Imidazolium‐Based Lipid Analogs as Lipofection Reagents

Three imidazolium-based lipid analogs with varying degrees of saturation were synthesized and evaluated for their potential to enhance gene transfer in the 1321N1 and HEK-293FT cell lines. The analogs were incorporated into DPPC/DOPE-based liposomes and compared with two established transfection reagents, Lipofectamine3000 and FuGENE 4 K. Cytotoxicity assessments, as determined by lactate dehydrogenase assays, revealed a lower toxicity profile for the fully saturated compound 1 compared with its mono- (2) and di-unsaturated (3) counterparts. Notably, longer incubation times (24 h) amplified cytotoxic responses, with C₁₇H₃₁-IMeNMe₃ (3) displaying the highest cell damage. Despite elevated toxicity relative to commercial reagents, compound 3 successfully delivered plasmid DNA using a PiggyBac transposon system, producing stable transfected cell lines after extended culture periods. C₁₅H₃₁-IMeNMe₃ (1) achieved stable transfection but required longer colony expansion than the commercial controls, whereas C₁₇H₃₃-IMeNMe₃ (2) did not yield transfected lines under the conditions tested. Overall, these results suggest that structural modifications, particularly the length of the alkyl chain and the number of double bonds, impact both cytotoxicity and transfection efficiency. The findings underscore the importance of rational lipid design, as stronger membrane interactions can enhance uptake while potentially heightening adverse effects. This study informs future development of safer, more effective nonviral delivery vectors.