title: Comparative study of poly (lactic-co-glycolic acid)-poly ethyleneimine-plasmid DNA microparticles prepared using double emulsion methods.
abstract: Controlled release of plasmid DNA (pDNA) from biodegradable poly lactic-co-glycolic acid (PLGA) microparticles has the potential to enhance transgene expression. However, barriers to this approach include limited encapsulation efficiency, pDNA damage during fabrication and confinement of the microparticles inside phagolysosomal compartments. Combining PLGA with poly ethyleneimine (PEI) can improve protection of pDNA during fabrication, increase encapsulation efficiencies and impart the PLGA microparticles with the capacity to escape the phagolysosomal compartments. This study compares three promising formulation methods for preparing PLGA PEI pDNA microparticles and evaluates for buffering capacity, cellular uptake, transfection efficiency and toxicity. In the first method, PLGA PEI pDNA microparticles are prepared by entrapping pDNA in blended PLGA/PEI using the double emulsion water-in-oil-in-water solvent evaporation technique (PA). In a second approach, PEI-pDNA polyplexes are prepared and then entrapped in PLGA microparticles using a double emulsion solvent evaporation method (PB). Microparticles prepared using formulation methods PA and PB are then compared against PLGA microparticles with PEI conjugated to the surface using carbodiimide chemistry (PC); 0.5% PVA is identified as the optimum concentration of surfactant for generating the strongest transfection efficiencies. N:P ratios of 5 and 10 are selected for preparation of each group. Gel electrophoresis demonstrates that all PLGA microparticle formulations have strong pDNA binding capacity. An MTT assay shows that in vitro cytotoxicity of PLGA PEI microparticles is significantly lower than PEI alone. PLGA PEI pDNA microparticles mediate higher cellular uptake efficiency and consequently higher transgene expression than unmodified PLGA microparticles in COS7 and HEK293 cells. Preparing PEI-pDNA polyplexes prior to entrapment in PLGA microparticles (PB) results in the highest pDNA loading. This is 2.5-fold higher than pDNA loading in unmodified PLGA microparticles. PLGA PEI pDNA microparticles prepared using method PB generates the strongest transfection efficiencies, which are 500-fold higher than unmodified PLGA pDNA microparticles in HEK293 cells and 1800-fold higher in COS-7 cells. The highest transfection efficiencies generated from microparticles prepared using method PB is achieved using an N:P ratio of 5.
authors.affiliation: Division of Pharmaceutics, College of Pharmacy, University of Iowa, Iowa City, IA 52242, USA.
authors.forename: Xue-Qing
authors.initials: XQ
authors.lastname: Zhang
authors.forename: Janjira
authors.initials: J
authors.lastname: Intra
authors.forename: Aliasger K
authors.initials: AK
authors.lastname: Salem
mesh_terms: D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D020099:Coated Materials, Biocompatible / Q000737:chemistry*; D004247:DNA / Q000008:administration & dosage* / Q000235:genetics; D016503:Drug Delivery Systems; D004655:Emulsions / Q000737:chemistry; D019344:Lactic Acid / Q000737:chemistry*; D008863:Microspheres; D010316:Particle Size; D010957:Plasmids / Q000737:chemistry* / Q000235:genetics; D011091:Polyesters; D011094:Polyethyleneimine / Q000737:chemistry*; D011100:Polyglycolic Acid; D011108:Polymers / Q000737:chemistry*; D012997:Solvents / Q000737:chemistry
chemical_list: D020099:Coated Materials, Biocompatible; D004655:Emulsions; D011091:Polyesters; D011108:Polymers; D012997:Solvents; D011100:Polyglycolic Acid; D019344:Lactic Acid; C033616:poly(lactide); D011094:Polyethyleneimine; D004247:DNA