Dalal Rishad J, Kumar Ramya, Ohnsorg Monica, Brown Mary, Reineke Theresa M
Department of Chemistry, University of Minnesota, Minneapolis, Minnesota 55455, United States.
University Imaging Centers, University of Minnesota, Minneapolis, Minnesota 55455, United States.
ACS Macro Lett. 2021 Jul 20;10(7):886-893. doi: 10.1021/acsmacrolett.1c00335. Epub 2021 Jun 25.
Cationic polymer vehicles have emerged as promising platforms for nucleic acid delivery because of their scalability, biocompatibility, and chemical versatility. Advancements in synthetic polymer chemistry allow us to precisely tune chemical functionality with various macromolecular architectures to increase the efficacy of nonviral-based gene delivery. Herein, we demonstrate the first cationic bottlebrush polymer-mediated pDNA delivery by comparing unimolecular, synthetically defined bottlebrush polymers to their linear building blocks. We successfully synthesized poly(2-(dimethylamino)ethyl methacrylate) (pDMAEMA) bottlebrushes through ring-opening metathesis polymerization to afford four bottlebrush polymers with systematic increases in backbone degree of polymerization ( = 13, 20, 26, and 37), while keeping the side-chain degree of polymerization constant ( = 57). Physical and chemical properties were characterized, and subsequently, the toxicity and delivery efficiency of pDNA into HEK293 cells were evaluated. The bottlebrush-pDNA complex (bottleplex) with the highest , BB_37, displayed up to a 60-fold increase in %EGFP+ cells in comparison to linear macromonomer. Additionally, we observed a trend of increasing EGFP expression with increasing polymer molecular weight. Bottleplexes and polyplexes both displayed high pDNA internalization as measured via payload enumeration per cell; however, quantitative confocal analysis revealed that bottlebrushes were able to shuttle pDNA into and around the nucleus more successfully than pDNA delivered via linear analogues. Overall, a canonical cationic monomer, such as DMAEMA, synthesized in the form of cationic bottlebrush polymers proved to be far more efficient in functional pDNA delivery and expression than linear pDMAEMA. This work underscores the importance of architectural modifications and the potential of bottlebrushes to serve as effective biomacromolecule delivery vehicles.
阳离子聚合物载体因其可扩展性、生物相容性和化学多功能性，已成为核酸递送的有前景的平台。合成聚合物化学的进展使我们能够通过各种大分子结构精确调节化学功能，以提高基于非病毒的基因递送的功效。在此，我们通过将单分子、合成定义的瓶刷聚合物与其线性构建块进行比较，展示了首个阳离子瓶刷聚合物介导的pDNA递送。我们通过开环易位聚合成功合成了聚（甲基丙烯酸2-（二甲氨基）乙酯）（pDMAEMA）瓶刷，得到了四种瓶刷聚合物，主链聚合度系统增加（ = 13、20、26和37），同时保持侧链聚合度恒定（ = 57）。对其物理和化学性质进行了表征，随后评估了pDNA进入HEK293细胞的毒性和递送效率。与线性大分子单体相比，具有最高 的瓶刷-pDNA复合物（瓶刷复合物）BB_37显示%EGFP+细胞增加了60倍。此外，我们观察到随着聚合物分子量增加，EGFP表达有增加的趋势。通过每个细胞的有效载荷计数测量，瓶刷复合物和多聚复合物都显示出高pDNA内化；然而，定量共聚焦分析表明，瓶刷比通过线性类似物递送的pDNA更能成功地将pDNA转运到细胞核内和细胞核周围。总体而言，以阳离子瓶刷聚合物形式合成的典型阳离子单体，如DMAEMA，在功能性pDNA递送和表达方面比线性pDMAEMA高效得多。这项工作强调了结构修饰的重要性以及瓶刷作为有效生物大分子递送载体的潜力。
