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

总结 利用免疫系统消除癌细胞多年来一直是一个目标，但它已成为 很明显，肿瘤会主动抑制免疫细胞的功能。虽然免疫调节抑制 受体，如通过PD-1检查点阻断疗法，具有巨大的前景，这种治疗是 仅对部分患者有效。决定对肿瘤免疫反应的因素仍然存在 很大程度上不确定。然而，我们的数据表明，T细胞的代谢需求可能是一个关键因素， 免疫治疗的成功我们已经表明，效应T细胞（Teff）的激活需要高速率的葡萄糖 和合成代谢，但癌细胞和肿瘤微环境可以抑制Teff代谢途径。 这可能代表了肿瘤介导的免疫抑制的基本机制。更好地了解 肿瘤微环境对T细胞代谢的影响并改善免疫疗法，我们已经 检查了来自肾透明细胞肾细胞癌的肿瘤浸润淋巴细胞（TIL 在一个实施方案中，本发明涉及一种用于在ccRCC肿瘤样品中检测癌症的方法，所述癌症是对PD-1阻断有响应并且具有预后免疫特征的癌症。 ccRCC与Von Hippel Lindau（VHL）肿瘤抑制因子的突变和缺失高度相关， 导致HIF 1 α和HIF 2 α的稳定，并诱导转录假缺氧反应， 改变肿瘤，促进免疫抑制微环境，对ccRCC CD 8产生负面影响 TIL功能与抗肿瘤免疫。我们发现ccRCC中CD 8 TIL丰富，但这些细胞均匀分布， PD-1高和功能抑制。此外，CD 8 TIL具有多种代谢损伤， 不能有效地摄取葡萄糖或进行糖酵解，并且具有小的、碎片化的线粒体， 产生高水平的活性氧（ROS）。重要的是，ROS的中和或提供 糖酵解终产物丙酮酸可部分拯救ccRCC CD 8 TIL功能。谷氨酰胺也是一个关键 营养素，以支持线粒体代谢的T细胞通过γ-氨基，我们在这里报告说， 该途径中的第一种酶谷氨酰胺酶1（GLS 1）的抑制或遗传缺失导致 糖酵解的代偿性增加可以增强细胞毒性CD 8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性糖酵解的积累， CD 8 TIL中线粒体和TIL糖酵解的拯救将增强T细胞对免疫疗法的应答。我们 将研究原发性ccRCC肿瘤和小鼠RCC模型，以：（1）确定线粒体是如何被激活的。 （2）研究是否促进葡萄糖代谢， 摄取或抑制GLS 1以增强糖代谢可改善CD 8的代谢和功能 TIL;和（3）测试PD-1阻断疗法如何影响ccRCC中的T细胞代谢和功能群体。 总之，这些研究将建立ccRCC TIL中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.