定，并诱导转录的假缺氧反应 改变肿瘤以促进免疫抑制微环境，从而对ccRCC CD8产生负面影响 TIL功能与抗肿瘤免疫。我们发现CD8TIL在ccRCC中含量丰富，但这些细胞是均匀分布的。 PD-1水平升高，功能抑制。此外，CD8TIL有多种代谢损伤， 不能有效地摄取葡萄糖或进行糖酵解，并有小的，碎裂的线粒体 产生高水平的活性氧(ROS)。重要的是，中和ROS或提供 糖酵解终产物丙酮酸可部分恢复ccRCC CD8 TIL功能。谷氨酰胺也是一个关键 通过谷氨酰胺分解来支持T细胞线粒体代谢的营养物质，我们在这里报道 抑制或基因缺失该途径中的第一种酶，谷氨酰胺酶1(GLS1)，会导致 糖酵解的代偿性增加，可增强细胞毒性CD8功能。这项提案将考验 假设ccRCC微环境损害糖酵解并导致功能障碍的积聚 CD8TIL中线粒体和TIL糖酵解的解救将增强T细胞对免疫治疗的应答。我们 将研究原发ccRCC肿瘤和小鼠RCC模型：(1)确定线粒体是如何 CcRCC CD8 TIL的激活和代谢紊乱；(2)研究是否促进血糖 摄取或抑制GLS1促进糖代谢可改善CD8的代谢和功能 以及(3)检测PD-1阻断治疗对慢性肾细胞癌T细胞代谢和功能群体的影响。 总之，这些研究将建立ccRCC TIL代谢功能障碍的机制，并测试是否 增强T细胞糖酵解的方法可以改善癌症的免疫治疗。

SUMMARY Exploiting the immune system to eliminate cancer cells has been a goal for many years, but it has become apparent that tumors actively suppress immune cell functions. While inhibition of immunomodulatory receptors, such as through PD-1 checkpoint blockade therapy, holds tremendous promise, this treatment is effective in only a portion of patients. Factors that determine immune responsiveness against tumors remain largely uncertain. Our data show, however, that the metabolic demands of T cells may be a critical factor in the success of immunotherapy. We have shown that effector T cell (Teff) activation requires high rates of glucose and anabolic metabolism yet cancer cells and the tumor microenvironment can inhibit Teff metabolic pathways. This may represent a fundamental mechanism of tumor-mediated immune suppression. To better understand the influence of the tumor microenvironment on T cell metabolism and improve immunotherapies, we have examined tumor infiltrating lymphocytes (TIL) from surgically-excised human clear cell Renal Cell Carcinoma (ccRCC) tumor samples, a cancer responsive to PD-1 blockade and with a prognostic immune signature. ccRCC is highly associated with mutations and loss of the Von Hippel Lindau (VHL) tumor suppressor, which leads to stabilization of HIF1α and HIF2α and induction of a transcriptional pseudo-hypoxic response that alters the tumor to promote an immune suppressive microenvironment that can negatively impact ccRCC CD8 TIL function and anti-tumor immunity. We found CD8 TIL are abundant in ccRCC, yet these cells are uniformly PD-1high and functionally suppressed. In addition, CD8 TIL had multiple metabolic impairments and were unable to efficiently uptake glucose or perform glycolysis and had small, fragmented mitochondria that produced high levels of Reactive Oxygen Species (ROS). Importantly, neutralization of ROS or provision of the glycolytic end-product pyruvate could partially rescue ccRCC CD8 TIL function. Glutamine is also a key nutrient to support mitochondrial metabolism for T cells through glutaminolysis and we report here that inhibition or genetic deletion of the first enzyme in this pathway, Glutaminase 1 (GLS1), leads to a compensatory increase in glycolysis that can enhance cytotoxic CD8 function. This proposal will test the hypothesis that the ccRCC microenvironment impairs glycolysis and leads to accumulation of dysfunctional mitochondria in CD8 TIL and that rescue of TIL glycolysis will enhance T cell response to immunotherapy. We will study primary ccRCC tumors and mouse RCC models to: (1) Determine how mitochondria are dysregulated and impair activation and metabolism of ccRCC CD8 TIL; (2) Investigate if promoting glucose uptake or inhibiting GLS1 to enhance glucose metabolism can improve the metabolism and function of CD8 TIL; and (3) Test how PD-1 blockade therapy impacts T cell metabolism and functional populations in ccRCC. Together, these studies will establish the mechanism of metabolic dysfunction in ccRCC TIL and test if approaches to enhance T cell glycolysis can improve cancer immunotherapy.