基于功能核酸/MOFs生物探针的血液miRNA指纹分析用于乳腺癌早期诊断

Breast cancer (BC) is among the most common cancers worldwide, and continues to pose great health threat to Chinese women. As survival rate of BC is closely associated with its diagnostic state, early screening and diagnosis are thus considered as the key to improve treatment efficiency. Large equipment-based techniques can hardly meet the requirement of generalized large-scale diagnosis of BC, spurring cost-effective, highly efficient, operation-friendly and noninvasive detection alternatives. After comprehensive literature review and summary of our previous research work, we propose that combined analysis of multiple miRNAs to create fingerprints would potentially improve the accuracy and specificity of BC diagnosis by blood test. We plan to accomplish the following tasks: a. Design of high-performance bio-probes using functional nucleic acids and metal-organic frameworks (MOFs) to overcome problems like high interference and low target concentrations in blood, so as to realize good sensitivity, selectivity and reproducibility for in-site miRNA detection; b. Screening of efficient combined miRNA markers to form specific fingerprints to allow for cross-reactive pattern recognition. It will circumvent technical issues resulted from variation in abundance quantification due to the lack of reliable inner reference and low specificity with a single miRNA biomarker. Approval and execution of the project as-proposed will definitely open up new perspectives for automotive BC diagnosis, and promote the development of new methods and new ideas to screen and monitor multiple cancers by blood miRNA analysis.

乳腺癌是世界上发病率较高的一类癌症，严重威胁我国女性的身心健康。乳腺癌的诊断与预后紧密相关，早发现、早诊断、早治疗是提高治愈率的关键。大型诊断设备无法满足常规化、大规模乳腺癌筛查，急需开发成本低、效率高、操作简便的非侵入式替代诊断技术。本项目在充分调研文献资料和总结课题组前期研究工作的基础上提出：联合检测多个miRNA形成指纹图谱可以提高临床血检诊断乳腺癌的准确性和特异性。拟完成如下开创性工作：a. 针对血液样本干扰组分多、靶标浓度低等问题，设计功能核酸/MOFs多级信号放大检测探针，用于提高miRNA即时检测的灵敏度、选择性和重复性; b.筛选高效的miRNA组合标志物形成指纹图谱，交叉响应、模式识别，避免因缺少统一性分子内标而造成的丰度测定差异和单个miRNA特异性差等关键性问题。本项目有望为智能化乳腺癌筛查研究提供新方法和新思路，为血液miRNA联合诊断多种癌症的新技术奠定基础。

血清样本中miRNA靶标的多重检测有望为乳腺癌等重大慢性病疾病的诊断提供重要标志物，高效的信号放大技术是构建超灵敏检测平台的关键，可以有效降低生物样本中的背景干扰。本项目针对血液样本干扰组分多、靶标浓度低等问题，设计功能核酸/MOFs多级信号放大检测探针，提高了miRNA即时检测的灵敏度和选择性；利用功能核酸/MOFs多级信号放大检测探针构建的鲍曼不动杆菌的荧光感应传感器，可以检测到血液样本中浓度低至10 CFU/mL的病原菌；为了解决纳米材料多重检测miRNA的稳定性问题，将LbaCas12a作为信号放大器，开发出多功能CRISPR-Cas12a平台，可以同时检测4个miRNA标志物，不受其它共存非靶标miRNA的干扰；利用该平台的通用性，我们相继开发了基于核酸编码技术和CRISPR-Cas解码技术的小分子检测平台和细菌耐药基因多重筛查技术。本项目的执行为血液miRNA联合诊断多种癌症的新技术奠定了基础，也为基于纳米/生物材料的信号放大策略在临床场景中的应用提供了新思路。