基于液滴微流控的单细胞多维度分析新体系的建立及对大肠癌循环肿瘤细胞异质性的解析

循环肿瘤细胞（circulating tumor cell, CTC）是预测转移复发的重要标志物。但CTC群体存在高度异质性，因此，亟需从单细胞水平分辨CTC中的转移活性细胞亚群，但目前缺少从分子和功能结合的多维度水平解析CTC异质性的方法。申请团队前期建立了基于液滴微流控的单细胞捕获分选机器人，可实现对全血中稀有靶细胞的自动化单细胞分选，同时，我们及他人研究显示，侵袭是转移活性细胞重要的功能表型，此外，前期构建了针对EpCAM和转移性大肠癌细胞表面蛋白的多个核酸适配体分子信标探针。本研究拟在上述工作基础上，创建基于液滴微流控的集成化单细胞分析系统，自动化完成基于探针标记的CTC单细胞分选、靶蛋白定量分析、侵袭能力分析，并结合单细胞外显子测序技术，实现对单细胞CTC在基因、蛋白、侵袭功能的多维度分析。以大肠癌为研究对象，解析CTC异质性与临床的相关性，为临床诊疗提供新的研究体系和检测方法

Circulating tumor cell (CTC) is an important biomarker for predicting metastasis and recurrence. It is urgent to distinguish the sub-populations of metastatic CTC at the single-cell level due to its high heterogeneity. However, currently there is no method to analyze the CTC heterogeneity based on the combination of molecule and function with the multidimensional level. Our project team previously established a robot system based on droplet microfluidics for capturing and sorting single-cell, which realized automatically sorting rare target cells at single-cell level from patients’ whole blood. At the same time, our research and other teams have indicated that invasion is an important functional phenotype of metastasis activity. Additionally, we have designed multiple aptamer molecular beacons targeting EpCAM and the proteins of metastatic colorectal cancer cell surface. Grounded on the work above, this study intends to build an integrated single-cell analysis system based on droplet microfluidics, which realizes sorting of the single CTC with probe labeling, quantitative analysis of target proteins and analysis of invasion ability. Combined with single-cell exome sequencing technology, the multidimensional single-cell analysis system in gene, protein and invasion function is proposed. Taking colorectal cancer as the research object, this study analyzes the correlation between CTC heterogeneity and clinic, and provides a new research system and detection method for clinical diagnosis and treatment.