Poly(lactic-co-glycolic) Acid Microparticles Research Articles

Fabrication and Characterization of Antibody-Loaded Cationic Porous PLGA Microparticles for Sustained Antibody Release.

Poly lactic-co-glycolic acid (PLGA) microparticles have been formulated to allow the sustained release of numerous drugs, including antibodies. It is well-known that antibodies are susceptible to chemical and physical stress; therefore, it is necessary to be loaded on PLGA microparticles under mild conditions. In the present study, we constructed cationic porous PLGA microparticles that could be electrostatically adsorbed with infliximab as a model antibody. Cationic porous PLGA microparticles were prepared using the double emulsion method by adding polyethyleneimine and ammonium bicarbonate. After antibody loading, surface pores closure was achieved by mild heating. The size of the optimized formulation was approximately 5 μm, exhibiting a positive charge. The loaded antibody was gradually released from the formulation over 56 days. Based on a tumor necrosis factor (TNF)-α inhibition assay, the released infliximab maintained its pharmacological activity. Collectively, we successfully loaded antibodies into PLGA microparticles while maintaining activity and demonstrating long-acting properties.

Intramuscularly Administered PLGA Microparticles for Sustained Release of Rivastigmine: In Vitro, In Vivo and Histological Evaluation

Rivastigmine is an acetylcholinesterase (AchE) and butyrylcholinesterase (BchE) inhibitor drug approved by the US Food and Drug Administration (FDA) for the treatment of mild to moderate dementia of Alzheimer's type. However, its first-pass metabolism and gastrointestinal side effects negatively affect the tolerability and efficacy of oral therapy. These adverse effects could be avoided with the use of a sustained -release formulation as an intramuscular (IM) administration system. The objective of this work was to develop polylactic co-glycolic acid (PLGA) microparticles for the sustained release of rivastigmine and to evaluate its stability during storage, tissue tolerance, in vitro release, and in vivo pharmacokinetics after its IM administration. The microparticles were made by the solvent evaporation emulsion method. A series of formulation parameters (the type of polymer used, the amount of polymer used, the initial amount of rivastigmine, and the volume of PVA 0.1% w/v) were studied to achieve an encapsulation efficiency (EE) and a rivastigmine load of 54.8 ± 0.9% and 3.3 ± 0.1%, respectively. The microparticles, whose size was 56.1 ± 2.8 μm, had a spherical shape and a smooth surface. FT-IR analysis showed that there is no chemical interaction between rivastigmine and the polymer. PLGA microparticles maintain rivastigmine retained and stable under normal (5 ± 3 °C) and accelerated storage (25 ± 2 °C and 60 ± 5 % RH) conditions for at least 6 months. The microparticles behaved as a sustained release system both in vitro and in vivo compared to non-encapsulated rivastigmine. The IM administration of the formulation in rats did not produce significant tissue damage. However, it is necessary to reproduce the experiments with multiple doses to rule out a negative effect in terms of tolerability in chronic treatment. To the best of our knowledge, this study is the only one that has obtained the sustained release of rivastigmine from PLGA microparticles after IM administration in an in vivo model.

Biocompatible polymeric microparticles serve as novel and reliable vehicles for exogenous hormone manipulations in passerines

