PEPTIDE-MEDIATED GENE DELIVERY - INFLUENCE OF PEPTIDE STRUCTURE ON GENE-EXPRESSION

Cationic peptides possessing a single cysteine, tryptophan, and lysine repeat were synthesized to define the minimal peptide length needed t o mediate transient gene expression in mammalian cells. The N-terminal cysteine in each peptide was either alkylated or oxidatively dimerize d to produce peptides possessing lysine chains of 3, 6, 8, 13, 16, 18, 26, and 36 residues. Each synthetic peptide was studied for its abili ty to condense plasmid DNA and compared to polylysine(19) and cationic lipids to establish relative in vitro gene transfer efficiency in Hep G2 and COS 7 cells. Peptides with lysine repeats of 13 or more bound D NA tightly and produced condensates that decreased in mean diameter fr om 231 to 53 nm as lysine chain length increased. In contrast, peptide s possessing 8 or fewer lysine residues were similar to polylysine(19) , which bound DNA weakly and produced large (0.7-3 mu m) DNA condensat es. The luciferase expression was elevated 1000-fold after HepG2 cells were transfected with DNA condensates prepared with alkylated Cys-Trp -Lys(18) (AlkCWK(18)) versus polylysine(19). The gene transfer efficie ncies of AlkCWK(19) and cationic lipids were equivalent in HepG2 cells but different by 10-fold in COS 7 cells. A 40-fold reduction in parti cle size and a 1000-fold amplification in transfection efficiency for AlkCWK(18) DNA condensates relative to polylysine(19) DNA condensates suggest a contribution from tryptophan that leads to enhanced gene tra nsfer properties for AlkCWK(18). Tryptophan-containing cationic peptid es result in the formation of small DNA condensates that mediate effic ient nonspecific gene transfer in mammalian cells. Due to their low to xicity, these peptides may find utility as carriers for nonspecific ge ne delivery or may be developed further as low molecular weight DNA co ndensing agents used in targeted gene delivery systems.