Investigating TOX2 as a novel regulator of T cell memory differentiation

项目摘要 最近的科学进步使免疫疗法成为许多癌症患者的一个有希望的选择。 免疫疗法可以增强免疫系统识别和摧毁癌细胞的能力。然而， 并不是所有的患者都对免疫治疗有反应，因此更好地了解了其潜在的机制。 免疫细胞效应功能是提高应答率所必需的。与卡尔·琼博士的团队合作， 我们的实验室最近研究了一名慢性淋巴细胞性白血病患者，他在接受CAR-T治疗后完全缓解。我们 发现绝大多数CAR+细胞的TET2基因发生了破坏，并进行了实验 TET2基因的敲除可增强CAR T细胞的体外增殖能力和抗肿瘤活性。 此外，具有TET2基因敲除的T细胞显示TOX2的表达增加，TOX2是TET2基因的成员之一。 HMG-box家族的DNA结合蛋白。TOX2是一种免疫特异性转录因子，表达于 人类的脾和扁桃体。有趣的是，TOX2唯一已知的功能是它正向调节 转录因子T-BET抑制PD-1在自然杀伤细胞发育中的作用 蛋白。由于TET2基因敲除的CAR T细胞中TOX2 mRNA水平的增加，除了其能力外， 为了下调一个关键的抑制性受体，我假设TOX2是T细胞效应器的正向调节因子 由于其上调T-BET的能力而发挥作用。为了验证这一假设，我将追求三个目标。目标1是 操控TOX2在人T细胞中的表达，以确定它是否促进T细胞分化和 效应器功能。目标2是确定TOX2与基因组的结合模式，目的是识别 它的转录靶点，在mRNA和蛋白质水平上。关于HMG-box如何 蛋白质在体内结合染色质。因此，这一目标也将提供对 HMG蛋白的染色质功能。目的3检测慢性淋巴细胞性白血病患者外周血中TOX2基因的表达水平。 接受CAR T治疗，以确定较高的TOX2水平是否预示着对CAR T的更好反应 心理治疗。本研究可能提高TOX2作为改善CAR T细胞功能的新靶点。一种理解 的分子机制可以激活TOX2，最终改善患者的反应 去接受免疫治疗。

PROJECT ABSTRACT Recent scientific advancements have made immunotherapy a promising option for many cancer patients. Immunotherapy can bolster the ability of the immune system to recognize and destroy cancer cells. However, not all patients respond to immunotherapy, thus a better understanding of the mechanisms that underlie immune cell effector function is required to improve response rates. In collaboration with Dr. Carl June's group, our lab recently studied a CLL patient who had gone into complete remission following CAR-T therapy. We found that an overwhelming majority of the CAR+ cells had a disruption in the TET2 gene, and experimental knockdown of TET2 resulted in enhanced proliferative capacity and anti-tumor activity of CAR T cells in vitro. Furthermore, T cells with the TET2 knockdown displayed increased expression of TOX2, a member of the HMG-box family of DNA-binding proteins. TOX2 is an immune-specific transcription factor expressed in the human spleen and tonsils. Interestingly, the only known function of TOX2 is that it positively regulates the transcription factor T-BET during the development of natural killer cells, by repressing the inhibitory PD-1 protein. Owing to the increase in TOX2 mRNA levels in TET2 knockdown CAR T cells, in addition to its ability to downregulate a key inhibitory receptor, I hypothesize that TOX2 is a positive regulator of T cell effector function due to its ability to upregulate T-BET. To test this hypothesis, I will pursue three aims. Aim 1 is to manipulate TOX2 expression in human T cells to determine whether it improves T cell differentiation and effector function. Aim 2 is to determine the pattern of TOX2 binding to the genome, with the goal of identifying its transcriptional targets, at both the mRNA and protein levels. Very little is known about how HMG-box proteins bind chromatin in vivo. Thus, this aim will also provide a more fundamental understanding of the chromatin functions of HMG proteins. Aim 3 is to determine levels of TOX2 mRNA in CLL patients who have received CAR T therapy, to determine whether higher TOX2 levels are predictive of better response to CAR T therapy. This study could elevate TOX2 as a novel target for improving CAR T cell function. An understanding of the molecular mechanisms could make it possible to activate TOX2, ultimately improving patient responses to immunotherapy.