Glycolysis and Glutaminase in CD8 T cell differentiation and anti-tumor immunity

摘要 免疫疗法改变了癌症治疗，改善了临床结果，但它并不能治愈大多数 病人。抗PD1单抗和嵌合抗原受体(CAR)T细胞等治疗 通过增强产生癌症特异性干扰素γ的Th1CD4T细胞的活性而发挥作用 细胞毒性CD8T细胞(CTL)。然而，增强的效应器功能必须与抗病毒能力相平衡。 癌症T细胞能够长期存活。免疫治疗的一个关键目标是增强效应器功能，同时 维持T细胞的寿命和记忆力。现在从Rathmell实验室的工作中可以清楚地看到，效应器T细胞 利用高比率的糖酵解，而记忆细胞则利用线粒体途径。在这里，我建议测试细胞 代谢作为增强免疫治疗中效应T细胞和记忆T细胞的手段。T细胞 从根本上改变激活后的代谢，增加糖酵解和谷氨酰胺氧化 (谷氨酰胺分解)以支持分化、效应器功能和最终产生长期记忆 依靠线粒体。调节T细胞谷氨酰胺代谢可能增强其疗效 通过增强T细胞效应器功能或记忆能力的免疫疗法。Rathmell实验室已经证明 代谢程序有氧糖酵解对于炎症和炎症中的效应T细胞(TEF)的功能是必不可少的 在肿瘤中。谷氨酰胺分解补充糖酵解通过将谷氨酰胺转化为三羧酸来为T细胞提供燃料 酸循环中间体α-酮戊二酸(AKG)。使用条件基因敲除谷氨酰胺分解酶 谷氨酰胺酶(GLS)，将谷氨酰胺转化为谷氨酸，以及目前正在临床使用的GLS抑制剂 作为抗癌药物的试验，我们发现抑制GLS导致代偿性增加 糖酵解，增强Th1和CTL的Tef功能和分化。除了增加效应器之外 然而，我发现抑制GLS功能也会增加抑制性受体的表达，并慢性 GLS缺乏最终会抑制T细胞。相反，短暂的GLS抑制增强了TEF功能，而 也启动了线粒体代谢的记忆样分化，并导致了T细胞持久性的改善 在活体内。在这项提议中，我将检验这样一个假设，即短暂的GLS抑制可以增强TEFF功能和 维持T细胞存活和记忆以提高抗癌免疫治疗效果，而慢性GLS 抑制会导致代偿性糖酵解、终末分化和衰竭。我将：(1)测试如何 短暂和慢性GLS抑制通过确定记忆T细胞形成的差异来影响CTL的命运， 评估代偿性糖酵解对Tef表型的贡献，并建立线粒体 GLS抑制的后果；和(2)检测GLS抑制对小鼠免疫治疗效果的影响 CD19靶向CAR T细胞和抗PD1治疗。这些研究将展示一种新的方法来 改进抗癌免疫治疗，突出利用GLS抑制调节T细胞的策略 新陈代谢和分化。

Summary Immunotherapy has transformed cancer treatment and improved clinical outcomes, but it does not cure most patients. Treatments such as anti-PD1 monoclonal antibodies and chimeric antigen receptor (CAR) T cells function by boosting the activity of cancer-specific interferon gamma-producing Th1 CD4 T cells and cytotoxic CD8 T cells (CTL). Increased effector function, however, must be balanced with the ability of anti- cancer T cells to persist long-term. A key goal of immunotherapy is to enhance effector function while maintaining T cell longevity and memory. It is now clear from work in the Rathmell lab that effector T cells utilize high rates of glycolysis while memory cells utilize mitochondrial pathways. Here I propose to test cell metabolism as a means to enhance both effector and memory T cell populations in immunotherapy. T cells radically alter their metabolism upon activation and increase glycolysis and glutamine oxidation (glutaminolysis) to support differentiation, effector function, and eventual generation of long-term memory that depend on mitochondria. Modulation of T cell glutamine metabolism may augment the efficacy of immunotherapy by enhancing both T cell effector function or memory capacity. The Rathmell Lab has shown that the metabolic program aerobic glycolysis is essential for effector T cell (Teff) function in inflammation and in tumors. Glutaminolysis complements glycolysis to fuel T cells by converting glutamine to the tricarboxylic acid cycle intermediate alpha-ketoglutarate (aKG). Using a conditional knockout of the glutaminolysis enzyme Glutaminase (GLS), which converts glutamine to glutamate, and an inhibitor of GLS that is currently in clinical trials as an anti-cancer agent, we have found that inhibition of GLS leads to a compensatory increase in glycolysis that enhances Th1 and CTL Teff function and differentiation. In addition to increasing effector function, however, I found that GLS inhibition also increases expression of inhibitory receptors, and chronic GLS deficiency ultimately suppresses T cells. In contrast, transient GLS inhibition enhanced Teff function while also priming mitochondrial metabolism for a memory-like differentiation, and led to improved T cell persistence in vivo. In this proposal, I will test the hypothesis that transient GLS inhibition can augment Teff function and maintain T cell survival and memory to boost anti-cancer immunotherapy efficacy, whereas chronic GLS inhibition will drive compensatory glycolysis, terminal Teff differentiation, and exhaustion. I will: (1) Test how transient versus chronic GLS inhibition affects CTL fate by determining differences in memory T cell formation, assessing the contribution of compensatory glycolysis to Teff phenotypes, and establishing the mitochondrial consequences of GLS inhibition; and (2) Test the effect of GLS inhibition on the immunotherapy efficacy of CD19-targeted CAR T cells and anti-PD1 treatment. These studies will demonstrate a new approach to improve anti-cancer immunotherapy and highlight a strategy of using GLS inhibition to modulate T cell metabolism and differentiation.