Brush-like branched biodegradable polyesters, part III. Protein release from microspheres of poly(vinyl alcohol)-graft-poly(D,L-lactic-co-glycolic acid).

Brush-like branched polyesters, obtained by grafting poly(lactic-co-glycolic acid), PLGA, onto water-soluble poly(vinyl alcohol) (PVAL) backbones, were investigated regarding their utility for the microencapsulation of proteins. Poly(vinyl alcohol)-graft-poly(lactic-co-glycolic acid), PVAL-g-PLGA, offers additional degrees of freedom to manipulate properties such as e.g. molecular weight, glass transition temperature and hydrophilicity. PLGA chain length was varied at a constant molecular weight (M(w)) of the PVAL backbone and secondly M(w) of the PVAL backbone was varied keeping the PLGA chain lengths constant. The most striking feature of these polymers is their high M(w). Microencapsulation of hydrophilic macromolecules, such as bovine serum albumin, ovalbumin, cytochrome c and FITC-dextran using a w/o/w double emulsion technique was investigated. Surface morphology, particle size, encapsulation efficiencies and protein release profiles were characterized as well. Microencapsulation of model compounds was feasible at temperatures of 0-4 degrees C with yields typically in the range of 60-85% and encapsulation efficiencies of 70-90%. Both, encapsulation efficiency and initial protein release (drug burst) were strongly affected by the glass transition temperature, T(g), of the polymer in contact with water, whereas the in vitro protein release profile depended on the PVAL-g-PLGA structure and composition. In contrast to PLGA, protein release patterns were mostly continuous with lower initial drug bursts. Shorter PLGA chains increased drug release in the erosion phase, whereas initial pore diffusion was affected by the M(w) of PVAL backbone. Release profiles from 2 to 12 weeks could be attained by modification of composition and molecular weight of PVAL-g-PLGA and merit further investigations under in vivo conditions. The in vitro cytotoxicity of PVAL-g-PLGA is comparable to PLGA and therefore, this new class of biodegradable polyesters has considerable potential for parenteral drug delivery systems.     

Cisplatin delivery from poly(acrylic acid-co-methyl methacrylate) microparticles.

To develop a platform for tumor chemotherapy, poly(acrylic acid-co-methyl methacrylate) microparticles have been synthesized. Carboxylate containing monomers were included to complex therapeutic agents, specifically cisplatin. Microparticles were prepared by free radical emulsion polymerization in aqueous media. Particle diameter, zeta-potential, in vitro cytotoxicity, and in vivo acute toxicity were characterized for both cisplatin-loaded microparticles and unloaded microparticles. In vitro cytotoxicity and FT-IR were used to characterize cisplatin released from cisplatin-loaded microparticles. Acrylic acid feed mole fraction determined several key microparticle properties, including particle size, zeta-potential, and yield. A burst release of cisplatin (40%) in the first day was followed by a zero-order release phase. The interaction between cisplatin and microparticles allowed approximately 20% additional cisplatin release in the next five days. Cisplatin-loaded and unloaded microparticles are non-toxic (LC50>15 mM) to the cell line used in in vitro tests. Cisplatin released from cisplatin-loaded microparticles retained activity, but that activity was slightly lower than freshly prepared cisplatin. Other than a slight reduction in cisplatin activity, microparticles exhibited low in vivo acute toxicity (LD50>170 mg/kg), which suggests that this hydrogel particulate system and the hydrogel complexation mechanism should further be studied for drug delivery.     

Mucoadhesive ocular insert based on thiolated poly(acrylic acid): development and in vivo evaluation in humans.