The administration of exogenous hormones emerged as an essential tool for field studies in endocrinology. However, working with wild animals remains challenging, because under field conditions not every available method meets the necessary requirements. Achieving a sustained elevation in hormone levels, while simultaneously minimising handling time and invasiveness of the procedure is a difficult task in field endocrinology.Facing this challenge, we have investigated the suitability of biocompatible polymeric microparticles, a novel method for drug-administration, as a tool to manipulate hormones in small songbirds. We chose the insulin-like growth factor-1 (IGF-1) as target hormone, because it receives great interest from the research community due to its important role in shaping life-history traits. Moreover, its short half-life and hydrophilic properties imply a major challenge in finding a suitable method to achieve a sustained, systemic long-term release. To study the release kinetics, we injected either IGF-1 loaded polylactic-co-glycolic acid (PLGA) microparticles or dispersion medium (control group) in the skin pocket of the interscapular region of captive bearded reedlings (Panurus biarmicus). We collected blood samples for 7 consecutive days plus an additional sampling period after two weeks and complemented these with an in vitro experiment.Our results show that in vitro, PLGA microparticles allowed a stable IGF-1 release for more than 15 days, following a burst release at the beginning of the measurement. In vivo, the initial burst was followed by a drop to still elevated levels in circulating IGF-1 until the effect vanished by 16 days post-treatment.This study is the first to describe the use of PLGA-microparticles as a novel tool for exogenous hormone administration in a small passerine. We suggest that this method is highly suitable to achieve the systemic long-term release of hydrophilic hormones with short half-life and reduces overall handling time, as it requires only one subcutaneous injection.

Cellular Affinity of Particle-Stabilized Emulsion to Boost Antigen Internalization.

The cellular affinity of micro-/nanoparticles is the precondition for cellular recognition, cellular uptake, and activation, which are essential for drug delivery and immune response. The present study stemmed from the observation that the effects of charge, size, and shape of solid particles on cell affinity are usually considered, but we seldom realize the essential role of softness, dynamic restructuring phenomenon, and complex interface interaction in cellular affinity. Here, we developed poly-lactic-co-glycolic acid (PLGA) nanoparticle-stabilized Pickering emulsion (PNPE) that overcame the shortcomings of rigid forms and simulated the flexibility and fluidity of pathogens. A method was set up to test the affinity of PNPE to cell surfaces and elaborate on the subsequent internalization by immune cells. The affinity of PNPE to bio-mimetic extracellular vesicles (bEVs)-the replacement for bone marrow dendritic cells (BMDCs)-was determined using a quartz crystal microbalance with dissipation monitoring (QCM-D), which allowed real-time monitoring of cell-emulsion adhesion. Subsequently, the PNPE was used to deliver the antigen (ovalbumin, OVA) and the uptake of the antigens by BMDCs was observed using confocal laser scanning microscope (CLSM). Representative results showed that the PNPE immediately decreased frequency (ΔF) when it encountered the bEVs, indicating rapid adhesion and high affinity of the PNPE to the BMDCs. PNPE showed significantly stronger binding to the cell membrane than PLGA microparticles (PMPs) and AddaVax adjuvant (denoted as surfactant-stabilized nano-emulsion [SSE]). Furthermore, owing to the enhanced cellular affinity to the immunocytes through dynamic curvature changes and lateral diffusions, antigen uptake was subsequently boosted compared with PMPs and SSE. This protocol provides insights for designing novel formulations with high cell affinity and efficient antigen internalization, providing a platform for the development of efficient vaccines.

The Acute Immune Responses of the Common Carp Cyprinus carpio to PLGA Microparticles-The Interactions of a Teleost Fish with a Foreign Material.

Poly lactic-co-glycolic acid (PLGA) particles safely and effectively deliver pharmaceutical ingredients, with many applications approved for clinical use in humans. In fishes, PLGA particles are being considered as carriers of therapeutic drugs and vaccine antigens. However, existing studies focus mainly on vaccine antigens, the endpoint immune responses to these (e.g., improved antibody titres), without deeper understanding of whether fishes react to the carrier. To test whether or not PLGA are recognized by or interact at all with the immune system of a teleost fish, we prepared, characterized and injected PLGA microparticles intraperitoneally into common carp. The influx, phenotype of inflammatory leukocytes, and their capacity to produce reactive oxygen species and phagocytose PLGA microparticles were tested by flow cytometry, qPCR, and microscopy. PLGA microparticles were indeed recognized. However, they induced only transient recruitment of inflammatory leukocytes that was resolved 4 days later whereas only the smallest µm-sized particles were phagocytosed. The overall response resembled that described in mammals against foreign materials. Given the similarities between our findings and those described in mammals, PLGA particles can be adapted to play a dual role as both antigen and drug carriers in fishes, depending on the administered dose and their design.

