Institute of Polymer Science & Technology, Madrid, Spain.
Eur J Pharm Biopharm. 2012 Nov;82(3):465-74. doi: 10.1016/j.ejpb.2012.08.002. Epub 2012 Aug 23.
Nonviral methods for gene delivery are becoming ever more prevalent along with the need to design new vectors that are highly effective, stable in biological fluids, inexpensive, and facile to produce. Here, we synthesize our previously reported monomer N-ethyl pyrrolidine methacrylamide (EPA) and evaluate its effectiveness in gene vector applications when copolymerized with 1-vinylimidazole (VI). A range of these novel linear cationic copolymers were synthesized via free radical polymerization with low molecular weights (oligomers) and low polydispersities showing two pK(a) values as the two co-monomers are cationic. DNA-polymer polyplexes had average sizes between 100 and 250nm and zeta-potentials between 10 and 25mV, and a strong dependence of composition on the size on the zeta-potential was observed. The cytotoxicity of the homopolymers, oligomers, and polyplexes toward human fibroblasts and 3T3 mouse fibroblasts was evaluated using the MTT and AlamarBlue™ assays, proving that formulations could be made with toxicity as low as low molecular weight linear poly (dimethylaminoethyl methacrylate) (PDMAEMA). The transfection capability of the polyplexes measured using the G-luciferase marker gene far superseded PDMAEMA when evaluated in biological conditions. Furthermore, blood compatibility studies showed that these new oligomers exhibit no significant hemolysis or platelet activation above PBS controls. These new EPA based oligomers with low toxicity and ease of scalability show high transfection abilities in serum conditions, and blood compatibility showing its potential for systemic gene delivery applications.
随着设计高效、在生物流体中稳定、廉价且易于生产的新型载体的需求不断增加，非病毒基因传递方法变得越来越流行。在这里，我们合成了之前报道的单体 N-乙基吡咯烷甲基丙烯酰胺（EPA），并评估了其与 1-乙烯基咪唑（VI）共聚时在基因载体应用中的有效性。通过自由基聚合合成了一系列新型线性阳离子共聚物，分子量低（低聚物），多分散性低，显示出两个 pK(a) 值，因为两个共聚单体都是阳离子的。DNA-聚合物聚集体的平均粒径在 100nm 到 250nm 之间，zeta 电位在 10mV 到 25mV 之间，并且观察到组成与粒径和 zeta 电位之间存在很强的依赖性。使用 MTT 和 AlamarBlue™测定法评估了这些同聚物、低聚物和聚集体对人成纤维细胞和 3T3 鼠成纤维细胞的细胞毒性，证明可以用低毒性制成低分子量线性聚（二甲氨基乙基甲基丙烯酰胺）（PDMAEMA）。使用 G-荧光素酶标记基因测量的聚集体的转染能力在生物条件下远远超过了 PDMAEMA。此外，血液相容性研究表明，这些新的低聚物在 PBS 对照之上没有明显的溶血或血小板活化。这些新的基于 EPA 的低聚物具有低毒性和易于规模化的特点，在血清条件下具有高转染能力，并且血液相容性显示出其在系统基因传递应用中的潜力。