The aim of the study was to develop a mucoadhesive ocular insert for the controlled delivery of ophthalmic drugs and to evaluate its efficacy in vivo. The inserts tested were based either on unmodified or thiolated poly(acrylic acid). Water uptake and swelling behavior of the inserts as well as the drug release rates of the model drugs fluorescein and two diclofenac salts with different solubility properties were evaluated in vitro. Fluorescein was used as fluorescent tracer to study the drug release from the insert in humans. The mean fluorescein concentration in the cornea/tearfilm compartment as a function of time was determined after application of aqueous eye drops and inserts composed of unmodified and of thiolated poly(acrylic acid). The acceptability of the inserts by the volunteers was also evaluated. Inserts based on thiolated poly(acrylic acid) were not soluble and had good cohesive properties. A controlled release was achieved for the incorporated model drugs. The in vivo study showed that inserts based on thiolated poly(acrylic acid) provide a fluorescein concentration on the eye surface for more than 8 h, whereas the fluorescein concentration rapidly decreased after application of aqueous eye drops or inserts based on unmodified poly(acrylic acid). Moreover, these inserts were well accepted by the volunteers. The present study indicates that ocular inserts based on thiolated poly(acrylic acid) are promising new solid devices for ocular drug delivery.     

Pancreatin enhanced erosion of and macromolecule release from 2,2-bis(2-oxazoline)-linked poly(epsilon-caprolactone).

The degradation and erosion of solvent cast films and injection molded bars prepared from poly(epsilon-caprolactone) (PCL) and 2,2'-bis(2-oxazoline) linked poly(epsilon-caprolactone) (PCL-O) were evaluated in simulated gastric fluid (SGF) (pH 1.2, pepsin present) and in simulated intestinal fluid (SIF) (pH 7.5, pancreatin present). After incubation of the polymer films (10 mg) and bars (70 mg) in the medium, the resulting decrease in molecular weight (degradation) was determined by size exclusion chromatography and the weight loss of the preparations was measured. In addition, the effect of pancreatin on FITC-dextran (MW 4400) release from PCL and PCL-O microparticles, prepared by w/o/w double emulsion technique, was studied. No degradation or weight loss was observed for either PCL or PCL-O films in SGF (12 h incubation, 37 degrees C). When compared to PBS pH 7.4, pancreatin hardly enhanced the weight loss of PCL films and bars. In contrast, pancreatin enhanced substantially erosion of PCL-O films and bars. Unlike PCL preparations, the PCL-O preparations showed surface erosion in SIF. Pancreatin increased considerably FITC-dextran release from both PCL and PCL-O microparticles. In conclusion, the present results demonstrate the enzyme sensitivity of the novel PCL-O polymer. In addition, the results show that pancreatin present in intestinal fluid may substantially affect drug release from PCL based preparations.     

Micelles of methoxy poly(ethylene oxide)-b-poly(epsilon-caprolactone) as vehicles for the solubilization and controlled delivery of cyclosporine A.

The commercial formulation of Cyclosporine A (CsA) for intravenous administration contains Cremophor EL, a low molecular weight surfactant known to be toxic. In this study, micelles of methoxy poly(ethylene oxide)-b-poly(epsilon-caprolactone) (PEO-b-PCL) were investigated as alternative vehicles for the solubilization and delivery of CsA. PEO-b-PCL block copolymers having identical PEO chain lengths and PCL molecular weights of 5000, 13,000, or 24,000 g mol(-)(1) were synthesized and assembled into polymeric micelles using a co-solvent evaporation method. PEO-b-PCL micelles were then compared to Cremophor EL micelles for their functional properties in drug delivery including micellar size, thermodynamic stability, core viscosity, CsA encapsulation, and in vitro CsA release. Among different PCL block lengths, optimum solubilization was achieved by utilizing polymeric micelles having a PCL block of 13,000 g mol(-)(1). CsA reached an aqueous solubility of 1.3 mg/mL in the presence of PEO-b-PCL micelles. This concentration is comparable to injectable CsA levels in its Cremophor EL formulation (0.5-2.5 mg/mL). In contrast to the Cremophor EL formulation, the in vitro rate of CsA release was significantly sustained by the polymeric micellar carrier. Within 12 h, only 5.8% of CsA was released from polymeric micelles while Cremophor EL micelles released 77% of their drug content. Accordingly, viscosity of the PEO-b-PCL micellar core was found to be significantly higher than Cremophor EL micelles. The results points to a potential for PEO-b-PCL micelles as nanoscopic drug carriers for efficient solubilization and controlled delivery of CsA.     

