Song Yanyan, Lou Bo, Zhao Peng, Lin Chao
The Institute for Biomedical Engineering and Nanoscience, Tongji University School of Medicine, Tongji University , Shanghai 200092, P. R. China.
Mol Pharm. 2014 Jul 7;11(7):2250-61. doi: 10.1021/mp4006672. Epub 2014 Jun 10.
A folate-decorated, disulfide-based cationic dextran conjugate having dextran as the main chain and disulfide-linked 1,4-bis(3-aminopropyl)piperazine (BAP) residues as the grafts was designed and successfully prepared as a multifunctional gene delivery vector for targeted gene delivery to ovarian cancer SKOV-3 cells in vitro and in vivo. Initially, a new bioreducible cationic polyamide (denoted as pSSBAP) was prepared by polycondensation reaction of bis(p-nitrophenyl)-3,3'-dithiodipropanoate, a disulfide-containing monomer, and BAP. It was found that the pSSBAP was highly efficient for in vitro gene delivery against MCF-7 and SKOV-3 cell lines. Subsequently, two cationic dextran conjugates with different amounts of BAP residues (denoted as Dex-SSBAP6 and Dex-SSBAP30, respectively) were synthesized by coupling BAP to disulfide-linked carboxylated dextran or coupling pSSBAP-oligomer to p-nitrophenyl carbonated dextran. Both two conjugates were able to bind DNA to form nanosized polyplexes with an improved colloidal stability in physiological conditions. The polyplexes, however, were rapidly dissociated to liberate DNA in a reducing environment. In vitro transfection experiments revealed that the polyplexes of Dex-SSBAP30 efficiently transfected SKOV-3 cells, yielding transfection efficiency that is comparable to that of linear polyethylenimine or lipofectamine 2000. AlamarBlue assay showed that the conjugates had low cytotoxicity in vitro at a high concentration of 100 mg/L. Further, Dex-SSBAP30 has primary amine side groups and thus allows for folate (FA) conjugation, yielding FA-coupled Dex-SSBAP30 (Dex-SSBAP30-FA). It was found that Dex-SSBAP30-FA was efficient for targeted gene delivery to SKOV-3 tumor xenografted in a nude mouse model by intravenous injection, inducing a higher level of gene expression in the tumor as compared to Dex-SSBAP30 lacking FA and comparable gene expression to linear polyethylenimine as one of the most efficient polymeric vectors for intravenous gene delivery in vivo. Disulfide-based cationic dextran system thus has a high potential for intravenous gene delivery toward cancer gene therapy.
设计并成功制备了一种以葡聚糖为主链、以二硫键连接的1,4 - 双（3 - 氨丙基）哌嗪（BAP）残基为接枝的叶酸修饰的基于二硫键的阳离子葡聚糖共轭物，作为一种多功能基因传递载体，用于在体外和体内将基因靶向传递至卵巢癌SKOV - 3细胞。首先，通过含二硫键的单体双（对硝基苯基）-3,3'-二硫代二丙酸酯与BAP的缩聚反应制备了一种新型的生物可还原阳离子聚酰胺（记为pSSBAP）。发现pSSBAP对MCF - 7和SKOV - 3细胞系的体外基因传递具有高效性。随后，通过将BAP偶联到二硫键连接的羧化葡聚糖上，或将pSSBAP - 低聚物偶联到对硝基苯基碳酸酯葡聚糖上，合成了两种具有不同数量BAP残基的阳离子葡聚糖共轭物（分别记为Dex - SSBAP6和Dex - SSBAP30）。这两种共轭物都能够结合DNA，在生理条件下形成具有改善的胶体稳定性的纳米级多聚体。然而，这些多聚体在还原环境中会迅速解离以释放DNA。体外转染实验表明，Dex - SSBAP30的多聚体能够有效地转染SKOV - 3细胞，产生的转染效率与线性聚乙烯亚胺或脂质体2000相当。AlamarBlue检测表明，在100 mg/L的高浓度下，共轭物在体外具有低细胞毒性。此外，Dex - SSBAP30具有伯胺侧基，因此可以进行叶酸（FA）偶联，得到FA偶联的Dex - SSBAP30（Dex - SSBAP30 - FA）。发现通过静脉注射，Dex - SSBAP30 - FA对于将基因靶向传递至裸鼠模型中移植的SKOV - 3肿瘤是有效的，与缺乏FA的Dex - SSBAP30相比，在肿瘤中诱导了更高水平的基因表达，并且与线性聚乙烯亚胺（作为体内静脉基因传递最有效的聚合物载体之一）具有相当的基因表达。因此，基于二硫键的阳离子葡聚糖系统在癌症基因治疗的静脉基因传递方面具有很高的潜力。
