4DN Interrogation of T Cell Exhaustion in Cancer

项目摘要/总结 增强T细胞识别和杀死肿瘤细胞的能力的免疫疗法已经发生了变革 免疫疗法在治疗人类癌症中是有效的，但免疫疗法并非对所有患者或癌症都有效，因此 需要进行研究以探究T细胞对癌症的持久应答的分子基础。一个关键的障碍， 肿瘤浸润性T细胞的持续活化是T细胞“耗竭”的发展，这导致肿瘤浸润性T细胞的持续活化。 抑制性表面受体的稳定表达、对肿瘤抗原的不良反应和低细胞增殖， 体内T细胞的持久性。然而，迄今为止，研究基因调控机制一直是困难的 由于缺乏敏感的基因组工具来研究， 患者的免疫细胞。我们最近开发了一套高通量表观基因组技术 能够测量三维（3D）基因组构象和单细胞染色质 来自人类肿瘤的原代T细胞的可及性。在拟议的研究中，我们的目标是利用这些方法， 确定4D核组（4DN）组织和可及性的变化，这些变化是人类发育的基础。 T细胞衰竭。在目标1中，我们将定义在人类T细胞耗竭中发生的3D基因组相互作用， 晚期皮肤癌患者。耗竭相关基因组构象将在 几种癌症类型，以确定一个共识耗尽概况，这些发现将与 染色质可及性和基因表达数据，以确定3D变化的转录效应。在目标2中， 将使用一种新的嵌合体， 抗原-受体（CAR）-T细胞模型。在目标3中，我们将使用CRISPR/Cas9基因组干扰这些相互作用， 在原代T细胞中进行编辑，再加上单细胞表观基因组读出，以设计出改进的，持久的，下一代 免疫疗法的一代。如果成功，这些发现将对未来的设计产生直接影响。 免疫治疗策略，这将对癌症患者的临床护理产生重大影响。最后我们 将通过免费分发协议和数据以及发布自定义数据来促进这些发现的传播。 软件工具，我们将使用这些研究作为4DN网络的协作启动点。我们预计 这些结果将导致对T细胞耗竭的分子调控的新见解，并作为一种新的研究方法。 Satpathy博士有效的研究计划，以建立他的独立实验室在接口， 免疫学和基因组科学。

PROJECT ABSTRACT/SUMMARY Immunotherapies that enhance the ability of T cells to recognize and kill tumor cells have been transformational in the treatment of human cancer, but immunotherapy is not effective in all patients or cancers, and therefore studies interrogating the molecular basis for durable T cell responses to cancer are needed. A critical barrier for the sustained activation of tumor-infiltrating T cells is the development of T cell ‘exhaustion,’ which leads to the stable expression of inhibitory surface receptors, poor response to tumor antigens, and low cell proliferation and persistence of T cells in vivo. However, to date, it has been difficult to study the gene regulatory mechanisms that control the development of T cell exhaustion in humans, due to a lack of sensitive genomic tools to study primary immune cells from patients. We recently developed a suite of high-throughput epigenomic technologies that enable the measurement of three-dimensional (3D) genome conformation and single-cell chromatin accessibility in primary T cells from human tumors. In the proposed research, we aim to utilize these methods to identify changes in 4D nucleome (4DN) organization and accessibility that underlie the development of human T cell exhaustion. In Aim 1, we will define 3D genome interactions that occur in human T cell exhaustion in patients with advanced skin cancer. Exhaustion-associated genome conformation will be compared across several cancer types to identify a consensus exhaustion profile, and these findings will be integrated with chromatin accessibility and gene expression data to identify transcriptional effects of 3D changes. In Aim 2, we will determine the dynamics and reversibility of regulatory 3D interactions in exhaustion using a novel chimeric antigen-receptor (CAR)-T cell model. In Aim 3, we will perturb these interactions using CRISPR/Cas9 genome editing in primary T cells, coupled with single-cell epigenomic read-outs, to engineer improved, durable, next- generation immunotherapies. If successful, these findings will have a direct impact on the future design of immunotherapy strategies, which will have a significant impact on the clinical care of cancer patients. Finally, we will facilitate the dissemination of these findings by freely distributing protocols and data and releasing custom software tools, and we will use these studies as a collaborative launch point in the 4DN network. We anticipate that these results will lead to novel insights into the molecular regulation of T cell exhaustion and serve as an effective research program for Dr. Satpathy to establish his independent laboratory at the interface of immunology and genome science.