新型双联肽-藤黄酸前体药物构建及其抗膀胱癌作用研究

To find a highly selective treatment of tumor cells is particularly important. Tumor specific endo-protease activated prodrug is a promising treatment strategy. Legumain is a newly discovered cysteine protease that highly expressed in the tumor microenvironment, coupled with the digestion reaction on a substrate with a unique high selectivity requirement, making it an attractive drug activation enzyme candidate. This subject will construct an innovative double coupling peptides based on legumain cutting the substrate and tat peptide penetrating the cell membrane.Then, garcinia acid will be linked to the double coupling peptides to create a novel drugs. The drug precursor will be specifically cut and released in tumor tissue, and be carried by the tat peptide into the cancer cells, so that the effective concentration of anti-tumor effect of Garcinia acid will greatly reduced. The successful implementation of this project will provide an efficient specific and initiative targeted therapy for the bladder cancer.

找到一种对肿瘤细胞具有高选择性的治疗方法尤为重要，内切蛋白酶激活型肿瘤前体药物就是其中一种极具前景的治疗策略。Legumain是新发现的半胱氨酸蛋白酶,在肿瘤组织微环境特异性高表达，而且其酶切反应中对反应底物有独特高选择性要求，使它成为一个富有吸引力的抗肿瘤前药激活酶的候选。本课题通过将legumain切割底物与穿膜肽Tat相连接，制备创新性双联肽分子；继而将新型药物藤黄酸与双联肽分子偶联，形成双联肽靶标前体药物。该前体药物特异地在肿瘤组织中被切割释放，从而具有靶向性；同时穿膜肽Tat增加了药物对肿瘤细胞的穿透杀伤效果，使藤黄酸抗肿瘤作用需要的药物浓度有效降低。本研究的成功实施将为膀胱癌的高效特异性主动靶向治疗提供实验基础。

藤黄素是新型植物来源抗肿瘤药物，具有来源多、作用广、毒副作用小等优点，其抗肿瘤活性有待提高、相关作用机制有待进一步阐明。本研究成功合成了新型藤黄酸-穿膜肽TAT，制备了双联肽靶标前体药物，构建了靶向高效藤黄酸导入体系；相关作用机制研究发现，藤黄酸能够有效促进起抗癌作用的长片段非编码RNA（lncRNA）GAS5表达，而GAS5能够作为转录调节因子结合转录因子E2F4，进而促进E2F4锚定至EZH2启动子区域，通过下调EZH2转录活性促进膀胱癌细胞凋亡及在体生长；另一方面，藤黄酸能够有效抑制起促癌作用的lncRNA SPRY4-IT1表达，减少SPRY4-IT1作为“miRNA海绵”对miR-101-3p的吸附，进而增强miR-101-3p对EZH2 mRNA 3’UTR活性的抑制作用，在转录后水平下调EZH2表达，从而在细胞和动物实验水平有效抑制膀胱癌生长及侵袭转移。本研究有效提高了藤黄素的临床抗肿瘤应用价值，同时证实了GAS5/E2F4/EZH2及SPRY4-IT1/miR-101-3p/EZH2通路对膀胱癌生物学活性的调控作用，进一步解析了膀胱癌发生发展机制。