Development of Long-Acting Injectable Ketamine Loaded PLGA Microparticles as a Non-opioid Analgesic

Background/Objective: Ketamine, a psychedelic, is a noncompetitive N-methyl-D-aspartate receptor antagonist that may also bind to mu opioid receptors. Historically, it has been used as an anesthetic (KetalarÒ), although now has found uses as a novel, quick acting, antidepressant for treatment-resistant depression (SpravatorÒ) and could be used as an adjuvant to opioid analgesia providing opioid-sparing effects. One major advantage over opioids is Ketamine does not suffer from respiratory depression and maintains patent airways during anesthesia. Ketamine is only available as a short-acting injectable solution or a nasal spray. Our goal is to develop a long-acting injectable form in a biodegradable matrix poly(lactic-co-glycolic) acid (PLGA) that does not have a burst release and provides 5-7 days of steady-state plasma levels. 
 Methods: A mechanistic approach towards development of a long-acting injectable began with a solubility screen of Ketamine. Based on these results, experiments began with an oil in water emulsification with two theoretical drug loadings (25% and 40%) and two processing conditions – (1) aqueous extraction and (2) aqueous extraction, intermediate drying, and a 25% Ethanol wash. The formulations were characterized for drug loading, drug release, and crystallinity and imaged using scanning electron microscopy (SEM). 
 Results: Minimal differences were noted in the release profiles between formulations. Although, a significant difference was noted between the two processing conditions, where the extra intermediate drying step and 25% ethanol wash resulted in a significant slowing of the drug release rate. 
 Conclusion and Implications: The difference in release kinetics is hypothesized to be due to densification of the PLGA matrix, based on the increase in surface roughness/wrinkling in the SEM images, crystallinity increase, and on their respective powder x-ray diffraction patterns. Our preliminary results demonstrate the feasibility of a longer acting Ketamine using PLGA. Further refinement of these formulations and rodent pharmacokinetic studies will be done in future.

Buprenorphine loaded PLGA microparticles: Characterization of a sustained-release formulation

Buprenorphine is a short-acting analgesic drug. Its use in veterinary medicine requires several injections per day to alleviate pain in animals. We, therefore, designed a poly-lactic-co-glycolic acid (PLGA) based depot formulation of the opioid to prolong the analgesic effect. We characterized our novel microparticulate depot formulation with emphasis on a potential future product development in the present work.Microparticles showed low residual moisture levels and did meet bacterial endotoxin requirements defined by the European Pharmacopeia. Depot formulation showed physicochemical stability over 6 months at 4 °C, indicating sufficient shelf life. Reconstituted formulation stored at 4 °C showed unchanged release properties for 24 h. Terminal sterilization by x-rays with a dose of 30 kGy revealed the sensitivity of buprenorphine towards radiation, resulting in substantial drug degradation. However, microparticle properties were not affected. Our experiments indicate that sterilization of the final product is not possible, therefore requiring aseptic manufacturing protocols.In conclusion, our novel buprenorphine depot formulation based on PLGA microparticles can be stored for at least 6 month as a lyophilisate and can be used for at least 24 h after reconstitution. Pilot experiments suggest that scale-up and a future aseptic production should be feasible. We conclude that the novel depot formulation shows promising attributes, a prerequisite for future industrial production and commercialization.

Co-delivery of artemether and piperine via core-shell microparticles for enhanced sustained release

