Efficient in vitro siRNA delivery and intramuscular gene silencing using PEG-modified PAMAM dendrimers.
Efficient in vitro siRNA delivery and intramuscular gene silencing using PEG-modified PAMAM dendrimers.
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DOI：
10.1021/mp3001364
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发表时间：
2012-06-04
影响因子：
4.9
通讯作者：
Qi R
中科院分区：
文献类型：
--
作者：
Tang Y;Li YB;Wang B;Lin RY;van Dongen M;Zurcher DM;Gu XY;Banaszak Holl MM;Liu G;Qi R
尽管siRNA技术已被广泛用作基因敲减的工具，但在实现有效递送和体内功效方面仍存在实质性挑战。具体地，靶基因在体内沉默的低效率是siRNA疗法的临床应用的关键限制步骤。聚酰胺-胺（PAMAM）树枝状大分子被广泛用作药物和基因递送的载体;然而，通过PAMAM树枝状大分子的体内siRNA递送仍有待仔细研究。在这项研究中，研究了G5和G6 PAMAM树枝状聚合物的有效性，其表面胺的8%与MPEG-5000缀合，用于siRNA体外递送和小鼠体内肌肉内递送。将PEG修饰的树枝状聚合物的结果与母体树枝状聚合物以及Lipofectamine 2000和INTERFERin进行比较。两种PEG修饰的树枝状聚合物保护siRNA不被RNase消化，并在原代血管平滑肌细胞（VSMC）和小鼠腹腔巨噬细胞中对FITC标记的siRNA产生高转染效率。PEG修饰的树枝状聚合物在体外实现了质粒（293 A细胞）和腺病毒介导的绿色荧光蛋白（GFP）表达（Cos 7细胞）的敲低，其效率与Lipofectamine 2000所示的相似。我们进一步证明，在体内，肌内输送的GFP-siRNA使用PEG修饰的树枝状大分子显着抑制GFP的表达在短暂的腺病毒感染的C57 BL/6小鼠和GFP转基因小鼠。
Although siRNA techniques have been broadly applied as a tool for gene knockdown, substantial challenges remain in achieving efficient delivery and in vivo efficacy. In particular, the low efficiency of target gene silencing in vivo is a critical limiting step to the clinical application of siRNA therapies. Poly(amidoamine) (PAMAM) dendrimers are widely used as carriers for drug and gene delivery; however, in vivo siRNA delivery by PAMAM dendrimers remains to be carefully investigated. In this study, the effectiveness of G5 and G6 PAMAM dendrimers with 8% of their surface amines conjugated to MPEG-5000 was studied for siRNA delivery in vitro and for intramuscular in vivo delivery in mice. The results from the PEG-modified dendrimers were compared to the parent dendrimers as well as Lipofectamine 2000 and INTERFERin. Both PEG-modifed dendrimers protect the siRNA from being digested by RNase and gave high transfection efficiency for FITC-labeled siRNA in the primary vascular smooth muscle cells (VSMC) and mouse peritoneal macrophages. The PEG-modified dendrimers achieved knockdown of both plasmid (293A cells) and adenovirus-mediated green fluorescence protein (GFP) expression (Cos7 cells) in vitro with efficiency similar to that shown for Lipofectamine 2000. We further demonstrated in vivo that intramuscular delivery of GFP-siRNA using PEG-modified dendrimer significantly suppressed GFP expression in both transiently adenovirus infected C57BL/6 mice and in GFP transgenic mice.
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