The effect of pegylation on the transfection activity of two homologous cationic cholesteryl cytofectins

Abstract

Polyethylene glycol is being used increasingly to improve circulation times and enhance bioavailability of therapeutic molecules and nano sized macromolecular assemblies. Here, two homologous cationic cholesteryl cytofectins 3β[N-(N’,N’-dimethylaminoethane)-carbamoyl] cholesterol (DC-Chol) and 3β[N- (N’,N’-dimethylaminopropane)-carbamoyl] cholesterol (Chol-T) with 2 and 3 carbon spacer elements, respectively, have been formulated into liposomes with near equimolar amounts of dioleoylphosphatidyl ethanolamine (DOPE) and 2 and 5% polyethylene glycol₂₀₀₀ (PEG₂₀₀₀). Pegylated liposomes (80 - 150 nm diameter) formed electrostatic complexes with plasmid DNA in the 1.5:1 – 2.5:1 range of liposome (positive): DNA (negative) charge ratio, which afforded protection to the DNA cargo against serum nuclease digestion. Plasmid pGL3-containing pegylated lipoplexes were only weakly cytotoxic in the human embryonic kidney cell line HEK 293. Gene transfer experiments in this cell line confirmed that the homologue with the 3 carbon spacer Chol-T, is associated with higher transgene activity, a trend which has been previously observed in unpegylated lipoplexes. Furthermore, only a 15% drop in transfection activity was recorded in increase of pegylation level from 2 to 5 mole percent.

Key words: Cationic cytofectin, gene transfer, polyethylene glycol, HEK 293.