Uncovering the metabolic underpinnings of T cell exhaustion

项目摘要/摘要 免疫疗法的成功，如阻断共抑制的“检查点”分子，已经改变了 癌症的治疗模式。然而，强大的反应仅限于一部分患者的事实 强调了进一步了解耗尽的T细胞生物学的必要性：是什么驱动了它们的分化， 维持它们的功能障碍，以及如何恢复它们的活力以根除肿瘤细胞。我们的实验室和其他人 已经发现代谢应激和线粒体功能障碍是T细胞衰竭的关键驱动因素，两者 从细胞外在和细胞内在的角度来看。我们最近报道了线粒体应激和反应 氧物种(ROS)的产生，在低氧环境下被驱动到无法忍受的水平 持久的抗原，足以使细胞偏离到最终耗尽的命运。两种抗氧化剂 药物和遗传可能会使T细胞分化从疲惫转向更有功能的命运。但ROS的产生究竟如何改变T细胞的命运和功能仍不清楚。最耐人寻味的 观察到ROS是通过线粒体功能障碍改变T细胞信号而升高的：因为过氧化氢是 更有效的酪氨酸磷酸酶抑制剂，仅提高ROS就能模拟TCR和其他 磷酸酪氨酸信号。ROS还戏剧性地重新编程细胞新陈代谢：通过抑制乌头酸酶，柠檬酸盐从线粒体中被驱动，在那里它被转化为乙酰辅酶A，作为从头开始的底物 脂肪生成作用。结果，虽然耗尽的细胞拥有功能失调的线粒体，但竞争能力很弱 葡萄糖，它们富含脂滴，抑制脂肪酸氧化和脂肪分解。虽然我们知道 线粒体应激可使T细胞耗尽，而最终耗尽的T细胞在代谢过程中 不充分，最终驱动和加强表型的机制仍不清楚。在这份提案中， 我们将确定T细胞衰竭的代谢基础：代谢应激如何干扰 信号、转录和分化。目标1：确定氧化应激如何改变T细胞信号 在磷酸酶抑制水平上级联反应。ROS作为酪氨酸抑制物在信号转导中发挥核心作用 磷酸酶。我们将在体内确定ROS在耗尽T细胞功能中的作用，并使用蛋白质组学和 转录转录技术，以确定对ROS诱导敏感的磷酸化级联反应。目标2： 确定ROS介导的代谢流量变化如何破坏T细胞功能。在这个目标中，我们将探索 增加脂肪储存在T细胞功能中的作用，并询问这些升高的脂质水平是否 代表“自重”或未开发的燃料来源。目的3：明确氧化应激反应引起的营养途径改变的重要性。我们的数据表明，SLC16A11类似地支持精疲力竭的T细胞摄取乳酸，并维持其功能失调状态。使用条件基因敲除小鼠和阻断抗体，我们将确定单羧酸代谢在疲惫的T细胞生物学中的重要性。

PROJECT SUMMARY/ABSTRACT The successes of immunotherapies like blockade of co-inhibitory `checkpoint' molecules have changed the treatment paradigm of cancer. However, the fact that robust responses are restricted to a subset of patients highlights the need to further understand the biology of exhausted T cells: what drives their differentiation, maintains their dysfunction, and how they may be reinvigorated to eradicate tumor cells. Our lab and others have revealed that metabolic stress and mitochondrial dysfunction are key drivers in T cell exhaustion, both from a cell extrinsic and cell intrinsic perspective. We recently reported that mitochondrial stress and reactive oxygen species (ROS) production, driven to intolerable levels under hypoxic environments in the face of persistent antigen, was sufficient to deviate cells into a terminally exhausted fate. Antioxidants both pharmacologic and genetic could bias T cell differentiation away from exhaustion to more functional fates. But precisely how ROS production alters T cell fate and function remains unclear. One of the more intriguing observations was elevating ROS via mitochondrial dysfunction altered T cell signaling: as peroxide is one of the more potent inhibitors of tyrosine phosphatases, elevating ROS alone mimicked TCR and other phosphotyrosine signals. ROS also dramatically reprograms cellular metabolism: by inhibiting aconitase, citrate is driven from the mitochondria where it is converted to acetyl-CoA, acting as a substrate for de novo lipogenesis. As a result, while exhausted cells possess dysfunctional mitochondria and compete poorly for glucose, they are loaded with lipid droplets and repress fatty acid oxidiation and lipolysis. While we know that mitochondrial stress can drive T cells to exhaustion and that terminally exhausted T cells are metabolically insufficient, the mechanisms that ultimately drive and enforce the phenotype remain unclear. In this Proposal, we will identify the metabolic underpinnings of T cell exhaustion: how metabolic stress can interfere with signaling, transcription, and differentiation. AIM 1: Determine how oxidative stress alters T cell signaling cascades at the level of phosphatase inhibition. ROS play central roles in signaling as inhibitors of tyrosine phosphatases. We will determine the role of ROS in exhausted T cell function in vivo, and use proteomics and transcriptomic technologies to identify the phosphorylation cascades susceptible to ROS induction. AIM 2: Identify how ROS-mediated changes in metabolic flux undermine T cell function. In this Aim, we will explore the role increased lipid storage plays in T cell function and ask whether these elevated levels of lipids represent `dead weight' or an untapped fuel source. AIM 3: Define the importance of altered nutrient pathways induced through oxidative stress responses. Our data suggest Slc16a11 similarly supports lactate uptake into exhausted T cells and maintains their dysfunctional state. Using a conditional knockout mouse and blocking antibodies, we will determine the importance of monocarboxylate metabolism in exhausted T cell biology.