Tumor microenvironment-directed epigenome remodeling of tumor infiltrating T cells

肿瘤浸润T细胞的肿瘤微环境定向表观基因组重塑

• 批准号: 420273313
• 负责人: Dr. Christian Schmidl
• 金额: --
• 依托单位: Leibniz-Institut für Immuntherapie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2019
• 资助国家: 德国
• 起止时间: 2018-12-31 至 2022-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/420273313?language=en
• 关键词: Tumor; microenvironment; directed; epigenome; remodeling

Tumor infiltrating lymphocytes (TILs) are often dysfunctional in the tumor microenvironment (TME), which prevents them from killing cancer cells. This dysfunctional state is thought to be molecularly fixed by a distinct epigenetic program, which hinders durable T cell reprogramming by immunotherapy, and could explain why currently only a minority of patients benefits from so-called checkpoint inhibitors. A chronic T cell receptor (TCR) stimulation was suggested as the main driver of epigenetic fixation of T cell dysfunction irrespective of signals from the TME. However, these studies are confounded by very strong TCR stimulation that likely overrides any other signal the T cells receive. Together with the fact that many studies document a significant impact of costimulatory and coinhibitory signals from the TME on T cell function, I want to challenge the current working model, and prove the hypothesis of tumor microenvironment-directed epigenome remodeling in T cells.I propose to employ a murine transplanted tumor model that allows us to provide low, intermediate and high TCR stimulation in combination with controlled, inducible expression of coinhibitory and costimulatory signals to T cells. By using single-cell chromatin mapping methods, we will identify signal-dependent epigenetic signatures in heterogeneous immune responses, and identify the underlying gene-regulatory programs. We will then delete candidate transcription factors and epigenetic modifiers to dissect their function in establishing and maintaining signal-specific T cell (dys-)functional states in vivo by using a novel single-cell CRISPR method that allows the investigation of hundreds to thousands of knockouts in parallel. With this we will identify concrete targets to enhance immunotherapy and prevent T cell dysfunction, but also provide basic mechanistic insights into the specific molecular pathways that regulate gene expression in T cells. Finally, we will develop high-throughput paired T cell receptor sequence and chromatin accessibility profiling from the same single cells, which allows investigating tumor-directed epigenome remodeling of T cells in primary tumor tissues in relation to their T cell receptor sequence. With the proposed work plan I will establish the concept of tumor microenvironment-directed epigenome remodeling in tumor infiltrating lymphocytes. The acquired data will significantly improve our basic understanding of T cell (dys-)function in tumors, and reveal pathways that establish, maintain, and overcome epigenetically “fixed” T cell states. With the newly developed methods from this proposal for paired T cell receptor and epigenome sequencing, we can leverage our findings from mouse models to primary patient samples. As an outlook, the proposal will set the base for developing molecular markers that predict response to immunotherapy, and reveal novel targets to enhance T cell function for clinical applications.

肿瘤浸润淋巴细胞（TIL）在肿瘤微环境（TME）中通常功能失调，这阻止了它们杀死癌细胞。这种功能障碍状态被认为是由一种独特的表观遗传程序在分子上固定的，这阻碍了免疫疗法对T细胞的持久重编程，并可以解释为什么目前只有少数患者受益于所谓的检查点抑制剂。慢性T细胞受体（TCR）刺激被认为是T细胞功能障碍的表观遗传固定的主要驱动力，而不管来自TME的信号如何。然而，这些研究被非常强的TCR刺激所混淆，这种刺激可能会覆盖T细胞接收的任何其他信号。结合许多研究证明了来自TME的共刺激和共抑制信号对T细胞功能的显著影响的事实，我想挑战当前的工作模型，并证明T细胞中肿瘤微环境指导的表观基因组重塑的假设。我建议采用小鼠移植肿瘤模型，该模型允许我们提供低、中和高TCR刺激，对T细胞的共抑制和共刺激信号的可诱导表达。通过使用单细胞染色质作图方法，我们将识别异质性免疫应答中的信号依赖性表观遗传特征，并识别潜在的基因调控程序。然后，我们将删除候选转录因子和表观遗传修饰剂，通过使用一种新的单细胞CRISPR方法来解剖它们在体内建立和维持信号特异性T细胞（dys-）功能状态的功能，该方法允许并行研究数百至数千个敲除。有了这个，我们将确定具体的目标，以加强免疫治疗和预防T细胞功能障碍，但也提供了基本的机制见解的具体分子途径，调节基因表达的T细胞。最后，我们将从相同的单细胞开发高通量配对T细胞受体序列和染色质可及性分析，这允许研究原发性肿瘤组织中T细胞与其T细胞受体序列相关的肿瘤定向表观基因组重塑。通过提出的工作计划，我将建立肿瘤微环境指导的肿瘤浸润淋巴细胞表观基因组重塑的概念。获得的数据将显着提高我们对肿瘤中T细胞（dys-）功能的基本理解，并揭示建立，维持和克服表观遗传“固定”T细胞状态的途径。通过这项配对T细胞受体和表观基因组测序提案中新开发的方法，我们可以利用我们从小鼠模型到主要患者样本的发现。作为展望，该提案将为开发预测免疫治疗反应的分子标记物奠定基础，并揭示增强T细胞功能的新靶点以供临床应用。