Artemether, a highly efficient antimalarial drug, possesses the potential to treat patients suffering from Plasmodium-falciparum (P.F). However, artemether can be degraded rapidly by stomach acids and cleared quickly from the body after oral administration, leading to poor therapeutic effects, which is a significant hindrance in the clinical cure of malaria. To overcome this problem, we developed two types of core-shell microparticles co-loaded with artemether and piperine based on polylactic-co-glycolic acid (PLGA) and chitosan (CS) by using the promising coaxial electrospray system (CES) for sustained drug release. 1.9 μm artemether-piperine loaded PLGA-chitosan (AP-PLGA-CS) microparticles and 1.3 μm artemether-piperine loaded PLGA (AP-PLGA) microparticles were successfully fabricated using optimized operation parameters of CES. X-ray diffractometer (XRD) and differential scanning calorimetry (DSC) results showed the presence of artemether and piperine within the polymer matrix when encapsulated in the microparticles after CES. Both prepared AP-PLGA-CS and AP-PLGA microparticles exhibited high drug encapsulation and drug loading efficiency. Moreover, both of them displayed enhanced sustained drug release behavior due to the PLGA or PLGA-CS shell, which protected the fast degradation of artemether from the acidic gastric juice. All results suggested that the AP-PLGA-CS and AP-PLGA microparticles not only embrace the great potential in the application of malaria treatment but also provide a promising platform to encapsulate multiple drugs in polymeric particles for drug delivery.

Scaffold-based osteogenic dual delivery system with melatonin and BMP-2 releasing PLGA microparticles

The growing safety problems about the use of bone morphogenetic protein 2 (BMP-2) is one of the recent issues that was improved by using low doses of BMP-2 with the support of other osteoinductive agents and/or using appropriate carriers. The aim of the present study is to investigate the effect of scaffold-based dual release system including melatonin (MEL) and BMP-2 loaded polylactic-co-glycolic acid (PLGA) microparticles on the osteogenic activity of pre-osteoblastic MC3T3-E1 cells. MEL and BMP-2 loaded microparticles were prepared by double emulsion solvent evaporation method in the average diameters of ~2 µm and ~11 µm, respectively and loaded into chitosan/hydroxyapatite (HAp) scaffolds. In vitro MC3T3-E1 culture studies were carried out comparatively with blank scaffolds, single (BMP-2 or MEL) releasing groups and dual (BMP-2 and MEL) releasing group. Microscopic observations and hematoxylin/eosin staining showed enhanced number of cells and dense ECM in dual release group. The expressions of differentiation markers, Runt-related transcription factor 2 (RUNX2) and alkaline phosphatase (ALP) and also mineralization were higher in dual release group than that of the other groups. Our findings showed that BMP-2 at low doses (~20 ng per scaffold) was sufficient in terms of osteogenic activity with controlled release systems where it was used in combination with MEL (~10 µg per scaffold).

Microparticle Encapsulation of a Prostate-targeted Biologic for the Treatment of Liver Metastases in a Preclinical Model of Castration-resistant Prostate Cancer

PRX302 is a highly potent, mutant bacterial pore-forming biologic protoxin engineered for selective activation by PSA, a serine protease expressed by benign and malignant prostate epithelial cells. Although being developed as a local therapy for benign prostatic hyperplasia and localized prostate cancer, PRX302 cannot be administered systemically as a treatment for metastatic disease due to binding to ubiquitously expressed glycosylphosphatidylinositol (GPI)-anchored proteins, which leads to poor accumulation within the tumor microenvironment. To overcome this limitation, poly-lactic-co-glycolic acid (PLGA) microparticles encapsulating the protoxin were developed, which are known to accumulate in the liver, a major site of metastasis for prostate cancer and other solid tumors. A highly sensitive and reproducible sandwich ELISA to quantify PRX302 released from microparticles was developed. Utilizing this assay, PRX302 release from different microparticle formulations was assessed over multiple days. Hemolysis assays documented PSA-dependent pore formation and lytic potential (i.e., function) of the released protoxin. MTT assays demonstrated that conditioned supernatant from PRX302-loaded, but not blank (i.e., unloaded), PLGA microparticles was highly cytotoxic to PC3 and DU145 human prostate cancer cells in the presence of exogenous PSA. Microparticle encapsulation prevented PRX302 from immediately interacting with GPI-anchored proteins as demonstrated in a competition assay, which resulted in an increased therapeutic index and significant antitumor efficacy following a single dose of PRX302-loaded microparticles in a preclinical model of prostate cancer liver metastasis with no obvious toxicity. These results document that PRX302 released from PLGA microparticles demonstrate in vivo antitumor efficacy in a clinically relevant preclinical model of metastatic prostate cancer.