Camptothecin derivative-loaded poly(caprolactone-co-lactide)-b-PEG-b-poly(caprolactone-co-lactide) nanoparticles and their biodistribution in mice.

Triblock copolymers of poly(caprolactone-co-lactide)-b-PEG-b-poly(caprolactone-co-lactide) (PCLLA-PEG-PCLLA) were synthesized by ring opening copolymerization of caprolactone and lactide in the presence of poly(ethylene glycol) (PEG). With such triblock copolymers, PCLLA-PEG-PCLLA nanoparticles entrapping 10-hydroxycamptothecin-10,20-diisobutyl dicarbonate (HCPT-1), a derivative of the antitumor drug 10-hydroxycamptothecin (HCPT), were prepared by nano-precipitation method and characterized by dynamic light scattering (DLS), transmission electron microscopy (TEM) and atomic force microscopy (AFM). The investigations on drug loading, in vitro release and body distribution in mice after intravenous (i.v.) administration were also carried out. It is found that the obtained nanoparticles showed smooth surface and spherical shape with the controllable size in the range of 70-180 nm, and drug loading content varied from 3.3% to 7.0% depending on the copolymer composition and preparation conditions. The in vitro release behavior exhibited a sustaining release manner and was affected by particle size as well as copolymer composition. The results of body distribution study in mice show that the blood concentration of HCPT-1 could be maintained for a long period and the tissue distribution was influenced by the particle size to some extent. These results suggest that the PCLLA-PEG-PCLLA nanoparticles seem to be a promising delivery system for poorly soluble antitumor drugs or their derivatives.     

In vitro and in vivo release of levonorgestrel from poly(ortho esters): II. Crosslinked polymers

The in vitro and in vivo release of levonorgestrel from crosslinked poly(ortho ester) cylindrical devices containing 30 wt.% drug and 7.1 wt.% Mg(OH)₂ was studied. For reasons not entirely understood, hydrolysis rates of crosslinked poly(ortho ester) are faster than hydrolysis of linear polymers. The in vitro release rate of levonorgestrel was about 10 μg/day for 160 days at which point the experiment was discontinued. The in vivo levonorgestrel release rate in rabbits from identical devices was about 20 μg/day. In the in vitro studies polymer erosion leads drug release but in vivo release of levonorgestrel is fast enough so that concomitant polymer erosion and drug release take place. SEM examination of devices explanted from rabbits are consistent with a surface erosion process.     

乳酸-羟基乙酸共聚物载药纳米粒的制备及体外释药性

背景:医用纳米粒作为药物传递的新型载体,目前已经成为医药领域研究的重点。目的:构建以生物可降解材料乳酸-羟基乙酸共聚物为载体,负载抗肿瘤药物5-氟尿嘧啶的载药纳米粒。方法:利用复乳-溶剂挥发法制备乳酸-羟基乙酸共聚物载药纳米粒。场发射扫描电子显微镜观察纳米粒表面形态;激光粒度分析仪测定粒径分布并计算成球率;紫外分光光度计测定5-氟尿嘧啶载药量、包封率,并对体外释药进行评估。结果与结论:纳米粒呈球性,平均粒径为(186±14)nm,成球率、载药量和包封率分别为70.8%、6.6%、28.1%,体外释药有突释现象,24h内5-氟尿嘧啶累积释药量达36.2%,10d达83.6%。提示成功制备乳酸-羟基乙酸共聚物载药纳米粒,其具有缓释效应。     

Folate-conjugated methoxy poly(ethylene glycol)/poly(epsilon-caprolactone) amphiphilic block copolymeric micelles for tumor-targeted drug delivery.

