Gene transfer by guanidinium-cholesterol: Dioleoylphosphatidyl-ethanolamine liposome-DNA complexes in aerosol

Background A major challenge of gene therapy is the efficient transfer of g enes to cell sites where effective transfection can occur. The impact of je t nebulization on DNA structural and functional integrity has been problema tic for the aerosol delivery of genes to pulmonary sites and remains a seri ous concern for this otherwise promising and noninvasive approach. Methods This study examined effects of cationic liposome-DNA formulation on both transfection efficiency (in vitro and in vivo) and jet nebulizer stab ility. The effects of nebulization and sonication on liposome-DNA particle size characteristics were examined. Electron microscopy of promising formul ations was performed using several fixation methods. Results The cationic lipid bis-guanidinium-tren-cholesterol (BGTC), in comb ination with the neutral co-lipid dioleoylphosphatidylethanolamine (DOPE), was found to have a degree of stability adequate to permit effective gene d elivery by the aerosol route. Optimal ratios of lipids and plasmid DNA were identified. Particle size analysis and ultrastructural studies revealed a remarkably homogeneous population of distinctly liposomal structures correl ating with the highest levels of transfection efficiency and nebulizer stab ility. Conclusions Optimizing gene delivery vectors for pulmonary aerosol delivery to respiratory sites must take into account factors other than transfectio n efficiency in vitro. Effects of liposome-DNA formulation on liposomal mor phology (i.e. particle size, multilamellar structure) appear to be relevant to stability during aerosolization. These studies have allowed us to ident ify formulations that hold promise for successful clinical application of a erosol gene delivery. Copyright (C) 1999 John Wiley & Sons, Ltd.