Poly(lactic-co-glycolic) Acid-Lipid Hybrid Microparticles Enhance the Intracellular Uptake and Antibacterial Activity of Rifampicin.

An urgent demand exists for the development of effective carrier systems that systematically enhance the cellular uptake and localization of antibiotic drugs for the treatment of intracellular pathogens. Commercially available antibiotics suffer from poor cellular penetration, restricting their efficacy against pathogens hosted and protected within phagocytic cells. In this study, the potency of the antibiotic rifampicin against intracellular small colony variants of Staphylococcus aureus was improved through encapsulation within a strategically engineered cell-penetrant delivery system, composed of lipid nanoparticles encapsulated within a poly(lactic-co-glycolic) acid (PLGA) nanoparticle matrix. PLGA-lipid hybrid (PLH) microparticles were synthesized through the process of spray drying, whereby rifampicin was loaded within both the polymer and lipid phases, to create a nanoparticle-in-microparticle system capable of efficient redispersion in aqueous biorelevant media and with programmable release kinetics. The ability of PLH particles to disintegrate into nanoscale agglomerates of the precursor nanoparticles was shown to be instrumental in optimizing rifampicin uptake in RAW264.7 macrophages, with a 7.2- and 1.6-fold increase in cellular uptake, when compared to the pure drug and PLGA microparticles (of an equivalent initial particle size), respectively. The enhanced phagocytosis and extended drug release mechanism (under the acidic macrophage environment) associated with PLH particles induced a 2.5-log reduction in colony forming units compared to initial colonies at 2.50 μg/mL rifampicin dose. Thus, the ability of PLH particles to reduce the intracellular viability of S. aureus, without demonstrating significant cellular toxicity, satisfies the requirements necessary for the safe and efficacious delivery of antibiotics to macrophages for the treatment of intracellular infections.

Fast dissolving glucose porogens for early calcium phosphate cement degradation and bone regeneration

Here, we demonstrate the in vivo efficacy of glucose microparticles (GMPs) to serve as porogens within calcium phosphate cements (CPCs) to obtain a fast-degrading bone substitute material. Composites were fabricated incorporating 20 wt% GMPs at two different GMP size ranges (100–150 μm (GMP-S) and 150–300 μm (GMP-L)), while CPC containing 20 wt% poly(lactic-co-glycolic acid) microparticles (PLGA) and plain CPC served as controls. After 2 and 8 weeks implantation in a rat femoral condyle defect model, specimens were retrieved and analyzed for material degradation and bone formation. Histologically, no adverse tissue response to any of the CPC-formulations was observed. All CPC-porogen formulations showed faster degradation compared to plain CPC control, but only GMP-containing formulations showed higher amounts of new bone formation compared to plain CPC controls. After 8 weeks, only CPC-porogen formulations with GMP-S or PLGA porogens showed higher degradation compared to plain CPC controls. Overall, the inclusion of GMPs into CPCs resulted in a macroporous structure that initially accelerated the generation of new bone. These findings highlight the efficacy of a novel approach that leverages simple porogen properties to generate porous CPCs with distinct degradation and bone regeneration profiles.

Preparation and Characterization of Terpenoid-Encapsulated PLGA Microparticles and its Antibacterial Activity against <i>Enterococcus faecalis</i>

