In vitro and in vivo transfections using siRNA lipoplexes prepared by mixing siRNAs with a lipid-ethanol solution

Many methods, such as the dry thin film and ethanol injection methods, have been developed for the preparation of cationic liposomes for siRNA delivery; however, these methods often require special equipment. In this study, we evaluated a siRNA transfection method, in which siRNA/cationic liposome complexes (siRNA lipoplexes) can be prepared by simply mixing an siRNA solution with lipid-ethanol solution. We used dimethyldioctadecylammonium bromide, 1,2-dioleoyl-3-trimethylammonium-propane or 11-((1,3-bis(dodecanoyloxy)-2-((dodecanoyloxy)methyl)propan-2-yl)amino)-N,N,N-trimethyl-11-oxoundecan-1-aminium bromide as a cationic lipid, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine or cholesterol as a neutral helper lipid, and poly(ethylene glycol) cholesteryl ether as a dispersant to prepare the lipid-ethanol solution. A solution of phosphate-buffered saline containing siRNA was then quickly added to a small volume of the lipid-ethanol solution, and the mixture (siRNA lipoplexes) was directly transferred into the cells or injected into mice. siRNA lipoplexes showed high gene knockdown efficiency in MCF-7, Colon 26, and LLC cells regardless of the type of cationic lipids. Furthermore, these siRNA lipoplexes exhibited significant gene knockdown in the lungs of mice after systemic injection. These results suggest that our siRNA lipoplex preparation method has the potential to be widely applicable for in vitro and in vivo siRNA transfections.