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High functionalization of dendrimer/cyclodextrin conjugate as a novel siRNA carrier

树枝状聚合物/环糊精缀合物作为新型 siRNA 载体的高功能化

基本信息

批准号：
18590144
负责人：
ARIMA Hidetoshi
金额：
$ 2.61万
依托单位：
Kumamoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

项目摘要

We previously reported the potential use of starburst polyamidoamine dendrimer (dendrimer, generation 3, G3)conjugate with a-cyclodextrin (α-CDE) as a novel gene delivery carrier. In the present study, the potential of α-CDE and their sugar-appended α-CDEs as novel and cell-specific carriers of small interfering RNA(siRNA) and short hairpin RNA-expressing vector(shpDNA)was evaluated in vitro and in vivo. The RNAi effects of the siRNA complex with cc-CDE were preferable to those of the complexes with commercially-available siRNA carriers in NIH3T3 cells stably expressing pGL3 luciferase. In addition, the siRNA complex with a-CDE showed negligible cytotoxicity up to higher charge ratios. However, we found that a-CDE did not have the potential as shpDNA carriers because of low RNAi effect. When the solution containing the binary complex of siRNA with α-CDE was injected into tumor tissues of mice bearing Colon-26 cells stably expressing the GL3 luciferase gene, the sufficient RNAi effects were provided, although the complex of Lipofectamine^TM2000 with siRNA showed the off-target effect. Therefore, α-CDE was found to provide the sequence-specific gene silencing effect by the complexation with siRNA. On the other hand, we prepared mannosylated α-CDE(M-α-CDE)and lactosylated α-CDE(L-α-CDE)to delivery siRNA and shpDNA specifically to antigen-presenting cells and hepatocytes, respectively. However, we found that L-α-CDE was able to bind to asialoglycoprotein receptor expressing on hepatocytes, although M-a-CDE could not bind to mannose receptor expressing on antigen-presenting cells. Both L-α-CDE and M-α-CDE had negligible cytotoxicity. These results suggest that L-α-CDE has the potential for hepatocyte-specific siRNA and shpDNA carriers.

我们之前报道了星爆聚酰胺胺树枝状聚合物（树枝状聚合物，第3代，G3）与α-环糊精（α-CDE）缀合物作为新型基因递送载体的潜在用途。在本研究中，α-CDE 及其附加糖的 α-CDE 作为小干扰 RNA（siRNA）和短发夹 RNA 表达载体（shpDNA）的新型细胞特异性载体的潜力在体外和体内进行了评估。在稳定表达pGL3荧光素酶的NIH3T3细胞中，siRNA与cc-CDE复合物的RNAi效果优于与市售siRNA载体的复合物。此外，带有a-CDE的siRNA复合物在较高电荷比下表现出可忽略不计的细胞毒性。然而，我们发现a-CDE由于RNAi效应较低，不具备作为shpDNA载体的潜力。当将含有siRNA与α-CDE的二元复合物的溶液注射到携带稳定表达GL3荧光素酶基因的Colon-26细胞的小鼠肿瘤组织中时，提供了足够的RNAi效果，尽管Lipofectamine^TM2000与siRNA的复合物显示出脱靶效果。因此，发现α-CDE通过与siRNA复合提供序列特异性基因沉默效应。另一方面，我们制备了甘露糖基化α-CDE（M-α-CDE）和乳糖基化α-CDE（L-α-CDE），分别将siRNA和shpDNA特异性递送至抗原呈递细胞和肝细胞。然而，我们发现L-α-CDE能够与肝细胞上表达的脱唾液酸糖蛋白受体结合，尽管M-a-CDE不能与抗原呈递细胞上表达的甘露糖受体结合。 L-α-CDE 和 M-α-CDE 的细胞毒性均可忽略不计。这些结果表明L-α-CDE具有作为肝细胞特异性siRNA和shpDNA载体的潜力。