癌细胞外泌体的生物分析新方法与临床样品检测研究

Realizing the early diagnosis of cancer is one of the scientific problems of effective cancer treatment. Nowadays, the exosomes secreted by cancer cells are regared as one of the important cancer markers by medical community. Nevertheless, the challenges in separation and detection of exosomes still exist. In this project, the detection of rectal cancer cells is used as model system to construct a micro-nanostructure interface that can specifically recognize exosomes secreted by cancer cells and achieve efficient specific enrichment and immobilization of exosomes. The recognition process of exosomes on the surface of specific micro-nano structures is monitored and its recognition mechanism is studied. The probe is constructed by performing nanometer/enzyme-labeled on antibody molecule (anti-CD147) which can specifically identify the surface antigens of the exosomes of rectal cancer cells. Electrochemical signal amplification is achieved by nanomaterial, enzyme and nanomaterial-enzyme synergy which achieve highly sensitive and highly specific detection of exosomes. The methods of electrochemical and QCM, that have high sensitivity and high specificity to exosomes, are developed, which can realize the efficient enrichment and high sensitive detection of exosomes. Thus, the project provides new methods and new ideas for the early diagnosis of cancer and postoperative monitoring.

如何实现癌症的早期诊断，是癌症有效治疗的科学难题之一。当今医学界认为癌细胞分泌的外泌体是重要的癌症标志物之一，但外泌体的分离和检测仍存在技术挑战。本项目拟以直肠癌细胞的检测为模型体系，构建高效癌细胞外泌体特异识别的微纳结构界面，实现外泌体的高效特异性富集和固定；监测外泌体在特异性微纳结构表面的识别过程，研究其识别机制；对特异性识别直肠癌细胞分泌的外泌体表面抗原的抗体分子（anti-CD147）进行纳米材料/酶标记，构建检测探针；实现电化学信号纳米材料放大、酶放大以及纳米材料-酶协同放大，实现外泌体的高灵敏、高特异性检测，发展癌细胞外泌体的高灵敏、高特异性的电化学、QCM的分析新方法，实现外泌体高效富集和高灵敏检测，为癌症的早期诊断和术后监控的提供新方法和新思路。

癌症是严重威胁人类健康的重大疾病，研究表明原发癌往往并不具有致死性，肿瘤的侵袭和转移才是肿瘤发生和演进过程中最危险的阶段。癌细胞来源的外泌体在癌症生长和转移中起到了至关重要的作用，可以作为一种新型的监测癌症转移的通讯工具。因此，以外泌体为分析对象研究肿瘤转移过程具有重要的意义。本课题以肿瘤来源外泌体作为工具，研究肿瘤来源外泌体的检测方法以及以其作为“探路者”，研究其对受体细胞的侵袭及侵袭后内含物释放细胞生理特性的影响。我们首先建立了一系列传感器实现了对外泌体及细胞内相关物质如核酸、蛋白质等的高灵敏高特异性检测；同时构建了相应的可视化传感技术，实现其可视化监测；将外泌体作为肿瘤转移的可视化通讯工具，建立了癌症转移过程监测方法，利用设计的一系列DNA分子器件，实现了癌细胞来源外泌体miRNAs侵袭成纤维细胞表型转变过程监测和癌细胞来源外泌体miRNAs对上皮间充质转化调控过程监测，直接观察到高转移性癌细胞分泌的外泌体在正常细胞内的miRNAs传递，很好的认知外泌体对癌细胞转移的作用。克服了传统研究方法不直观及无法实现长时间监测等的缺点。