Amphiphilic block copolymers composed of methoxy poly(ethylene glycol) (MPEG) and poly(epsilon-caprolactone) (PCL) were synthesized and then conjugated with folic acid to produce a folate-receptor-targeted drug carrier for tumor-specific drug delivery. Folate-conjugated MPEG/PCL micelles containing the anticancer drug paclitaxel were prepared by micelle formation in aqueous medium. The size of the folate-conjugated MPEG/PCL micelles formed was about 50-130 nm, depending on the molecular weight of block copolymers, and was maintained at less than 150 nm even after loading with paclitaxel. The in vitro release profile of the paclitaxel from the MPEG/PCL micelles exhibited no initial burst release and showed sustained release. Paclitaxel-loaded folate-conjugated MPEG/PCL micelles (PFOL50) exhibited much higher cytotoxicity for cancer cells, such as MCF-7 and HeLa cells, than MPEG/PCL micelles without the folate group (PMEP50). Confocal image analysis revealed that fluorescent paclitaxel-loaded PFOL50 micelles were endocytosed into MCF-7 cells through the interaction with overexpressed folate receptors on the surface of the cancer cells.     

Comparison of poly(acryl starch) and poly(lactide-co-glycolide) microspheres as drug delivery system for a rotavirus vaccine.

Drug delivery systems allowing controlled release of antigen are of particular interest in the development of vaccines. We have compared poly(acrylic starch) microspheres (PAS) and poly(lactide-co-glycolide) microspheres (PLG) as drug delivery systems for a rotavirus vaccine. The polymers are both biodegradable but have different degradation mechanisms and antigen release profiles. PAS are enzymatically degraded and have a continuous fast antigen release rate compared to the hydrolytically degraded PLG which release the incorporated antigen in a pulsatile manner. In this study mice were immunised intramuscularly and orally on three occasions with formalin-inactivated rotavirus (FRRV) incorporated in PAS and PLG and with FFRV alone. Serum and faeces samples were collected and analysed by ELISA for rotavirus specific IgG and IgA antibodies. A neutralising assay was also conducted on both serum and faeces antibodies. The two different polymer drug delivery systems induced different immune responses depending on administration route. PAS elicited significant antibody levels and neutralising effect after oral administration while PLG showed high antibody levels after intramuscular administration. The immune response appears to be dependent on the differences in antigen release and degradation mechanism for the two polymer systems.     

Degradation of and drug release from a novel 2,2-bis(2-oxazoline) linked poly(lactic acid) polymer.

The degradation rate of poly(lactic acid) (PLA) is typically modified by copolymerization of the glycolide with lactide. In the present study, the degradation rate of PDLLA was modified by a novel linking of PLA with 2,2'-bis(2-oxazoline). This modification resulted in formation of a more rapidly degrading poly(ester amide) (PEA) for controlled drug release. The hydrolytic degradation of PDLLA and PEA films was studied in PBS (pH 7.4, USP XXIV, 37 degrees C); the resulting decrease in molecular weight was determined by size exclusion chromatography and the weight loss of films was measured. Drug releases of guaifenesin (mw 198.2), timolol (mw 332.4), sodium salicylate (mw 160.1) and FITC-dextran (mw 4400) from PDLLA and PEA films and microspheres were examined in PBS (pH 7.4, 37 degrees C). The degradation rate of PEA was substantially greater than that of PDLLA. The release profiles of all small model drugs (mw <332.4) from PDLLA films were biphasic or triphasic, while the release profiles of small model drugs from PEA films varied extensively. Due to the faster weight loss of PEA, FITC-dextran (mw 4400) was released substantially more rapidly from PEA microspheres than from PDLLA microspheres. In conclusion, all model drugs, except guaifenesin, were released faster from PEA preparations than from PDLLA preparations.     