Exploration of natural compound for the treatment of dental-related problems are gaining of interest for enhancing therapeutic efficacy of the drugs delivery system. In this study, we have prepared terpenoid, which have been isolated from Myrmecodia pendens Merr & Perry from Papua Island, Indonesia, to be encapsulated in Polylactic-co-glycolic acid (PLGA), as the most widely used biodegradable polymer for biomedical applications, through one step single-emulsion method followed by subsequent coating by poly (vinyl alcohol) (PVA). The resultant of terpenoid-loaded PLGA microparticles were characterized systematically through scanning electron microscope and Fourier-transform infrared spectroscopy. In vitro drug release test was evaluated through dialysis method. Antibacterial test was conducted against Enterococcus faecalis as a model for persistent bacteria that causes root canal infections. The results showed that terpenoid-loaded PLGA microparticles were developed in spherical morphology with an average particle size of around 1-2μm. Terpenoid released from PLGA compartment at pH 6.5 and temperature of 37°C through a controlled-release profile mechanism with enhanced prolonged release. The bacterial assay result showed that terpenoid-loaded PLGA microparticles could reduce Enterococcus faecalis, effectively. Eventually, these result show that terpenoid-loaded PLGA microparticles as unique natural product-based extract could be developed as a potential naturally-based drug for dental-related diseases applications.

An Artificial Antigen-Presenting Cell Delivering 11 Immune Molecules Expands Tumor Antigen-Specific CTLs in Ex Vivo and In Vivo Murine Melanoma Models.

Antigen-presenting cells expand antigen-specific T cells ex vivo and in vivo for tumor immunotherapy, but are time-consuming to generate and, as live cells, raise biosafety concerns. An alternative is found in cell-free artificial antigen-presenting cells (aAPC), but these only present two or three kinds of immune molecules. Here, we describe a multipotent artificial antigen-presenting cell (MaAPC) that delivered 11 kinds of immune moleclues. This MaAPC simulated natural APCs through the concurent coupling of target antigens (H-2Kb/TRP2180-188-Ig dimers and H-2Db/gp10025-33-Ig dimers), costimulatory molecules (anti-CD28, anti-4-1BB, and anti-CD2), and "self-marker" CD47-Fc onto surface-modified polylactic-co-glycolic acid microparticles (PLGA-MP). These PLGA-MPs also encapsulated cytokines (IL2 and IL15), a chemokine (CCL21), and checkpoint inhibitors (anti-CTLA-4 and anti-PD-1). Culture of MaAPCs with naïve T cells for 1 week elevated the frequencies of TRP2180-188-specific and gp10025-33-specific CTLs to 51.0% and 43.3%, respectively, with enhanced cytotoxicity. Three infusions of MaAPCs inhibited subcutaneous melanoma growth in a mouse model and expanded TRP2180-188 and gp10025-33-specific CTLs 59-86-fold in peripheral blood, 76-77-fold in spleen, and 205-212-fold in tumor tissue, in an antigen-specific manner. Compared with conventional aAPCs carrying two or three immune molecules, the 11-signal MaAPCs exerted greater impact on T cells, including activation, proliferation, cytotoxicity, differentiation to memory CTLs or regulatory T cells and cytokines profiles, without detected side effects. Such MaAPCs could be used to individualize tumor immunotherapy.

Encapsulation of Photosystem I in Organic Microparticles Increases Its Photochemical Activity and Stability for Ex Vivo Photocatalysis.

Photosystem I (PSI) is a pigment binding multisubunit protein complex involved in the light phase of photosynthesis, catalyzing a light-dependent electron transfer reaction from plastocyanin to ferredoxin. PSI is characterized by a photochemical efficiency close to one, suggesting its possible application in light-dependent redox reaction in an extracellular context. The stability of PSI complexes isolated from plant cells is however limited if not embedded in a protective environment. Here we show an innovative solution for exploiting the photochemical properties of PSI, by encapsulation of isolated PSI complexes in PLGA (poly lactic-co-glycolic acid) organic microparticles. These encapsulated PSI complexes were able to catalyze light-dependent redox reactions with electron acceptors and donors outside the PLGA microparticles. Moreover, PSI complexes encapsulated in PLGA microparticles were characterized by a higher photochemical activity and stability compared with PSI complexes in detergent solution, suggesting their possible application for ex vivo photocatalysis.

