Profiling and leveraging bystander T cells within the tumor microenvironment

项目概要/摘要 癌症仍然是美国死亡率的主要原因，每4例死亡中就有1例是由癌症引起的。免疫疗法 开创了癌症治疗的新时代，利用免疫系统的效力来限制癌症， 传统疗法的许多副作用这些免疫疗法主要针对免疫 肿瘤抗原特异性反应（tAg）。尽管tAg特异性T细胞免疫疗法已被证明是有效的， 尽管这些药物是对抗癌症的工具，但它们的成功取决于环境，在大多数实体瘤中显示出有限的功效。 这在很大程度上是由于慢性T细胞受体（TCR）与同源tAg结合，导致永久性细胞凋亡。 功能障碍实体瘤包括大多数癌症病例和死亡，因此，开发其他治疗方法是必要的。 免疫疗法中的细胞模式。最近，tAg-非特异性"旁观者" T 在实体瘤中观察到tAg细胞，并且其数量经常超过tAg特异性T细胞。尽管它们在肿瘤中的功能 目前还不清楚，旁观者T细胞一旦被炎症激活，可以在许多情况下发挥细胞毒性效应功能。 的context。我独特的方法利用T细胞与定义的T细胞受体（TCR），使我能够确定 决定旁观者T细胞进入肿瘤的机制，以及它们是否保持对肿瘤的反应能力。 刺激一旦肿瘤驻留。为了了解肿瘤微环境的异质性，我将使用 多种实体瘤动物模型。我的目标有两个：第一，我想测试旁观者T细胞 如果它们由于不能识别tAg而免于失调，则它们可以迁移到肿瘤。二我 想要测试旁观者T细胞的二分效应是否可以在治疗上被利用来改善抗- 肿瘤反应。在体内平衡时，旁观者T细胞可以简单地拒绝其他免疫细胞接近靶标， 阻碍抗原（Ag）特异性免疫应答。一旦被炎症激活，旁观者T细胞迅速 获得效应子功能并以类似先天的方式直接杀死靶细胞。为了实现这些目标，我将 利用我在28色流式细胞术方面的专业知识来询问细胞表型，激活和功能能力。 我将用三维免疫荧光补充我的流式细胞术，这将揭示亚细胞。 肿瘤内免疫细胞的解剖结构。假设：我的中心假设是，旁观者T细胞 在实体瘤中保持功能性，并且可以特异性地在癌症中进行治疗。我来测试一下 目的1：检验旁观者T细胞被募集到 CXCR3在肿瘤中起作用，并在肿瘤微环境中保持功能。目的2：检验抗- 肿瘤免疫应答可以通过激活的旁观者T细胞或靶向清除组织驻留的T细胞来增强。 旁观者我提出的实验将阐明旁观者T细胞在肿瘤中的作用， 干预措施，目标是制定干预措施，以改善癌症的结果，这是符合 NCI和NIH的使命。

PROJECT SUMMARY/ABSTRACT Cancer remains a leading cause of mortality within the US, responsible for 1 in 4 deaths. Immunotherapies have ushered in a new age of cancer treatment, leveraging the potency of the immune system to restrict cancer without many of the side effects of conventional therapies. These immunotherapies have primarily targeted immune responses specific for tumor antigens (tAg). Although tAg-specific T cell immunotherapies have proven powerful tools in combatting cancer, their success is context-dependent, displaying limited efficacy in most solid tumors. This is largely owed to chronic T cell receptor (TCR) binding to cognate tAg, leading to permanent cellular dysfunction. Solid tumors comprise the majority of cancer cases and deaths, thus, it is essential to exploit other cellular modalities in immunotherapies. Recently, it has become apparent that tAg-nonspecific “bystander” T cells are observed and often outnumber tAg-specific T cells in solid tumors. Although their function within tumors is unknown, bystander T cells can exert cytotoxic effector function once activated by inflammation in a number of contexts. My unique approach leveraging T cells with defined T cell receptors (TCRs) allows me to determine the mechanisms that dictate bystander T cell entry into the tumor and if they maintain the ability to respond to stimulation once tumor-resident. To appreciate the heterogeneity of tumor microenvironments, I will employ multiple animal models of solid tumors. My objectives are two-fold: First, I want to test how bystander T cells migrate to the tumor and if they are spared from dysregulation due to their inability to recognize tAg. Second, I want to test if the dichotomous effects of bystander T cells can be therapeutically leveraged to improve anti- tumor responses. At homeostasis, bystander T cells can simply deny other immune cells access to targets, hindering antigen (Ag)-specific immune responses. Once activated by inflammation, bystander T cells rapidly acquire effector function and directly kill target cells in an innate-like manner. To achieve these objectives, I will employ my expertise in 28-color flow cytometry to interrogate cell phenotype, activation, and functional capacity. I will complement my use of flow cytometry with 3-dimensional immunofluorescence, which will uncover sub- anatomic immune cell organization within the tumor. Hypothesis: My central hypothesis is that bystander T cells in solid tumors remain functional and can be therapeutically leveraged in cancer specifically. I will test this hypothesis through two independent aims: AIM 1: Test the hypothesis that bystander T cells are recruited into tumors by CXCR3 and remain functional in the tumor microenvironment. AIM 2: Test the hypothesis that anti- tumor immune responses can be enhanced by activated bystander T cells or targeted depletion of tissue-resident bystanders. My proposed experiments will elucidate the role of bystander T cells in tumor both with and without interventions, with the goal of developing interventions to improve cancer outcomes, which is in alignment with the mission of the NCI and NIH.