Star-shaped copolypeptide-mediated transfection of 3D organoids

While gene therapy offers hope for treating genetic disorders and cancer, efficient and targeted gene delivery continues to be a significant hurdle. This study introduces star-shaped block copolypeptides as novel gene carriers capable of directly transfecting entire cell clusters within three-dimensional (3D) organoid models. Star-shaped poly(L-lysine)-block-poly(L-alanine) (s-PLL-PLA) polypeptides with various arm numbers were synthesized and their potentiality as non-viral gene carriers was evaluated. Our experimental results revealed that the transfection efficiency of s-PLL-PLA/plasmid polyplexes rivaled that of the commercial standard, Lipofectamine 2000, also highlighting the importance of using 3D organoids to mimic the structure and function of native tissues. The s-PLL-PLA polypeptides exhibited amphiphilicity and steric hindrance imposed by the presence of rigid, hydrophobic PLA segment on the star architecture, rendering the improved biocompatibility and transfection efficiency due to the charge shielding and less dense packing with the plasmids. By targeting HSP90 AB1, an oncogene associated with aggressive cancer progression and poor patient outcomes, the s-PLL-PLA/short hairpin RNA (shRNA) polyplexes exhibited potent anticancer efficacy via the effective suppression of MDA-MB-231 breast cancer cell proliferation and then the induction of apoptosis. These findings strongly suggest that s-PLL-PLA polypeptides hold promise as effective non-viral gene delivery systems for inducing cancer cell apoptosis.