Anti-tumor drug delivery of pH-sensitive poly(ethylene glycol)-poly(L-histidine-)-poly(L-lactide) nanoparticles

pH-sensitive poly(ethylene glycol)-poly(l-histidine)-poly(L-lactide) (PEG-PH-PLLA) nanoparticles were prepared and used as carriers for anti-tumor drug delivery. The morphology and properties of the nanoparticles such as pH sensitivity, zeta potential and mean diameters were investigated. The cytotoxicity of PEG-PH-PLLA nanoparticles was evaluated. Doxorubicin (DOX) was encapsulated in the nanoparticles to explore the release profile. The drug-loaded nanoparticles were incubated with HepG2 cells to study the in vitro anti-tumor effect. The results showed the sizes of both blank nanoparticles and drug-loaded nanoparticles in pH 7.4 were smaller than those of nanoparticles in pH 5.0, and the mean diameter of drug-loaded nanoparticles was much bigger than that of blank nanoparticles. The PEG-PH-PLLA nanoparticles were nontoxic to both NIH 3T3 fibroblasts and HepG2 cells. The release profile showed that the release of DOX in pH 5.0 was much faster than that in pH 7.4. The in vitro experiments demonstrated that the anti-tumor effect of drug-loaded nanoparticles was preferable to free doxorubicin. The pH-sensitive PEG-PH-PLLA nanoparticles are promising carriers for anti-tumor drug delivery.     

Mucoadhesive drug carrier based on interpolymer complex of poly(vinyl pyrrolidone) and poly(acrylic acid) prepared by template polymerization.

To develop a new mucoadhesive drug carrier, poly(vinyl pyrrolidone) (PVP)/poly(acrylic acid) (PAA) interpolymer complexes were prepared by the template polymerization of acrylic acid using PVP as a template polymer. Fourier transform infrared results showed that the interpolymer complexes were formed by hydrogen bonds between the carboxyl groups of PAA and the carbonyl groups of PVP. The adhesive forces of the PVP/PAA interpolymer complexes were higher than that of commercial Carbopol 971. Moreover, the adhesive force and the release rate can be controlled by changing the mole ratios of PVP and PAA. The release rates of ketoprofen from the PVP/PAA interpolymer complexes showed pH-dependency, and were slower at lower pH. The release rate of ketoprofen from the complex seemed to be mainly controlled by the dissolution rate of the complex above a pK(a) of PAA (4.75) and by the diffusion rate below the pK(a). The prepared complex appears to be an adequate carrier for the mucoadhesive drug delivery system.     

Influence of formulation parameters on the characteristics of poly(D, L-lactide-co-glycolide) microspheres containing poly(L-lysine) complexed plasmid DNA.

This study describes the influence of polymer type, surfactant type/concentration, and target drug loading on the particle size, plasmid DNA (pDNA) structure, drug loading efficiency, in vitro release, and protection from DNase I degradation of poly(D, L-lactide-co-glycolide) (PLGA) microspheres containing poly(L-lysine) (PLL) complexed pDNA. PLGA microspheres containing pDNA-PLL were prepared using the water-in-oil-in-water (w-o-w) technique with poly(vinyl alcohol) (PVA) and poly(vinyl pyrrolidone) (PVP) as surfactants in the external aqueous phase. A complex ratio of 1:0.33 (pDNA-PLL, w/w) enhanced the stability of pDNA during microsphere preparation. Higher pDNA-PLL loading efficiency (46.2%) and supercoiled structure (64.9%) of pDNA were obtained from hydrophobic PLGA (M(w) 31000) microspheres compared with hydrophilic PLGA or low-molecular-weight PLGA microspheres. The particle size decreased from 6.6 to 2.2 microm when the concentration of PVA was increased from 1 to 7%. At the same concentration of surfactant, PVA stabilized microspheres showed higher pDNA-PLL loading efficiency (46.2%) than PVP stabilized microspheres (24.1%). Encapsulated pDNA in PLGA microspheres was protected from enzymatic degradation and maintained in the supercoiled form. The pDNA-PLL microspheres showed in vitro release of 95.9 and 84.9% within 38 days from the low-molecular-weight PLGA and hydrophilic PLGA microspheres, respectively, compared to 54.2% release from the hydrophobic, higher-molecular-weight PLGA microspheres. The results suggest loading and release of pDNA-PLL complex can be influenced by surfactant concentration and polymer type.     

