高灵敏纳米荧光探针用于动态可视化监测microRNA调控癌细胞凋亡及其在药效评估的应用

Targeting microRNA (miRNA)-based tumor therapy is a research hotspot in the current field of precise treatment of tumors. There are different expression levels of miRNA in different tumors or different subtypes of tumors, and miRNA has been identified to show carcinogenic effect or carcinostatic effect. Therefore, to achieve the goal of developing the miRNA-based precision therapy for tumors, it is urgent to monitor the effect of changes in the expression of miRNA on apoptosis in real time. The traditional biological analysis methods of miRNA and apoptosis are in the dead cell state. And fluorescence imaging can only detect miRNA or apoptosis in living cells, which can’t reflect the real changes of both. For the purpose of dynamically visualizing the effect of changes in the expression of miRNA on apoptosis, it is necessary to construct highly sensitive and specific fluorescent probes for real-time in situ detection of intracellular miRNA and apoptosis. A new fluorescent nanoprobe with multiple detection functions will be constructed in this project. The nanoprobe will be prepared by assembling biocompatible nanomaterials with recognition groups targeting miRNA and apoptosis-related protein markers. The nanoprobe can detect intracellular miRNA and apoptosis in situ with high sensitivity and selectivity, and then dynamically and visually monitor the effect of changes in the expression of miRNA on apoptosis in the models of different tumors or different subtypes of tumors. This probe can provide important information for the development of novel anti-cancer drugs targeting miRNA, the evaluation of therapeutic efficacy and the formulation of precise personalized treatment options.

MicroRNA（miRNA）靶向治疗是当前肿瘤精准治疗领域的研究热点。由于不同类型或不同亚型的肿瘤细胞中的miRNA表达量不同，发挥的致癌或抑癌作用也不同，因此要发展基于miRNA的肿瘤精准治疗，迫切需要实时监测miRNA表达量变化对细胞凋亡的影响。目前发展的方法大都是在细胞裂解后检测miRNA和细胞凋亡，或者对细胞内miRNA或细胞凋亡的单一检测，很难在活细胞水平上实时监测miRNA表达量变化对细胞凋亡的影响。因此，构建高灵敏度、高特异性的分析方法实现活细胞内miRNA和细胞凋亡的实时原位检测是非常必要的。本项目拟设计miRNA和凋亡标志物特异识别的高灵敏、高特异性的新型纳米荧光探针。建立生物学模型，动态可视化监测不同类型或不同亚型肿瘤细胞模型中，miRNA表达量的变化对细胞凋亡的影响，为研发以miRNA为靶点的抗癌新药、评估肿瘤治疗疗效及制定精准的个性化治疗方案提供了重要信息。

发展高灵敏、高特异性的新型纳米荧光探针，实现在活细胞水平实时监测miRNA表达量变化对细胞凋亡的影响，对研发以miRNA为靶点的抗癌新药、评估肿瘤治疗疗效及制定精准的个性化治疗方案，具有十分重要的研究意义。本工作建立了简单、具有多重检测功能的荧光检测策略，实现了在活细胞水平对miRNA和凋亡蛋白的高灵敏度、高特异性检测，能够实时原位监测miRNA表达量变化对细胞凋亡的影响；将Au-Se键和多色成像技术引入同一纳米平台，构建了能有效抗生物硫醇干扰，稳定性更高的金硒双色探针dSe-SNAP，避免出现假阳性结果，能有效区分正常细胞和癌细胞；基于AuNPs设计了新型双色纳米探针，通过识别并监测两种生物标志物的表达水平，实现了以可视化方法研究肿瘤微环境变化对癌细胞迁移和侵袭性质的影响。在国家自然科学基金资助下，在ACS Sensors、Biomaterials Science、Pharmaceutics、Biosensors等学术期刊上发表标注基金资助 SCI 论文4篇，影响因子均大于5.0；申请国家发明专利1件；指导硕士研究生3名。总体来说，较好地完成了项目申请书中的预期研究成果目标。