靶向治疗肿瘤的新型阳离子聚（β-氨基酯）/RNA自组装纳米粒的研究

The development of efficient and low-toxic non-viral delivery systems with high targeting ability is one of the most urgent and challenging key issues for targeting therapy of cancers through RNA interference (RNAi) technology. In this subject, a series of novel cationic poly (β-amino esters) derivatives (PAEs) is synthesized based on our previous work, siRNA or shRNA targeting Survivin, Notch1 and Snail genes are designed, and human mammary carcinoma cell lines such as MCF7 and MDA-MB-231 are chosen as in vitro models. The relationship between the structures of PAEs and the RNAi efficiency and cytotoxicity of PAEs/RNA self-assembly nanoparticles (PAEN)with one kind or several kinds of RNA simultaneously is revealed by cell biological technologies such as flow cytometry and Western Blot, and obtain non-viral RNA delivery systems with high targeting ability, high RNAi efficiency, low toxicity and potential application value. The intracellular trafficking and the mechanisms of synergistic effect when combined with chemotherapeutics of PAEN are illustrated. The RNAi and anti-tumor efficacy and preliminary biological safety of PAEN in mice human mammary carcinoma xenograft and in situ tumor models are evaluated. This research will be valuable to achieve the clinical application of RNAi-based gene therapy of cancer as soon as possible.

发展靶向性强、转染效率高和毒性低的非病毒输送系统是RNA干扰（RNAi）技术用于靶向治疗肿瘤领域最为迫切并具挑战性的关键课题之一。本项目以现有工作为基础，再合成一系列新型阳离子聚（β-氨基酯）衍生物，以基因Survivin、Notch1 和Snail等为靶点的siRNA或shRNA，选择人乳腺癌细胞MCF7和MD-MBA-231等，采用流式细胞术、Western Blot等技术，揭示阳离子聚（β-氨基酯）衍生物结构与其和RNA自组装纳米粒（含一种或多种RNA共输送）干扰效率和细胞毒性关系，优化获得靶向性强、干扰效率高、毒性低且具潜在应用价值的RNA非病毒输送系统；阐明新型阳离子聚合物/RNA自组装纳米粒在细胞内转运过程及与化疗药物协同增效作用机制，评价其在人乳腺癌裸小鼠移植瘤模型和原位瘤模型上的干扰效率和抗肿瘤效果及初步生物安全性等，为基于RNAi的肿瘤基因治疗早日实现临床应用奠定基础

发展靶向性强、转染效率高和毒性低的非病毒输送系统是RNA干扰（RNAi）技术用于靶向治疗肿瘤领域最为迫切并具挑战性的关键课题之一。本项目设计合成了一系列对肿瘤组织过表达的基质金属蛋白酶或肿瘤细胞内涵体/溶酶体酸性环境和具有刺激响应性的新型聚(β-氨基酯)衍生物，构建针对肿瘤凋亡抑制基因Survivin的shRNA表达载体和肿瘤转移相关的Twist和Snail基因的siRNA，制备能同时包载化疗药物阿霉素（DOX）或紫杉醇（PTX）和基因药物的纳米粒，显著下调了人乳腺癌耐药细胞中凋亡抑制蛋白Survivin和小鼠乳腺癌4T1细胞中Twist和Snail表达水平，分别使耐药细胞中DOX和PTX的IC50值降低了83倍和62倍，抑制了4T1细胞的迁移能力，在荷耐药肿瘤动物体内抑瘤率达95%以上，在荷转移性肿瘤动物体内使肺转移灶数量减少90%以上，同时不造成明显的器官损伤。相关研究在国际重要学术刊物上发表高质量SCI论文17篇，包括国际权威期刊ACS Nano（1篇，IF：13.94），J Am Chem Soc（1篇，IF：13.86），Adv Funct Mater（1篇，IF：12.12）和Adv Drug Deliv Rev（1篇，IF：11.76）等。申请发明专利2项，获授权1项。共培养硕士生3人，博士生4人。该研究成果为RNAi靶向治疗肿瘤提供了新的思路和手段，对基于RNAi的肿瘤基因治疗早日实现从实验室到临床应用的跨越具有重要的参考价值。