Paclitaxel releasing films consisting of poly(vinyl alcohol)-graft-poly(lactide-co-glycolide) and their potential as biodegradable stent coatings.

Although substantial progress in catheter and stent design has contributed to the success of percutaneous transluminal angioplasty (PTA) of atherosclerotic disease, the incidence of restenosis caused by in-stent neointimal hyperplasia remains a serious problem. Therefore, stents with a non-degradable polymer coating showing controlled release of active ingredients have become an attractive option for the site-specific delivery of anti-restenotic agents. Biodegradable coatings using polyesters, namely poly(lactic-co-glycolic acid) (PLGA) and different poly(vinyl alcohol)-graft-poly(lactic-co-glycolic acid) (PVA-g-PLGA) as paclitaxel-eluting stent coating materials were investigated here to evaluate their influence on the release kinetic. Whereas PLGA showed sigmoid release behavior, the paclitaxel release from PVA-g-PLGA films was continuous over 40 days without initial drug burst. Wide angle X-ray diffraction confirmed that paclitaxel is dissolved in the polymer matrix. Paclitaxel crystallization can be observed at a drug load of > or =10%. The effect of drug loading on polymer degradation was studied in films prepared from PVA300-g-PLGA30 with paclitaxel loadings of 5% and 15% over a time period of 6 weeks. The results suggest a surface-like erosion mechanism in films. A model stent (Jostent peripheral) coated with Parylene N, a poly(p-xylylene) (PPX) derivate, was covered with a second layer of PVA300-g-PLGA15, and PVA300-g-PLGA30 by using airbrush method. Morphology of coated stents, and film integrity after expansion from 3.12 to 5 mm was investigated by scanning electron microscopy (SEM). The devices resisted mechanical stress during stent expansion and merit further investigation under in vivo conditions.     

Mucoadhesive drug carriers based on complexes of poly(acrylic acid) and PEGylated drugs having hydrolysable PEG-anhydride-drug linkages.

We have designed a new mucoadhesive drug delivery formulation based on H-bonded complexes of poly(acrylic acid) (PAA) or poly(methacrylic acid) (PMAA) with the poly(ethylene glycol) (PEG), of a (PEG)-drug conjugate. The PEGylated prodrugs are synthesized with degradable PEG-anhydride-drug bonds for eventual delivery of free drug from the formulation. In this work we have used indomethacin as the model drug which is PEGylated via anhydride bonds to the PEG. The complexes are designed first to dissociate as the formulation swells in contact with mucosal surfaces at pH 7.4, releasing PEG-indomethacin, which then hydrolyses to release free drug and free PEG. We found that as MW of PAA increases, the dissociation rate of the complex decreases, which results in decreased rate of release of the drug. On the other hand, the drug release from PEG-indomethacin alone and from solid mixture of PEG-indomethacin+PAA was much faster than that from the H-bonded complexes. Due to the differences in the thermal stability, PMAA complex exhibited slightly faster drug release than that of the PAA complex of comparable MW. These H-bonded complexes of degradable PEGylated drugs with bioadhesive polymers should be useful for mucosal drug delivery.     

Incorporation and release behavior of hydrophobic drug in functionalized poly(D,L-lactide)-block-poly(ethylene oxide) micelles.

