LGA-PEI nanoparticles enable efficient nucleic acid delivery to primary murine T cells

Non-viral engineering of primary T cells is constrained by low transfection efficiency and significant cargo-induced toxicity. This study evaluates a LGA-PEI (lactic-co-glycolic acid-modified polyethyleneimine) nanoparticle system designed to overcome the recalcitrance of murine T cells, particularly unstimulated, murine T cells. We characterized LGA-PEI polyplexes for size, surface charge, and pH-dependent stability, noting rapid disassembly at pH 5.0 to facilitate endosomal escape. Optimization of nanoparticle concentrations enabled transfection efficiencies of 16.5 % in CD3-sensitized T cells and 30.0 % in stimulated primary mouse T cells, maintaining high viability (85 %). Mechanistic analysis suggests that cell death is primarily driven by initial membrane disruption rather than post-internalization metabolic stress. We identified a direct correlation between the number of internalized plasmids and GFP expression, confirming efficient post-internalization events. LGA-PEI-mediated delivery of shRNA targeting Sorcin achieved significant gene knockdown. Transfection with LGA-PEI-encapsulated scrambled control plasmids (shRNA) exhibited increased CD25 and FoxP3 marker expression, along with cytokine production alterations. These findings demonstrate that the LGA-PEI platform provides a high-efficiency, low-toxicity alternative for T cell modulation, though the observed phenotypic alterations necessitate careful immunotoxicity screening in therapeutic applications.