基于发光氧化石墨烯的端粒酶活性影像学分析及其对肿瘤边界精准定位

Telomerase is a key biological enzyme to control cell proliferation, which is highly expressed in tumor cells, but it is normally inactive in somatic cells. Thus, the reactivation of telomerase is considered as a broad- spectrum marker of tumorigenesis. However, it remains an atelic challenge to use its in-situ assay in living animals as a powerful tool for the precise diagnosis and auxiliary surgery of cancers. In this project, we will develop a Nanofluorescence imaging method by fluorescent graphene oxide to detect the telomerase activity of living cells for the precise location of living tumor boundary. 1) Synthesize the fluorescent graphene oxide carrier that can efficiently penetrate the cell membrane and into the nucleus, and then enhance the loading efficiency of telomerase primer and targeting ability of tumor cells by surface modification. 2) To explore the assemble method between the telomerase primers and the fluorescent graphene oxide carrier, and then to design and construct the internal labeled fluorescent nanoprobes sensitively toward the telomerase activity. 3) To develop the quantitative analysis method of telomerase activity in whole cells based on the nanoprobe, establishing the corresponding relationship between the expression level of telomerase activity and the degree of tumor cell deterioration. 4) Through the animal live tumor model, combined with the microsurgery technology, establisht the visualizational imaging method for the precise location of living tumor boundary. The expected results of this project will provide new technical means for early diagnosis and accurate surgery of cancer.

端粒酶活性是控制细胞增殖和永生化的关键因素，在肿瘤细胞内高表达，而正常体细胞内活性被抑制。因此，端粒酶的重新激活被视为广谱的肿瘤发生发展标志物，但通过活体内端粒酶活性的原位影像检测实现肿瘤诊断及精准手术仍然是一个挑战。本项目拟以发光氧化石墨烯为载体，发展全细胞内端粒酶活性的荧光成像方法，通过荧光可视化影像分析实现肿瘤边界的精准定位。1）合成筛选能高效穿透细胞膜并进入细胞核的发光氧化石墨烯载体，通过表面改性提高其对端粒酶引物的负载效率及肿瘤细胞的靶向能力；2）探索端粒酶引物与载体组装方式，设计并构建端粒酶活性敏感响应的氧化石墨烯荧光内标纳米探针；3）发展基于该探针的全细胞内端粒酶活性的定量分析方法，建立端粒酶活性表达水平与肿瘤细胞恶化程度的对应关系；4）通过动物活体肿瘤模型，结合显微手术技术，建立肿瘤边界精准定位的影像学可视化方法。预期成果将为癌症的早期诊断和精准手术提供新的技术手段。

细胞内端粒酶活性表达水平与肿瘤细胞的恶变程度紧密相关，因此建立细胞内端粒酶活性的定量检测方法对监测癌症发生发展过程具有重要意义。本项目以细胞内端粒酶活性的定量检测为目标牵引，发展了全细胞内端粒酶活性的荧光成像方法，通过荧光可视化影像分析实现肿瘤边界的精准定位。主要发现点和成果如下：1）合成筛选了能高效穿透细胞膜并进入细胞核的发光氧化石墨烯载体，通过表面改性提高其对端粒酶引物的负载效率及肿瘤细胞的靶向能力；2）探索了端粒酶引物与载体组装方式，设计并构建端粒酶活性敏感响应的氧化石墨烯荧光内标纳米探针；3）发展了基于该探针的全细胞内端粒酶活性的定量分析方法，建立端粒酶活性表达水平与肿瘤细胞恶化程度的对应关系；4）通过动物活体肿瘤模型，结合显微手术技术，建立了肿瘤边界精准定位的影像学可视化方法。上述研究成果在国际期刊发表 SCI 论文 11 篇，其中高影响力期刊有Angew. Chem. Int. Ed.（1 篇），Chem. Commun. （1篇）Sensor. Actuat. B-Chem.（1 篇），Chinese Chemical Letters （1篇）。授权发明专利一项。