New basic membrane-destabilizing peptides for plasmid-based gene delivery in vitro and in vivo.
We have designed new basic amphiphilic peptides, ppTG1 and ppTG20 (20 amino acids), and evaluated their efficiencies in vitro and in vivo as single-component gene transfer vectors. ppTG1 and ppTG20 bind to nucleic acids and destabilize liposomes consisting of 1-palmitoyl-2-oleoylphosphatidylcholine (POPC) and cholesterol (3:1 mol/mol) at pH 5 and pH 7. Complexes of plasmid DNA and ppTG1 gave rise to efficient transfection in a variety of human and murine cell lines at low charge ratios ([+/-] between 1 and 2). In cell culture experiments, such vectors were superior to the membrane-destabilizing peptide KALA. In comparison with cationic lipid-, dendrimer-, and polymer-based transfection agents like Superfect, polyethylenimine (PEI), and Lipofectin, ppTG1 vectors showed good transfection efficiencies, especially at low DNA doses. Moreover, we demonstrated for the first time successful gene transfer in living animals with a single-component peptide vector. In the mouse, intravenous injection of a luciferase expression plasmid complexed with ppTG1 or ppTG20 led to significant gene expression in the lung 24 hours after injection. Structure-function studies with ppTG1, ppTG20, and sequence variants suggest that the high gene transfer activity of these peptides is correlated with their propensity to exist in alpha-helical conformation, which seems to be strongly influenced by the nature of the hydrophobic amino acids.
Rittner
Karola
K
TRANSGENE S.A., 11 rue de Molsheim, F-67000 Strasbourg, France. rittner@transgene.fr
Benavente
Annie
A
Bompard-Sorlet
Albine
A
Heitz
Frédéric
F
Divita
Gilles
G
Brasseur
Robert
R
Jacobs
Eric
E
D000595:Amino Acid Sequence; D000818:Animals; D000900:Anti-Bacterial Agents; D004268:DNA-Binding Proteins / Q000138:chemical synthesis* / Q000737:chemistry; D018014:Gene Transfer Techniques; D006367:HeLa Cells; D006801:Humans; D008081:Liposomes; D018942:Macrolides; D051379:Mice; D009842:Oligopeptides / Q000737:chemistry / Q000235:genetics*; D010455:Peptides / Q000737:chemistry / Q000235:genetics*; D010957:Plasmids / Q000235:genetics*; D014162:Transfection
D000900:Anti-Bacterial Agents; D004268:DNA-Binding Proteins; D008081:Liposomes; D018942:Macrolides; D009842:Oligopeptides; D010455:Peptides; C465384:ppTG1 peptide; C465383:ppTG20 peptide; C057620:bafilomycin A