Inflammatory processes in the development of cholangiocarcinoma in Primary Sclerosing Cholangitis – Do the T cells play a role?

原发性硬化性胆管炎胆管癌发展中的炎症过程 â T 细胞发挥作用吗？

• 批准号: 426654902
• 负责人: Professor Dr. Nicola Gagliani, Ph.D.
• 金额: --
• 依托单位: Klinik und Poliklinik für Allgemein-, Viszeral- und Thoraxchirurgie
• 依托单位国家: 德国
• 项目类别: Clinical Research Units
• 财政年份: 2019
• 资助国家: 德国
• 起止时间: 2018-12-31 至 2023-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/426654902?language=en
• 关键词: Inflammatory; processes; development; cholangiocarcinoma; Primary

Extrahepatic cholangiocarcinoma (CCA) is the most dreaded comorbidity in Primary Sclerosing Cholangitis (PSC). The prognosis is very poor and as of yet the pathogenesis is unknown, but there is evidence that the immune system and the microbiota contribute towards promoting this disease. One suggested prerequisite for carcinogenesis in PSC-associated CCA (PSC-CCA) is a strong association of chronic biliary inflammation with biliary metaplasia and dysplasia, typical of PSC. In addition, differences in the composition of the intestinal and biliary microbiota of patients with PSC have been shown. Finally, the impact of the microbiota on T cell function, which is often determined by the secretion of cytokines and the expression of co-inhibitory receptors, has recently been implicated in carcinogenesis. On the basis of all this, we hypothesize that the PSC specific microbiota favours the pro-tumorigenic function of T cells, ultimately leading to carcinogenesis in PSC patients. The first objective of our study is to define the functional phenotype of human tumour infiltrating T cells in PSC-CCA by using single-cell RNA/TCR sequencing in combination with Cellular Indexing of Transcriptome and Epitopes by sequencing (CITE-seq) and RNA seq of total tissue. Results will allow us to identify new cell populations and their cell relationships with either a pro- or anti-tumorigenic functional phenotype. In addition, by defining the co-inhibitory receptor profile of these populations, we will set the basis for designing ad hoc immune therapies (e.g. anti-PD1). Finally, we will infer the potential interaction among the cells and the tissue by matching differentially expressed genes identified in the single cell RNA analysis to expression patterns of known interaction partners (e.g. cytokine-receptor pairings) in the surrounding tissue, identified by total tissue RNA sequencing. The second objective is to identify specific PSC-CCA microbiota able to shape the T cell response. Therefore, we will first sequence the faecal and biliary microbiota from PSC-CCA patients, and then test the microbiota - T cell interaction by transferring human microbiota into germ-free mice. The third objective is to develop a specific mouse model that exhibits a PSC phenotype and develops extrahepatic CCA. We will achieve this by using the bile duct injection technique to introduce carcinogenic substances directly into the biliary tree of Mdr2-/- mice. Additionally, using the Cre-Lox-system, we will develop a transgenic mouse model with a conditional overexpression of the oncogene kras and deletion of the tumour suppressor gene pten in cholangiocytes. In summary, the results of this project will contribute towards the understanding of the pathogenesis of PSC-associated CCA. They will reveal immunological mechanisms and will help the clinical translation of possible therapeutic targets in order to directly treat patients suffering from this disease.

肝外胆管癌（CCA）是原发性硬化性胆管炎（PSC）最可怕的合并症。预后非常差，发病机制尚不清楚，但有证据表明免疫系统和微生物群有助于促进这种疾病。PSC相关CCA（PSC-CCA）中致癌的一个先决条件是慢性胆道炎症与PSC典型的胆道化生和发育不良密切相关。此外，已经显示出PSC患者的肠道和胆道微生物群组成的差异。最后，微生物群对T细胞功能的影响，这通常是由细胞因子的分泌和共抑制受体的表达决定的，最近已经涉及到致癌作用。基于所有这些，我们假设PSC特异性微生物群有利于T细胞的促肿瘤发生功能，最终导致PSC患者的致癌作用。我们研究的第一个目的是通过使用单细胞RNA/TCR测序结合通过测序的转录组和表位的细胞索引（CITE-seq）和总组织的RNA seq来定义PSC-CCA中人肿瘤浸润T细胞的功能表型。结果将使我们能够确定新的细胞群和它们的细胞与促或抗肿瘤功能表型的关系。此外，通过定义这些人群的共抑制受体谱，我们将为设计特定免疫疗法（例如抗PD 1）奠定基础。最后，我们将通过将在单细胞RNA分析中鉴定的差异表达基因与通过总组织RNA测序鉴定的周围组织中已知相互作用伴侣（例如，精氨酸-受体配对）的表达模式相匹配来推断细胞和组织之间的潜在相互作用。第二个目标是鉴定能够塑造T细胞应答的特定PSC-CCA微生物群。因此，我们将首先对PSC-CCA患者的粪便和胆汁微生物群进行测序，然后通过将人微生物群转移到无菌小鼠中来测试微生物群- T细胞相互作用。第三个目标是开发一种表现出PSC表型并发生肝外CCA的特定小鼠模型。我们将通过使用胆管注射技术将致癌物质直接引入Mdr 2-/-小鼠的胆管树来实现这一点。此外，使用Cre-Lox系统，我们将开发一种转基因小鼠模型，在胆管细胞中癌基因kras的条件性过表达和肿瘤抑制基因pten的缺失。总之，本研究的结果将有助于对PSC相关CCA发病机制的理解。它们将揭示免疫学机制，并将有助于临床翻译可能的治疗靶点，以直接治疗患有这种疾病的患者。