Development of novel poly(ethylene glycol)-based vehicles for gene delivery

The purpose of this research was to develop and characterize a gene delivery vehicle with a poly(ethylene glycol) (PEG) backbone with the aim of overcoming limitations, such as cytotoxicity and rapid clearance, associated with current commonly used non-viral carriers. PEG was functionalized with DNA-binding peptides (DBPs) to make a vehicle (DBP-PEG) capable of condensing DNA. Complexes of plasmid DNA and DBP-PEG were formed and characterized by measuring particle size, zeta potential, and transfection efficiency as a function of N:P charge ratios (DBP-PEG amino groups:DNA phosphate). Dynamic light scattering showed that DBP-PEG was able to condense DNA efficiently resulting in a population of particles in the range of 250-300 nm. Neutral or slightly positive zeta potentials were measured for charge ratios of 3.5:1 and greater. DBP-PEG/DNA complexes, made with plasmids encoding the green fluorescent protein (GFP) and beta-Galactosidase (beta-Gal) genes, were used to transfect Chinese hamster ovary (CHO) cells. DBP-PEG/DNA was capable of transfecting cells and maximum transfection efficiency was observed for N:P ratios from 4:1 to 5:1, corresponding to zeta potentials from -4 to +1.6 mV. The effect of the DBP-PEG vehicle on cell viability was assayed. DBP-PEG was associated with a higher percentage of viable cells ( approximately 95%) than either polyethylenimine (PEI) or poly-L-lysine (PLL), and with transfection efficiency greater than PLL, but with somewhat lower than PEI. The results of this work demonstrate that PEG can be used as the backbone for gene delivery vehicles.