title: Characterization of liposome-mediated gene delivery: expression, stability and pharmacokinetics of plasmid DNA.
abstract: We have characterized a new synthetic gene delivery system, termed DLS, which may be suitable for systemic gene therapy. DLS constitutes a lipopolyamine and a neutral lipid and associated plasmid DNA in the formation of lamellar vesicles (DLS-DNA). The ratio of lipids and lipid to DNA as well as the method of preparation were optimized to yield a high in vitro transfection efficiency compared with that previously reported for cationic lipid systems. DLS-DNA showed a rapid cellular uptake and distribution in the cytoplasmic and nuclear (especially in the nucleoli) compartments as determined by laser-assisted confocal microscopy. There was little or no plasmid DNA degradation over a period of 20 min, relatively slow plasma clearance, and effective and rapid cellular uptake of DLS-DNA following intravenous administration in mice. Supercoiled plasmid DNA could be detected in blood cells up to 1 h after injection. Systemic administration of DLS-DNA yielded transgene expression in mouse tissues, such as in lung or liver. The ratio of DLS:DNA and the procedure used to form DLS-DNA affected both the level and cellular specificity of expression of a luciferase reporter gene showing that in vitro transfection efficiency of DLS-DNA formulations cannot be easily extrapolated to an in vivo setting. Optimization of the formulation of a DNA delivery system was critical to obtain a defined structure resulting in a preparation with high reproducibility and stability, greater homogeneity of particle size and high efficacy following systemic gene transfer. In addition, the DLS system may be formulated for specific target tissues and may have a wide range of applications for gene therapy.
authors.affiliation: Laboratory of Tumor Cell Biology, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.
authors.forename: A R
authors.initials: AR
authors.lastname: Thierry
authors.forename: P
authors.initials: P
authors.lastname: Rabinovich
authors.forename: B
authors.initials: B
authors.lastname: Peng
authors.forename: L C
authors.initials: LC
authors.lastname: Mahan
authors.forename: J L
authors.initials: JL
authors.lastname: Bryant
authors.forename: R C
authors.initials: RC
authors.lastname: Gallo
mesh_terms: D000818:Animals; D004247:DNA / Q000032:analysis / Q000235:genetics / Q000493:pharmacokinetics; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D005998:Glycine / Q000031:analogs & derivatives; D006367:HeLa Cells; D006801:Humans; D017403:In Situ Hybridization; D008081:Liposomes; D008156:Luciferases / Q000235:genetics; D051379:Mice; D008807:Mice, Inbred BALB C; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D010714:Phosphatidylethanolamines; D010957:Plasmids; D013096:Spermine / Q000031:analogs & derivatives; D014162:Transfection; D019076:Transgenes / Q000235:genetics; D001616:beta-Galactosidase / Q000235:genetics
chemical_list: D008081:Liposomes; D010714:Phosphatidylethanolamines; C061134:dioctadecylamidoglycylspermine; D013096:Spermine; C020888:1,2-dielaidoylphosphatidylethanolamine; D004247:DNA; D008156:Luciferases; D001616:beta-Galactosidase; D005998:Glycine