基于类器官共培养体系的肝细胞癌反应性TCR筛选
Screening of Reactive TCRs against Hepatocellular Carcinoma Based on Organoid Co-culture System
孙倩 1张晓明1
作者信息
- 1. 苏州大学苏州医学院生命科学学院，苏州 215021
- 折叠
摘要
目的：从肝细胞癌患者肿瘤浸润T细胞中筛选肿瘤反应性T细胞受体，并建立基于类器官与TCR报告细胞共培养的体外功能验证体系。方法：收集肝细胞癌患者肿瘤及癌旁组织，建立患者来源的肿瘤类器官模型。对肿瘤浸润T细胞行单细胞TCR测序，依据克隆扩增分析筛选候选TCR序列。构建TCR慢病毒表达载体，转导至内源性TCR敲除且稳定表达NFAT-ZsGreen报告基因的Jurkat细胞中。将TCR-T报告细胞分别与自体肿瘤类器官及永生化B细胞共培养，通过检测ZsGreen荧光评估TCR激活状态。结果：成功构建HCC-60患者来源的肿瘤类器官及永生化B细胞系。单细胞测序显示肿瘤组织中存在T细胞克隆扩增，筛选出8条候选TCR。功能验证表明，仅候选TCR30在与肿瘤类器官共培养时诱导明显NFAT信号激活，而对永生化B细胞无显著反应。结论：成功建立基于患者来源类器官的TCR功能筛选平台，并初步鉴定出一条具有肿瘤反应性的候选TCR。该体系为个体化TCR-T细胞治疗靶点的筛选提供了实验基础。
Abstract
Objective：To screen tumor-reactive T cell receptors (TCRs) from tumor-infiltrating T cells of patients with hepatocellular carcinoma (HCC) and to establish an in vitro functional validation system based on a co-culture model of organoids and TCR reporter cells.Methods：Tumor and adjacent non-tumor tissues were collected from HCC patients, and patient-derived tumor organoid models were established. Single-cell TCR sequencing was performed on tumor-infiltrating T cells, and candidate TCR sequences were screened based on clonal expansion analysis. TCR lentiviral expression vectors were constructed and transduced into Jurkat cells with endogenous TCR knockout and stable expression of the NFAT-ZsGreen reporter gene. TCR-T reporter cells were co-cultured with autologous tumor organoids or immortalized B cells, and TCR activation was evaluated by detecting ZsGreen fluorescence.Results：Patient-derived tumor organoids and an immortalized B cell line were successfully established from the HCC-60 patient. Single-cell sequencing revealed clonal expansion of T cells in the tumor tissue, and eight candidate TCRs were screened. Functional validation demonstrated that only candidate TCR30 induced significant NFAT signal activation upon co-culture with tumor organoids, while no significant response was observed against immortalized B cells.Conclusion：A functional TCR screening platform based on patient-derived organoids has been successfully established, and one tumor-reactive candidate TCR has been preliminarily identified. This system provides an experimental foundation for the screening of individualized TCR-T cell therapy targets.关键词
肿瘤免疫/T细胞受体/肝细胞癌/类器官/TCR-T细胞/单细胞测序Key words
Tumor Immunology/T cell receptor/Hepatocellular carcinoma/Organoid/TCR-T cells/Single-cell sequencing引用本文复制引用
孙倩,张晓明.基于类器官共培养体系的肝细胞癌反应性TCR筛选[EB/OL].(2026-07-03)[2026-07-05].http://www.paper.edu.cn/releasepaper/content/202607-6.学科分类
肿瘤学/生物科学研究方法、生物科学研究技术
