EDAC-mediated O-acylisourea rearrangement for tertiary amine cationization of hyaluronic acid (HA) and its application as structural backbones in virus-inspired polyplexes

Cationic modification of hyaluronic acid (HA) is challenging due to its polyanionic nature, poor reactivity in water, and the instability of conventional coupling intermediates. This limits the development of HA-based components in non-viral gene delivery systems, which already suffer from amorphous morphology and mechanical fragility that reduce their transfection efficiency. Here, we reprogram a classically unfavorable EDAC-mediated rearrangement into a productive synthetic route, enabling direct cationization of hyaluronic acid (HA) through spontaneous O-acylisourea rearrangement. This water-based, catalyst-free process achieves up to 70 % substitution of HA's carboxyl groups-introducing cationic tertiary amine functionalities in water. The resulting aminated-hyaluronic acid (HAA) scaffolds act as rigid structural backbones in virus-inspired polymer-DNA nanoparticles termed as "Skeletoplexes", with enhanced stability and performance. When incorporated into polyplexes formed from diverse cationic systems-including poly(β-amino esters) and commercial vectors such as BrPERfect, Xfect, jetPEI, and Lipofectamine3000-HAA scaffolds improved <i>in vitro</i> transfection efficiency by up to 4-fold and <i>in vivo</i> gene expression by approximately 2-fold. These results establish a generalizable and green scaffold-based strategy that bridges the structural and functional gap between viral and non-viral gene delivery vectors.