肿瘤外泌体标志物核酸适体的筛选及其在液态活检中的应用

Tumor derived exosomes have been found to accumulate in human blood and carry molecular information of the parent tumor. Therefore, targeting cancer-cell-derived exosomes provides a promising tool for tumor biology and early disease detection without the need for an invasive biopsy. Unfortunately, the lack of highly efficient and sensitive isolation of cancer-cell-derived exosomes in the circulation remains challenging in liquid biopsy. In this project, aptamers against glypican-1 (GPC1), a specific surface marker of cancer exosomes, will be generated. A microfluidic chip functionalized with aptamers against tumor derived exosomes and nano-interface will be designed and fabricated for highly sensitive and selective tumor derived exosomes analysis from body fluids. Based on the specific marker of cancer exosomes, the proposed method is capable of efficient capture of targeted exosomes. On the other hand, using aptamers as affinity ligands rather than antibody avoids several antibody associated limitations such as protein inactivation, low enrichment efficiency, high cost and affected cell viability during release process. Moreover, microfluidics and nanostructured interface offer the advantages of high capture efficiency and increase immuno-detection sensitivity. The successful implementation of the project will deliver a useful tool to facilitate basic and clinical investigations of exosomes for non-invasive disease diagnosis and precise treatment.

肿瘤生长过程中不断释放外泌体到周围环境中，肿瘤外泌体的捕获及检测对肿瘤的早期检测及预后判断方面具有重大意义。如何高纯度、高效地分离和检测肿瘤细胞来源的外泌体是其应用于液态活检的瓶颈问题。本项目拟通过核酸适体筛选技术，获得多条特异性识别肿瘤外泌体标志物GPC1的核酸适体，结合微流控高通量富集方法及纳米生物界面构筑，实现体液中特异性肿瘤外泌体高灵敏、高选择性的快速富集检测。以肿瘤外泌体标志物为靶点，有望解决目前无法特异性捕获肿瘤来源外泌体的局限性；以核酸适体作为捕获探针，有望解决抗体易失活、捕获效率低、成本高、难以无损释放等问题；以微流控芯片及纳米生物界面为捕获平台，能够提高外泌体富集效率及检测灵敏度。项目的成功实施将为癌症扩散早期诊断和预警、治疗方案评估等提供重要的检测途径。

肿瘤生长过程中不断释放外泌体到周围环境中，对肿瘤外泌体的精准检测有助于肿瘤的早期检测及预后判断方面具有重大意义。如何高纯度、高效检测肿瘤细胞来源的外泌体是其应用于液态活检的瓶颈问题。本项目通过结合分子拥挤、机器学习、分子对接等发展了多种核酸适体筛选新方法，获得多条特异性识别肿瘤外泌体标志物PD-L1、EpCAM的核酸适体，结合微量热泳动及单分子液滴扩增实现了体液中特异性肿瘤外泌体高灵敏、高选择性的快速检测。项目的成果为外泌体生物功能研究、癌症扩散早期诊断和预警、免疫治疗方案评估等提供重要的化学测量技术。在项目实施过程中在国际化学期刊上发表SCI论文13篇，申请专利1项，培养优秀青年基金获得者1人，培养了生物分析化学博士2名，硕士2名。