Dissecting auto-reactive and tumor-specific T cells in tumor patients undergoing immune checkpoint inhibitor treatment and suffering from immune-related adverse events

基于检查点标志物的免疫疗法（ICI）在各种肿瘤实体中显示出显著的疗效，但也与显著的免疫相关不良事件（irAE）相关，导致发病率增加和治疗中止，甚至在最坏情况下是致命的。然而，这些ICI诱导的不良反应的病理机制迄今尚未得到充分研究，大多数机制假设和假说都是基于已知的抗肿瘤作用机制。鉴于免疫细胞群体中广泛的PD 1表达，我们假设捕获血液中循环白细胞群体的定量和功能变化可能会提供有关ICI治疗的癌症患者中irAE的原因和驱动机制的新见解。因此，我们在接受抗PD 1抗体治疗的患者中进行了一项纵向全血筛查研究，这项探索性研究的一个主要发现是在irAE诊断前经常观察到的不同活化T细胞群的显著增加，假设在irAE患者中显著升高的活化效应T细胞具有部分自身攻击性，（非恶性组织特异性），我们现在计划通过scRNAseq对转录组和TCR克隆性进行同时研究，除了它们的纵向定量（临床过程中的动态）。与分选的T细胞亚群的TCR批量测序的进一步联系将使我们能够追踪潜在的致病性自身反应性和治疗性T细胞的分化途径和起源群体。其次，使用扩增的TCR克隆的TCR序列和预测算法，将在包括T细胞功能分析的共培养实验中鉴定和验证候选肽，以评估其抗原特异性。鉴定的肿瘤特异性和自身反应性T细胞的T细胞转录组的比较将确定治疗相关的差异，以及根据不同的irAE影响的组织鉴定不同的组织相关的信号传导/免疫/炎症模式。和无关的激活的旁观者T细胞，并为开发灵活和可调节的ICI疗法提供基础。在此过程中，该项目概述的目的是阐明对irAE的发展和进展至关重要的新病理机制。此外，还可以深入了解PD 1如何控制人类的自身免疫过程以及T细胞在自身免疫性疾病早期阶段的作用。

Checkpoint inhibitor-based immunotherapies (ICI) show remarkable efficacy in various tumor entities, but are also associated with significant immune-related adverse events (irAE) leading to increase of morbidity and therapy discontinuation or even being fatal in the worst case. Nevertheless, pathomechanisms of these ICI-induced adverse effects have so far insufficiently investigated and most mechanistic assumptions and hypotheses are based on known mechanisms of action against tumors. Given the widespread PD1 expression among immune cell populations, we hypothesized that capturing quantitative and functional changes in circulating leukocyte populations in the blood might provide new insights regarding the causes and driving mechanisms of irAE in ICI-treated cancer patients. Thus, we conducted a longitudinal whole blood screening study in patients undergoing anti-PD1 antibody treatment.A major finding of this exploratory study was a marked increase in distinct activated T cell populations, which was often observed before diagnosis of irAEs.Assuming that the activated effector T cells, which were significantly elevated in irAE patients, are partially autoaggressive (non-malignant tissue-specific), we now plan to perform simultaneous studies on transcriptome and TCR clonality by scRNAseq in addition to their longitudinal quantification (dynamics in the clinical course). Further linkage with TCR bulk sequencing of sorted T cell subpopulations will allow us to trace the differentiation pathway and populations of origin of potential pathogenically autoreactive and therapeutic T cells. Second, using the TCR sequences of expanded TCR clones and prediction algorithms, candidate peptides will be identified and validated in co-culture experiments including T cell function analyses to assess their antigen specificity. Comparison of the T cell transcriptomes of identified tumor-specific and auto-reactive T cells will determine therapeutically-relevant differences, as well as identify different tissue-related signaling/immune/inflammatory patterns depending on the different irAE-affected tissues.The combination of these studies has the potential to reveal characteristic differences between tumor-specific, auto-aggressive, and unrelated activated bystander T cells and to provide a basis for the development of flexible and adjustable ICI therapies. In doing so, the project outlined aims to elucidate novel pathomechanisms that are critical to the development and progression of irAEs. Furthermore, insights into how PD1 controls autoimmune processes in humans and the role of T cells during the early phase of autoimmune diseases could be gained.