Efficient intracellular siRNA delivery by ethyleneimine-modified amphiphilic macromolecules.
Efficient intracellular siRNA delivery by ethyleneimine-modified amphiphilic macromolecules.
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DOI：
10.1002/mabi.201100064
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发表时间：
2011-09-09
影响因子：
4.6
通讯作者：
Uhrich, Kathryn E.
中科院分区：
文献类型：
--
作者：
Sparks, Sarah M.;Waite, Carolyn L.;Harmon, Alexander M.;Nusblat, Leora M.;Roth, Charles M.;Uhrich, Kathryn E.
为了实现治疗性基因沉默的前景，迫切需要能够结合和递送寡核苷酸（如短干扰RNA（siRNA））而无毒性的新材料。两亲性大分子（AM）用线性乙烯亚胺官能化以产生能够与siRNA络合的阳离子AM。在结构上，母体AM由粘酸骨架形成，其四羟基被12碳脂肪链烷基化以形成大分子的疏水组分。然后将该烷基化粘酸用作为亲水组分的聚（乙二醇）（PEG）单官能化。所得AM在疏水结构域内含有游离羧酸。在这项工作中，线性亚乙基亚胺与游离羧酸缀合以产生具有一个伯胺（1 N）或一个伯胺和四个仲胺（5 N）的AM。此外，合成了具有胺取代的AM，其既取代疏水结构域中的游离羧酸，又取代相邻的PEG，以产生具有一个伯胺和八个仲胺（9 N）的聚合物，四个仲胺位于AM疏水结构域的每一侧。所有胺官能化AM形成纳米胶束，但只有5 N和9 N AM具有阳离子zeta电位，其随着胺数目的增加而增加。在10 µM及以上浓度下，所有AM的固有细胞毒性均低于线性聚乙烯亚胺（L-PEI）。通过增加阳离子乙烯亚胺链的长度和胺的总数，在恶性胶质瘤细胞系中实现了成功的siRNA络合和细胞siRNA递送。此外，使用siRNA与9 NAM的复合物观察到萤火虫荧光素酶的siRNA诱导的沉默，并且与L-PEI相当，但在较高浓度（高于10 μM）下显示出更好的细胞活力。这项工作突出了阳离子AM作为siRNA递送的安全有效的合成载体的前景。具体地，鉴定了一种新型聚合物（9 N）用于有效地将siRNA递送至癌细胞，并将进一步评估。
New materials that can bind and deliver oligonucleotides such as short interfering RNA (siRNA) without toxicity are greatly needed to fulfill the promise of therapeutic gene silencing. Amphiphilic macromolecules (AMs) were functionalized with linear ethyleneimines to create cationic AMs capable of complexing with siRNA. Structurally, the parent AM is formed from a mucic acid backbone whose tetra-hydroxy groups are alkylated with 12-carbon aliphatic chains to form the hydrophobic component of the macromolecule. This alkylated mucic acid is then mono-functionalized with poly(ethylene glycol) (PEG) as a hydrophilic component. The resulting AM contains a free carboxylic acid within the hydrophobic domain. In this work, linear ethyleneimines were conjugated to the free carboxylic acid to produce an AM with one primary amine (1N) or one primary amine and four secondary amines (5N). Further, an AM with amine substitution both to the free carboxylic acid in the hydrophobic domain and also to the adjacent PEG was synthesized to produce a polymer with one primary amine and eight secondary amines (9N), four located on each side of the AM hydrophobic domain. All amine-functionalized AMs formed nanoscale micelles but only the 5N and 9N AMs had cationic zeta potentials, which increased with increasing number of amines. All AMs exhibited less inherent cytotoxicity than linear polyethyleneimine (L-PEI) at concentrations of 10 µM and above. By increasing the length of the cationic ethyleneimine chain and the total number of amines, successful siRNA complexation and cellular siRNA delivery was achieved in a malignant glioma cell line. In addition, siRNA-induced silencing of firefly luciferase was observed using complexes of siRNA with the 9N AM and comparable to L-PEI, yet showed better cell viability at higher concentrations (above 10 µM). This work highlights the promise of cationic AMs as safe and efficient synthetic vectors for siRNA delivery. Specifically, a novel polymer (9N) was identified for efficient siRNA delivery to cancer cells and will be further evaluated.
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