KIR2DL2 Immune Checkpoint as Modulator of T-Cell Effector Function

项目摘要 杀伤细胞免疫球蛋白样受体(KIR)主要由NK细胞表达，尽管它们的 在CD_4、CD_8和γδT细胞中也有表达。在CD8 T细胞中，KIR的表达是 在淋巴细胞成熟的后期被诱导，被认为调节特定的T细胞效应器功能。 在KIR受体中，KIR2DL2调节T细胞效应器的功能，因为KIR2DL2存在CD8 T细胞 T细胞受体后活化诱导的细胞死亡水平降低和干扰素-γ分泌减少 刺激。CD8T细胞中KIR2DL2表达的抑制功能的概念得到了 观察到，表达其同源配体人类白细胞抗原-C1的患者显示总存活率降低和 无法控制肿瘤的生长。我们的初步数据显示，KIR2DL2在体内的表达增加 在过继免疫治疗的患者和临床前模型中过继转移T细胞。使用 胰腺肿瘤模型中，我们发现表达KIR2DL2的嵌合抗原受体(CAR)-T细胞 在存在KIR2DL2‘S配体的情况下，细胞毒性明显低于KIR2DL2-。此外， KIR2DL2在CAR-T细胞中的表达与抗肿瘤疗效降低相关，这是一种HLA-I依赖的研究 在胰腺癌的小鼠模型中。根据这些初步发现，我们假设 KIR2DL2作为T细胞免疫检查点，调节T细胞效应器功能，导致 无效的免疫监控。因此，在T细胞制造期间靶向KIR2DL2可能 改善细胞输注后的T细胞功能。我们将通过1)定义调制来检验我们的假设 KIR2DL2影响CAR和TCR转基因T细胞抗肿瘤效应的机制我们 将决定KIR2DL2参与TCR转基因和CAR-T细胞效应器功能的整体效果。 此外，我们将确定KIR2DL2中的哪些区域负责其调节功能。 最后，我们将描述KIR2DL2信号相互作用组和触发的下游事件 通过免疫沉淀和蛋白质组学分析其与配体的相互作用。2)提高T细胞的性能 通过取消KIR2DL2功能来加强过继细胞免疫疗法(ACTs)。以防止其 抑制效应、KIR2DL2表达和/或信号的操纵将被进行并适应于 目前用于CAR-T细胞制造的方案。根据预期结果，我们将首次联系 KIR2DL2在治疗性T细胞中的生物学和分子功能拟议的研究将 增加我们对KIR2DL2生物学的机械理解，并将产生具有高性能的新型细胞产品 翻译潜力。

Project Summary Killer cell immunoglobulin-like receptors (KIR) are mainly expressed by NK cells, although their expression has also been described in CD4+, CD8+ and γδ T cells. Within CD8 T cells, KIR expression is induced at later stages of lymphocyte maturation and is thought to regulate specific T cell effector functions. Within KIR receptors, KIR2DL2 modulates T cell effector functions, as KIR2DL2+ CD8+ T cells present reduced level of activation-induced cell death, and poor IFN-γ secretion after T cell receptor (TCR) stimulation. The notion of a suppressive function of KIR2DL2 expression in CD8+ T cells is supported by the observation that patients that express its cognate ligand, HLA-C1, showed decreased overall survival and could not control tumor growth. Our preliminary data show that KIR2DL2 expression increases in vivo in adoptively transferred T cells in patients and in preclinical models of adoptive immunotherapy. Using a pancreatic tumor model, we found that chimeric antigen receptor (CAR)-T cells expressing KIR2DL2 were significantly less cytotoxic than their KIR2DL2- counterparts in presence of KIR2DL2’s ligand. Furthermore, KIR2DL2 expression in CAR-T cells was associated with reduced antitumor efficacy, in an HLA-I-dependent manner, in a murine model of pancreatic cancer. Based on these preliminary findings we hypothesize that KIR2DL2 behaves as a T cell immune checkpoint, modulating T cell effector function and leading to an ineffective immunosurveillance. Therefore, targeting KIR2DL2 during T cell manufacturing may improve T cell performance after cell infusion. We will test our hypothesis by 1) Defining the modulatory mechanisms whereby KIR2DL2 shapes CAR- and TCR-transgenic T cell antitumoral effector function. We will determine the overall effect of KIR2DL2 engagement in TCR-transgenic and CAR-T cell effector function. Additionally, we will determine which regions within KIR2DL2 are responsible for its modulatory function. Finally, we will characterize both the KIR2DL2 signaling interactome and the downstream events triggered by its ligand interaction by immunoprecipitation and proteomic analyses. 2) Improving T cell performance for the enhancement of adoptive cell immunotherapies (ACTs) by abrogating KIR2DL2 function. To prevent its inhibitory effect, manipulation of KIR2DL2 expression and/or signaling will be conducted and adapted to the current protocols for CAR-T cell manufacturing. Based on the anticipated results, we will link for the first time the biological and molecular function of KIR2DL2 within therapeutic T cells. The proposed studies will increase our mechanistic understanding of KIR2DL2 biology and will generate novel cell products with high translational potential.