Abstract：
Among non-viral gene delivery vectors, poly(β-amino ester)s (PAEs) are one of the most versatile candidates because of their wide monomer availability, high polymer flexibility, and superior gene transfection performance both in vitro and in vivo. Over two decades, PAEs have evolved from linear to highly branched structures, significantly enhancing gene delivery efficacy. Building on the proven efficient sets of monomers in highly branched PAEs (HPAEs), this work introduced a new class of cyclic PAEs (CPAEs) constructed via an A2 + B4 + C2 cyclization synthesis strategy and identified their markedly improved gene transfection capabilities in gene delivery applications. Two sets of cyclic PAEs (CPAEs) with rings of different sizes and topologies were obtained. Their chemical structures were confirmed via two-dimensional nuclear magnetic resonance and the photoluminescence phenomena, and their DNA delivery behaviours were investigated and compared with the HPAE counterparts. In vitro assessments demonstrated that the CPAEs with a macrocyclic architecture (MCPAEs), significantly enhanced DNA intracellular uptake and facilitated efficient gene expression while maintaining perfect biocompatibility. The top-performance MCPAEs have been further employed to deliver a plasmid coding dual single guide RNA-guided CRISPR-Cas9 machinery to delete COL7A1 exon 80 containing the c.6527dupC mutation. In recessive dystrophic epidermolysis bullosa (RDEB) patient-derived epidermal keratinocytes, MCPAEs facilitated the CRISPR plasmid delivery and achieved efficient targeted gene editing in multiple colonies.
摘要：
在非病毒基因递送载体中，聚(β-氨基酯)(PAEs)是最通用的候选物之一，因为它们具有广泛的单体可用性，高聚合物柔韧性，和优越的基因转染性能在体外和体内。二十多年来,PAEs已经从线性结构发展到高度支化结构，显著增强基因递送功效。在高度支化的PAEs（HPAEs）中证明有效的单体组的基础上，这项工作引入了通过A2+B4+C2环化合成策略构建的一类新的环状PAEs(CPAEs),并确定了它们在基因递送应用中显著提高的基因转染能力.获得了两组具有不同大小和拓扑结构的环的环状PAE（CPAE）。通过二维核磁共振和光致发光现象证实了它们的化学结构，和他们的DNA递送行为进行了调查，并与HPAE同行进行了比较。体外评估表明，具有大环结构的CPAE（MCPAE），显着增强DNA细胞内摄取并促进有效的基因表达，同时保持完美的生物相容性。性能最高的MCPAE已进一步用于递送编码双单指导RNA指导的CRISPR-Cas9机制的质粒，以删除包含c.6527dupC突变的COL7A1外显子80。在隐性营养不良性大疱性表皮松解症（RDEB）患者来源的表皮角质形成细胞中，MCPAE促进CRISPR质粒递送并在多个菌落中实现有效的靶向基因编辑。
