Ros Samantha, Freitag Jessica S, Smith David M, Stöver Harald D H
Department of Chemistry and Chemical Biology, McMaster University, Hamilton, Ontario L8S 4L8, Canada.
Fraunhofer Institute for Cell Therapy and Immunology IZI, 04103 Leipzig, Saxony, Germany.
ACS Omega. 2020 Apr 16;5(16):9114-9122. doi: 10.1021/acsomega.9b03734. eCollection 2020 Apr 28.
Synthetic polycations are studied extensively as DNA delivery agents because of their ease of production, good chemical stability, and low cost relative to viral vectors. This report describes the synthesis of charge-shifting polycations based on ,-(dimethylamino)ethyl acrylate (DMAEA) and 3-aminopropylmethacryamide (APM), called PAD copolymers, and their use for DNA delivery into HeLa cells. PAD copolymers of varying compositions were prepared by RAFT polymerization to yield polymers of controlled molecular weights with low dispersities. Model hydrolysis studies were carried out to assess the rate of charge-shifting of the polycations by loss of the cationic dimethylaminoethanol side chains. They showed reduction in the net cationic charge by about 10-50% depending on composition after 2 days at pH 7, forming polyampholytes comprising permanent cationic groups, residual DMAEA, as well as anionic acrylic acid groups. HeLa cells exposed for 4 h to PAD copolymers with the greatest charge-shifting ability showed comparable or higher viability at high concentrations, relative to the noncharge shifting polycations PAPM and polyethyleneimine (PEI) 2 days post-exposure. Cell uptake efficiency of PAD/60bp-Cy3 DNA polyplexes at 2.5:1 N/P ratio was very high (>95%) for all compositions, exceeding the uptake efficiency of PEI polyplexes of equivalent composition. These results suggest that these PAD copolymers, and in particular PAD containing 80 mol % DMAEA, have suitable rates of charge-shifting hydrolysis for DNA delivery, as PAD showed reduced cytotoxicity at high concentrations, while still retaining high uptake efficiencies. In addition, the polyampholytes formed during DMAEA hydrolysis in PAD copolymers can offer enhanced long-term cytocompatibility.
合成聚阳离子作为DNA递送剂被广泛研究，因为它们易于生产、具有良好的化学稳定性且相对于病毒载体成本较低。本报告描述了基于丙烯酸2-（二甲基氨基）乙酯（DMAEA）和3-氨丙基甲基丙烯酰胺（APM）的电荷转移聚阳离子（称为PAD共聚物）的合成及其用于将DNA递送至HeLa细胞的用途。通过可逆加成-断裂链转移（RAFT）聚合制备了不同组成的PAD共聚物，以得到具有低分散度的分子量可控的聚合物。进行了模型水解研究，以通过阳离子二甲基氨基乙醇侧链的损失来评估聚阳离子的电荷转移速率。结果表明，在pH 7条件下放置2天后，根据组成不同，净阳离子电荷减少了约10%-50%，形成了包含永久性阳离子基团、残留DMAEA以及阴离子丙烯酸基团的两性聚电解质。暴露于具有最大电荷转移能力的PAD共聚物4小时的HeLa细胞，在暴露后2天，相对于非电荷转移聚阳离子聚（3-氨丙基甲基丙烯酰胺）（PAPM）和聚乙烯亚胺（PEI），在高浓度下显示出相当或更高的活力。对于所有组成，在2.5:1的氮磷比下，PAD/60bp-Cy3 DNA多聚体的细胞摄取效率非常高（>95%），超过了同等组成的PEI多聚体的摄取效率。这些结果表明，这些PAD共聚物，特别是含有80摩尔%DMAEA的PAD，具有适合DNA递送的电荷转移水解速率，因为PAD在高浓度下显示出降低的细胞毒性，同时仍保持高摄取效率。此外，PAD共聚物中DMAEA水解过程中形成的两性聚电解质可以提供增强的长期细胞相容性。
