Formulation and in vitro transfection efficiency of poly (D, L-lactideco-glycolide) microspheres containing plasmid DNA for gene delivery

Abstract

The stability, in vitro release, and in vitro cell transfection efficiency of plasmid DNA (pDNA) poly (D,L-lactide-co-glycolide) (PLGA) microsphere formulations were investigated. PLGA microspheres containing free and polylysine (PLL)-complexed pDNA were prepared by a water-oil-water solvent extraction/evaporation technique. Encapsulation enhanced the retention of the supercoiled structure of pDNA as determined by gel electrophoresis. PLL complexation of pDNA prior to encapsulation increased both the stability of the supercoiled form and the encapsulation efficiency. Free pDNA was completely degraded after exposure to DNase, while encapsulation protected the pDNA from enzymatic degradation. Rapid initial in vitro release of pDNA was obtained from microspheres containing free pDNA, while the release from microspheres containing PLL-complexed pDNA was sustained for more than 42 days. Bioactivity of encapsulated pDNA determined by in vitro cell transfection using Chinese hamster ovary cells (CHO) showed that the bioactivity of encapsulated pDNA was retained in both formulations but to a greater extent with PLL-complexed pDNA microspheres. These results demonstrated that PLGA microspheres could be used to formulate a controlledrelease delivery system for pDNA that can protect the pDNA from DNase degradation without loss of functional activity.

Formulating Poly(Lactide-Co-Glycolide) Particles for Plasmid DNA Delivery

“…21 This results in PLL complexed release of DNA and delayed second phase release. 74 A higher percentage of supercoiled pDNA and protection against endonuclease activity of DNAse 1 is observed for the PLLcomplexed pDNA compared to uncomplexed pDNA. 21 The release of pDNA-PLL complex from PLGA particles is also dependent on the stabilizer type and concentration and the highest release is found using 7% PVA.…”

Innovative strategies for co-delivering antigens and CpG oligonucleotides

“…Other cationic microparticles that can potentially be used to deliver CpG ODN and antigens include biodegradable microparticles that have been functionalized with cetyltrimethylammoniumbromide (CTAB) [78], cetyldimethylethylammonium bromide (CDAB), dimethyl dioctadecyl ammonium bromide (DDAB) [79], 1,2-dioleoyl-1,3-trimethylammoniopropane (DOTAP), cationic DDAB [79], poly(L-lysine) (PLL) [80-83], polyamidoamine (PAMAM) dendrimers [28], polyethylenimine [72, 84-95] and chitosan [96]. The advantage of these cationic microparticles is that they also have the potential to provide sequential release of antigen and CpG ODN, which could be used to enhance the immune response further [28].…”

Poly(D,L‐lactide‐co‐glycolide acid) nanoparticles for DNA delivery: Waiving preparation complexity and increasing efficiency

“…10,11,[15][16][17] However, many variations of this preparation protocol are being used by different groups of researchers resulting with different NPs characteristics in terms of size, yield, loading efficiency, release kinetics, and protein expression levels. 10,[18][19][20][21][22] A particularly important parameter which differs from one study to another and which may affect the efficiency of NPs is sonication. In these studies different operation times and amplitudes of sonication are employed which set the energy load entering the emulsification system.…”