A novel N-acyl phosphatidylethanolamine-containing delivery vehicle for spermine-condensed plasmid DNA

A unique method for formulation of plasmid DNA with phospholipids has been devised for the purpose of producing vehicles that can mediate gene deliver y and transfection of living cells. The polycation, spermine, was used to c ondense plasmid DNA within a water-in-chloroform emulsion stabilized by pho spholipids. After organic solvent removal, the particles formed could be ex truded to a number average size of about 200 nm and retained DNA that was p rotected from nuclease digestion. This resulted in a relatively high protec ted DNA-to-lipid ratio of approximately 1 mu g DNA/mu mol lipid. The size d istribution of the preparation was relatively homogeneous as judged by ligh t microscopy and quasi-elastic light scattering. Electron microscopic studi es showed structural heterogeneity, but suggested that at least some of the plasmid DNA in this preparation was in the form of the previously observed spermine-condensed bent rods and toroids and was encapsulated within lipos omal membranes. Preparations with the fusogenic phospholipid composition, 1 ,2-dioleoyl-sn-glycero-3-phosphoethanolamine-N-dodecanoyl/1,2-dioleoyl-sn-g lycero-3-phosphocholine showed transfection activity for several cells line s, particularly OVCAR-3 cells. The transfection activity sedimented with th e lipid during centrifugation, confirming the association of active plasmid DNA with phospholipids. Transfection efficiency in culture was found to be of the same order of magnitude as cationic lipoplexes but much less toxic to the cells. Significant transfection of OVCAR-3 cells in tissue culture c ould also be observed, even in the presence of the intraperitoneal fluid fr om a mouse with an OVCAR-3 ascites tumor. These data indicate a new type of liposomal gene delivery system devoid of cationic lipids, phosphatidyletha nolamine, cationic polymers and viral components.