Polymeric nanoparticles containing conjugated phospholipase A2 for nonviral gene delivery.
Polyethylenimine (PEI) was conjugated to phospholipase A(2) (PLA(2)) in an effort to improve transfection efficiency. PLA(2) was conjugated to PEI using EDC as a coupling reagent. The activity of enzyme in the conjugate was measured. DNA condensation ability of the conjugate to polymer was determined. The resultant nanoparticles were characterized by dynamic and electrophoretic light scattering. Two reporter genes were used to evaluate transfection efficiency in human embryonic kidney (HEK293) and human hepatoma (HepG2) cell lines. Conjugate was shown to retain PLA(2) activity and its ability to condense plasmid DNA, resulting in nanoparticles of a similar size to native PEI. The results demonstrated at N/P ratios of 15 and 20 showed 13- and 8-fold increase in transfection efficiency, respectively, compared to the maximum transfection efficiency of PEI (N/P ratio of 5) in the whole range of N/P ratios tested, from 5 to 60 in HepG2 cells. Toxicity studies in HepG2 cells showed uncomplexed conjugate had similar toxicity as PEI, and when complexed with DNA the conjugate had a significantly reduced toxicity. The results clearly indicate the potential for this approach to improve efficiencies of nonviral gene delivery vectors.
Le
Huong T
HT
Department of Pharmaceutics, College of Pharmacy, University of Florida, Gainesville, Florida 32610, USA.
Rao
Gururaj A
GA
Hirko
Aaron C
AC
Hughes
Jeffrey A
JA
D002460:Cell Line; D002470:Cell Survival; D018014:Gene Transfer Techniques; D056945:Hep G2 Cells; D006801:Humans; D008962:Models, Theoretical; D053758:Nanoparticles / Q000737:chemistry*; D054467:Phospholipases A2 / Q000737:chemistry* / Q000378:metabolism; D011094:Polyethyleneimine / Q000737:chemistry*; D011108:Polymers / Q000138:chemical synthesis / Q000737:chemistry*; D014162:Transfection
D011108:Polymers; D011094:Polyethyleneimine; D054467:Phospholipases A2