title: Maintenance of transfection rates and physical characterization of lipid/DNA complexes after freeze-drying and rehydration.
abstract: It is well established that cationic liposomes form complexes with DNA and effectively transfect cells in vivo and ex vivo. Lipid/DNA complexes have proven safe and nonimmunogenic in clinical trials; however, they are known to aggregate readily in liquid formulations. This physical instability requires clinicians to prepare lipid/DNA complexes immediately prior to injection. In order to eliminate problems associated with this temporal requirement, we investigated the feasibility of preserving complexes as a dried preparation that could be tested, stored, and rehydrated as needed. To this end, our study evaluated the ability of different stabilizers to preserve transfection rates of complexes during acute freeze-drying stress. Our data show that complexes lyophilized in 0.5 M sucrose or trehalose possessed transfection rates similar to those of fresh preparations. In addition, dried complexes that exhibited full transfection activity upon rehydration had sizes comparable to nonlyophilized controls. Our work demonstrates that lipid/DNA complexes can be stabilized as dried powders that offer significant advantages over current liquid formulations. Furthermore, the correlation of transfection rates with maintenance of complex diameter suggests that size plays a critical role in lipid-based DNA delivery.
authors.affiliation: Department of Pharmaceutical Sciences, University of Colorado Health Sciences Center, Denver 80262, USA.
authors.forename: T J
authors.initials: TJ
authors.lastname: Anchordoquy
authors.forename: J F
authors.initials: JF
authors.lastname: Carpenter
authors.forename: D J
authors.initials: DJ
authors.lastname: Kroll
mesh_terms: D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D004247:DNA; D004337:Drug Carriers; D005229:Fatty Acids, Monounsaturated; D005612:Freeze Drying; D005947:Glucose; D007700:Kinetics; D008027:Light; D008081:Liposomes; D010714:Phosphatidylethanolamines; D000644:Quaternary Ammonium Compounds; D011994:Recombinant Proteins / Q000096:biosynthesis; D012542:Scattering, Radiation; D013395:Sucrose; D014162:Transfection / Q000379:methods* / Q000502:physiology*; D014199:Trehalose; D001616:beta-Galactosidase / Q000096:biosynthesis*
chemical_list: D004337:Drug Carriers; D005229:Fatty Acids, Monounsaturated; D008081:Liposomes; D010714:Phosphatidylethanolamines; D000644:Quaternary Ammonium Compounds; D011994:Recombinant Proteins; D013395:Sucrose; C020888:1,2-dielaidoylphosphatidylethanolamine; D004247:DNA; D014199:Trehalose; D001616:beta-Galactosidase; D005947:Glucose; C070046:1,2-dioleoyloxy-3-(trimethylammonium)propane