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

A major challenge of gene therapy is the efficient transfer of genes to cell sites where effective transfection can occur. The impact of jet nebulization on DNA structural and functional integrity has been problematic for the aerosol delivery of genes to pulmonary sites and remains a serious concern for this otherwise promising and noninvasive approach.

This study examined effects of cationic liposome-DNA formulation on both transfection efficiency (in vitro and in vivo) and jet nebulizer stability. The effects of nebulization and sonication on liposome-DNA particle size characteristics were examined. Electron microscopy of promising formulations was performed using several fixation methods.

The cationic lipid bis-guanidinium-tren-cholesterol (BGTC), in combination with the neutral co-lipid dioleoylphosphatidylethanolamine (DOPE), was found to have a degree of stability adequate to permit effective gene delivery 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 correlating with the highest levels of transfection efficiency and nebulizer stability.

Optimizing gene delivery vectors for pulmonary aerosol delivery to respiratory sites must take into account factors other than transfection efficiency in vitro. Effects of liposome-DNA formulation on liposomal morphology (i.e. particle size, multilamellar structure) appear to be relevant to stability during aerosolization. These studies have allowed us to identify formulations that hold promise for successful clinical application of aerosol gene delivery.