A class of cationic poly(alkylamidoamine)s (PAAAs) containing lipophilic methylene linkers were designed and examined as in vitro plasmid DNA (pDNA) delivery agents. The PAAAs were synthesized via step-growth polymerization between a diamine monomer and each of four different diacid chloride monomers with varying methylene linker lengths, including glutaryl chloride, adipoyl chloride, pimeloyl chloride, and suberoyl chloride, which served to systematically increase the lipophilicity of the polymers. The synthesized polymers successfully complexed with pDNA in reduced serum medium at N/P ratios of 5 and greater, resulting in polyplexes with hydrodynamic diameters of approximately 1 μm. These polyplexes were tested for in vitro transgene expression and cytotoxicity using HDFa (human dermal fibroblast), HeLa (human cervical carcinoma), HMEC (human mammary epithelial), and HUVEC (human umbilical vein endothelial) cells. Interestingly, select PAAA polyplex formulations were found to be more effective than Lipofectamine 2000 at promoting transgene expression (GFP) while maintaining comparable or higher cell viability. Transgene expression was highest in HeLa cells (∼90% for most formulations) and lowest in HDFa cells (up to ∼20%) as measured by GFP fluorescence. In addition, the cytotoxicity of PAAA polyplex formulations was significantly increased as the molecular weight, N/P ratio, and methylene linker length were increased. The PAAA vehicles developed herein provide a new delivery vehicle design strategy of displaying attributes of both polycations and lipids, which show promise as a tunable scaffold for refining the structure–activity–toxicity profiles for future genome editing studies.

中文翻译：

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亲脂性聚阳离子载体显示高质粒DNA传递到多种细胞类型

设计了一类含有亲脂性亚甲基接头的阳离子聚（烷基酰胺基胺）（PAAA），并作为体外质粒DNA（pDNA）传递试剂进行了检查。通过在二胺单体与四种不同的具有不同亚甲基接头长度的不同二酰氯单体（包括戊二酰氯，己二酰氯，庚二酰氯和辛二酰氯）之间逐步生长聚合来合成PAAA，它们可用来系统地提高其脂溶性聚合物。合成的聚合物成功地与pDNA在降低的血清培养基中以5或更高的N / P比率复合，从而形成流体动力学直径约为1μm的多链体。使用HDFa（人类皮肤成纤维细胞），HeLa（人类宫颈癌），HMEC（人类乳腺上皮）和HUVEC（人类脐静脉内皮）细胞。有趣的是，发现选择的PAAA复合物配方在促进转基因表达（GFP）的同时比Lipofectamine 2000更有效，同时保持了相当或更高的细胞活力。通过GFP荧光检测，转基因表达在HeLa细胞中最高（对于大多数制剂约为90％），而在HDFa细胞中最低（高达20％）。另外，随着分子量，N / P比和亚甲基接头长度的增加，PAAA多聚体制剂的细胞毒性显着增加。本文开发的PAAA载具提供了一种新的载具设计策略，可显示聚阳离子和脂质的属性，