RESUMEN
We have developed new, synthetic vector formulations that display high efficiency of gene transfer to vascular cells and tissues. The formulations comprise cationic liposomes and cationic, receptor-targeting peptides that self assemble on mixing with plasmid DNA into receptor-targeted nanocomplexes (RTNs). One such RTN formulation was optimal for transfection of primary smooth muscle cells (LYD-1), while a second was optimal for transfection of rabbit aortic explants (LYD-2). In both RTNs, the peptide was a 16-lysine motif linked to the targeting sequence CYGLPHKFCG via a short spacer sequence. The major difference between LYD-1 and LYD-2 lay in the cationic lipid component, where LYD-1 contained ditetradecyl trimethyl ammonium (DTDTMA), an unsaturated, cationic lipid with a 14-carbon alkyl tail, whereas LYD-2 contained 2,3-dioleyloxypropyl-1-trimethyl ammonium chloride (DOTMA), a cationic lipid with an 18-carbon unsaturated alkyl tail. LYD-2 transfections of aortic explants were effective with incubations performed at room temperature for as little as 30 minutes, with either saline or glucose-based solutions. Transgene expression in the explants peaked at 5 days and persisted for 14 days. The kinetics of transfected gene expression, along with the efficacy of transfection with short incubation times, indicate that these new formulations may be useful tools in the development of molecular therapies for cardiovascular diseases.
Asunto(s)
Aorta/citología , Músculo Liso Vascular/metabolismo , Nanopartículas/química , Receptores de Superficie Celular/genética , Animales , Células Cultivadas , Técnicas de Transferencia de Gen , Vectores Genéticos/genética , Humanos , Cinética , Liposomas/química , Masculino , Músculo Liso Vascular/citología , Técnicas de Cultivo de Órganos , Péptidos/química , Plásmidos/química , Plásmidos/genética , Reacción en Cadena de la Polimerasa , Conejos , Porcinos , Transfección/métodosRESUMEN
A range of monocationic and dicationic dioxyalkylglycerol cytofectins have been synthesised possessing methylene and short n-ethylene glycol spacers. The monocationic compounds were found to be effective in transfections when formulated as lipopolyplexes with peptide and DNA components, in particular with shorter PEG head groups which may have less effect on peptide targeting in the ternary complex.