The poly(ethylene oxide)-poly(lactide) (PEO-PLA) block copolymers containing a small quantity of carboxylic acid in the PLA block were synthesized. The microscopic characteristics of nanoparticles with carboxylic acid content in the copolymer were analyzed, and the effect of specific interactions between the copolymer and the model drug on the drug loading capacity and the release behavior were investigated systematically. The sizes of nanoparticles prepared by a dialysis method are within the range of 30-40 nm. The nanoparticles prepared from functionalized block copolymers have a very low critical micelle concentration (CMC) value as low as approximately 10(-3) mg/ml, which indicates a good stability of the nanoparticles in spite of the presence of carboxylic acid. The drug loading efficiency of nanoparticles dramatically increased when carboxylic acid content was increased in the block copolymer. This result may be attributed to the increase of interactions between the copolymer and the drug. The release rate of the drug was much slower from nanoparticles containing higher amounts of carboxylic acid in the copolymer, which might be associated with the enhanced interaction between the carboxylic group of copolymers and the drug. These experimental results suggest that the nanoparticles prepared from functionalized PEO-PLA block copolymers could be a good candidate for an injectable drug delivery carrier.     

In vitro and in vivo gene transfer with poly(amino acid) vesicles.

Non-viral gene delivery systems utilise either amine lipids or polyamines and although non-viral gene delivery systems are said to have a superior safety profile to viruses, the polyamines such as poly(L-lysine) are toxic when used without derivatisation and usually require specific receptor mediated uptake and/or endosomolytic agents to be effective. However, the conversion of poly(L-lysine) and poly(L-ornithine) polyamino acids into amphiphilic vesicle forming polymers reduces the toxicity of the polyamino acids and enables the resulting polyamino acid vesicles to deliver genes both in vitro and in vivo in the absence of receptor specific ligands and endosomolytic agents. The incorporation of a distearoylphosphatidylethanolamine poly(ethylene glycol)-galactosamine conjugate (with the galactosamine unit at the distal end of the poly(ethylene glycol) moiety) into the polyamino acid formulations improved in vitro gene transfer in the case of the amphiphilic poly(L-ornithine) (POP) although no in vivo targeting was detected with the galactosamine formulations. We conclude that the conversion of poly(L-lysine) and poly(L-ornithine) into amphiphilic colloid forming molecules reduces their toxicity, thus allowing these systems to be used for gene transfer in vivo. It is possible that this approach may be extended to other polyamines.     

Influence of formulation parameters on the characteristics of poly( -lactide-co-glycolide) microspheres containing poly( -lysine) complexed plasmid DNA

This study describes the influence of polymer type, surfactant type/concentration, and target drug loading on the particle size, plasmid DNA (pDNA) structure, drug loading efficiency, in vitro release, and protection from DNase I degradation of poly( -lactide-co-glycolide) (PLGA) microspheres containing poly( -lysine) (PLL) complexed pDNA. PLGA microspheres containing pDNA–PLL were prepared using the water-in-oil-in-water (w–o–w) technique with poly(vinyl alcohol) (PVA) and poly(vinyl pyrrolidone) (PVP) as surfactants in the external aqueous phase. A complex ratio of 1:0.33 (pDNA–PLL, w/w) enhanced the stability of pDNA during microsphere preparation. Higher pDNA–PLL loading efficiency (46.2%) and supercoiled structure (64.9%) of pDNA were obtained from hydrophobic PLGA (M_w 31 000) microspheres compared with hydrophilic PLGA or low-molecular-weight PLGA microspheres. The particle size decreased from 6.6 to 2.2 μm when the concentration of PVA was increased from 1 to 7%. At the same concentration of surfactant, PVA stabilized microspheres showed higher pDNA–PLL loading efficiency (46.2%) than PVP stabilized microspheres (24.1%). Encapsulated pDNA in PLGA microspheres was protected from enzymatic degradation and maintained in the supercoiled form. The pDNA–PLL microspheres showed in vitro release of 95.9 and 84.9% within 38 days from the low-molecular-weight PLGA and hydrophilic PLGA microspheres, respectively, compared to 54.2% release from the hydrophobic, higher-molecular-weight PLGA microspheres. The results suggest loading and release of pDNA–PLL complex can be influenced by surfactant concentration and polymer type.