Cationic charge regulation in nanogel networks for enhanced siRNA delivery

Cationic nanogels offer distinct advantages over conventional linear or branched polymeric vectors for siRNA delivery, particularly in terms of structural controllability and serum tolerance. However, the potential of modulating their internal charge motifs to further boost efficacy remains underexplored. Here, we engineer a PEGylated nanogel with a cationic network incorporating both permanently charged quaternary ammonium and pH-dependent protonatable tertiary amine groups, via copolymerization of (vinylbenzyl)trimethylammonium chloride (VBTMA, "T") and N-(4-vinylbenzyl)-N,N-dimethylamine (VBDMA, "D") with a reduction-responsive cross-linker N,N'-bis(acryloyl)cystamine (BAC). The rational combination of these monomers integrates distinct charge characteristics and pH-buffering capacity, leading to optimized siRNA loading, enhanced cellular uptake, improved endosomal escape, and reduced cytotoxicity. The PEG shell is designed to suppress protein corona formation in serum-containing environments, while the redox-sensitive crosslinker enables rapid nanogel dissociation and siRNA release upon exposure to intracellular glutathione (GSH). The optimally formulated nanogel, designated PEG₁₁₃-T₂₅D₇₅ (comprising 25% VBTMA and 75% VBDMA), exhibits superior delivery and transfection performance across a broad spectrum of cell lines. This study validates charge modulation as a viable strategy to simultaneously enhance delivery efficiency and mitigate cytotoxicity in nanogel-based vectors, thereby offering instructive design principles for the development of advanced polymeric vectors.