Modifying T Cell Responses by Combinatorial Targeting of Negative regulators

描述(由申请人提供)：T细胞在长期受到持久抗原的刺激后，通常会失去继续作出强健反应的能力。这是一个重大问题，因为既有肿瘤和慢性感染也可以通过诱导抗原特异性T细胞处于类似的耐受状态--即耗竭或无能--来逃避保护性免疫反应。该项目确定了有助于维持这种耐受性的新型调节分子。耐受的分子机制被理解为包括抑制分子的表达，如PD1、LAG3、CTLA4等，它们抑制T细胞内不同的激活信号。因此，最近的临床研究已经成功地使用抗体和抑制剂来对抗这种负面调节因子，以增强T细胞对肿瘤和慢性病毒感染的反应。这项建议评估了两个新发现的调节因子-PEAR1和EndoD1-协同抑制CD4T细胞的反应性，促进T细胞耐受的假说。据预测，抑制T细胞扫描中的这些分子可以提高免疫治疗的效果，例如对抗肿瘤--即使在仅针对已知调节因子是不够的情况下也是如此。这些假设将使用三个独立和互补的目标进行评估。1.通过使用T细胞特异性基因敲除方法，确定在自身免疫和移植排斥的背景下，这些新的调节剂如何单独和协同地影响体内T细胞的激活。除了关节炎模型的初步数据外，还将通过计数激活标记物的表达、细胞因子的产生以及免疫病理学来评估基因敲除对皮炎模型中T细胞的影响。2.阐明这些调节剂在耐受性T细胞中的生化靶点，以期开发一种合理的策略来确定可以最有效地靶向免疫治疗的途径的排列。为此，将使用分子分析和计算模型的组合。3.确定在肿瘤过继T细胞治疗过程中通过下调肿瘤特异性转基因T细胞和天然肿瘤浸润性淋巴细胞中这些调节因子的表达来针对这些调节因子的策略。预计这些实验将首次确定T细胞表达的PEAR1和EndoD1作为T细胞效应反应的抑制物所起的作用。这一点意义重大，因为它为治疗提供了新的靶点：(A)在对肿瘤和慢性感染的免疫背景下，增强抗原特异性T细胞的功能； (B)在自身免疫和移植物排斥反应期间耐受T细胞。此外，对这些分子与其他已知的T细胞激活负调控因子，如PD1、CTLA4、CBL-b和LAG3的协同性的拟议研究，将产生快速将这些靶点与正在进行的临床工作相结合所需的数据。因此，提案中还包括一种合理的战略，以指导此类组合治疗的设计，使用生化数据和计算模型。

DESCRIPTION (provided by applicant): T cells often lose their ability to continue making robust responses, when stimulated for a long time by a persistent antigen. This is a significant problem, because established tumors and chronic infections can also evade protective immune responses by inducing a similar state of tolerance in antigen-specific T cells - known as exhaustion or anergy. This project identifies novel regulatory molecules that help maintain such a tolerance. The molecular mechanics of tolerance is understood to include the expression of inhibitory molecules such as PD1, LAG3, CTLA4 etc. which dampen different intracellular activating signals in T cells. Accordingly, recent clinical studies have successfully used antibodies and inhibitors against such negative regulators to augment T cell responses to tumors and chronic viral infections. This proposal evaluates the hypothesis that two newly identified regulators - PEAR1 and EndoD1 - cooperatively inhibit the responsiveness of CD4+ T cells, contributing to T cell tolerance. It is projected that inhibiting these molecules in T cellscan improve the efficacy of immunotherapy, e.g. against tumors - potentially even in contexts where targeting known regulators alone is insufficient. These hypotheses will be evaluated using three independent and complimentary aims. 1. Determine how these novel regulators individually and synergistically affect T cell activation in vivo in the context of autoimmunity and graft rejection by using a T cell-specific knockdown approach. In addition to preliminary data in an arthritis model, the impact of the knockdowns on T cells in a dermatitis model will be evaluated by enumerating activation marker expression, cytokine production as well as immunopathology. 2. Elucidate the biochemical targets of these regulators in tolerant T cells, with a view to developing a rational strategy to identify permutations of pathways that can be most efficiently targeted for immunotherapy. A combination of molecular analyses and computational modeling will be used for this purpose. 3. Define a strategy to target these regulators during adoptive T cell therapy for tumors by knocking-down their expression in tumor-specific transgenic T cells as well as native tumor-infiltrating lymphocytes. It is expected that these experiments will, for the first time, define the role of T cell expressed PEAR1 and EndoD1 as inhibitors of T cell effector responses. This is significant because it offers new targets for therapeutically (a) enhancing the function of antigen-specific T cells in the context of immunity to tumors and chronic infections or (b) tolerising T cells during autoimmunity and graft rejection. Furthermore, the proposed studies on the cooperativity of these molecules with other known negative regulators of T cell activation, such as PD1, CTLA4, cbl-b and LAG3, will generate data required to rapidly integrate these targets with ongoing clinical efforts. A rational strategy to guide the design of such combinatoria treatments, using biochemical data and computational models, is therefore also included in the proposal.