Poly-L-lysine-graft-PEG-comb-type polycation copolymers for gene delivery

Polycations have been used for gene delivery in vitro quite successfully, however, in vivo applications suffered from serum effects that lower the overall gene drug efficiency. PEG polymers have been used extensively to minimize serum effects and create "stealth liposomes", biocompatible materials, and proteins with extended circulation. Here, we report our efforts towards creating "stealth polyplexes". A comb-type polycation, poly-L-lysine-graft-PEG copolymers were successfully prepared by ring opening of PEG-epoxide with E-amino lysine groups of linear poly-L-lysine. The ratios of PEG-epoxide to poly-L-lysine, PEG-epoxide size (2K, 3K, and 5B), and poly-L-lysine size (10K, 26K, and 38K) were varied. Copolymers with as little as 2% grafted PEG chains sterically stabilized DNA/copolymer complexes (polyplexes) even at charge neutrality. These polyplexes, formed with copolymers with various size of PEG chains grafted on various lenghths of poly-L-lysine backbone, remained relatively small, approximately 100 nm in saline With higher degree of grafting, the binding was significantly diminished. In addition, the morphology of polyplexes changed from thoroidal to more elongated, worm-like forms. Some globular structures were detected in cases of a lower degree of grafting. Finally, DNA release form polyplexes when exposed to negatively charged macromolecules, poly-L-aspartic acid sodium salt, is very structure dependant. Enhanced levels of luciferase expression observed with PLL-PEC polyplexes versus either free DNA or PLL polyplexes are encouraging and warrant further optimization of the polymeric gene delivery system.