Ramizol® encapsulation into extended release PLGA micro- and nanoparticle systems for subcutaneous and intramuscular administration: in vitro and in vivo evaluation

Objective: Novel antibiotic Ramizol® is advancing to clinical trials for the treatment of gastrointestinal Clostridium difficile associated disease. Despite this, previous studies have shown a rapid plasma clearance upon intravenous administration and low oral bioavailability indicating pure drug is unsuitable for systemic infection treatment following oral dosing. The current study aims to investigate the development of poly-lactic-(co-glycolic) acid (PLGA) particles to overcome this limitation and increase the systemic half-life following subcutaneous and intramuscular dosing.Significance: The development of new antibiotic treatments will help in combatting the rising incidence of antimicrobial resistance.Methods: Ramizol® was encapsulated into PLGA nano and microparticles using nanoprecipitation and emulsification solvent evaporation techniques. Formulations were analyzed for particle size, loading level and encapsulation efficiency as well as in vitro drug release profiles. Final formulation was advanced to in vivo pharmacokinetic studies in Sprague–Dawley rats.Results: Formulation technique showed major influence on particle size and loading levels with optimal loading of 9.4% and encapsulation efficiency of 92.06%, observed using emulsification solvent evaporation. Differences in formulation technique were also linked with subsequent differences in release profiles. Pharmacokinetic studies in Sprague–Dawley rats confirmed extended absorption and enhanced bioavailability following subcutaneous and intramuscular dosing with up to an 8-fold increase in Tmax and T1/2 when compared to the oral and IV routes.Conclusions: Subcutaneous and intramuscular dosing of PLGA particles successfully increased systemic half-life and bioavailability of Ramizol®. This formulation will allow further development of Ramizol® for systemic infection eradication.

Lipoxin A4 encapsulated in PLGA microparticles accelerates wound healing of skin ulcers.

Lipoxin A4 (LXA4) is involved in the resolution of inflammation and wound healing; however, it is extremely unstable. Thus, to preserve its biological activities and confer stability, we encapsulated LXA4 in poly-lactic-co-glycolic acid (PLGA) microparticles (LXA4-MS) and assessed its application in treating dorsal rat skin lesions. Ulcers were sealed with fibrin adhesive and treated with either LXA4-MS, unloaded microparticles (Un-MS), soluble LXA4, or PBS/glue (vehicle). All groups were compared at 0, 2, 7, and 14 days post-lesions. Our results revealed that LXA4-MS accelerated wound healing from day 7 and reduced initial ulcer diameters by 80%. Soluble LXA4, Un-MS, or PBS closed wounds by 60%, 45%, and 39%, respectively. LXA4-MS reduced IL-1β and TNF-α, but increased TGF-β, collagen deposition, and the number of blood vessels. Compared to other treatments, LXA4-MS reduced inflammatory cell numbers, myeloperoxidase (MPO) concentration, and metalloproteinase-8 (MMP8) mRNA in scar tissue, indicating decreased neutrophil chemotaxis. In addition, LXA4-MS treatment increased macrophages and IL-4, suggesting a positive impact on wound healing. Finally, we demonstrated that WRW4, a selective LXA4 receptor (ALX) antagonist, reversed healing by 50%, indicating that LXA4 must interact with ALX to induce wound healing. Our results show that LXA4-MS could be used as a pharmaceutical formulation for the treatment of skin ulcers.

Paracrine release of IL-2 and anti-CTLA-4 enhances the ability of artificial polymer antigen-presenting cells to expand antigen-specific T cells and inhibit tumor growth in a mouse model.

Accumulating evidence indicates that bead-based artificial antigen-presenting cells (aAPCs) are a powerful tool to induce antigen-specific T cell responses in vitro and in vivo. To date, most conventional aAPCs have been generated by coupling an antigen signal (signal 1) and one or two costimulatory signals, such as anti-CD28 with anti-LFA1 or anti-4-1BB (signal 2), onto the surfaces of cell-sized or nanoscale magnetic beads or polyester latex beads. The development of a biodegradable scaffold and the combined use of multiple costimulatory signals as well as third signals for putative clinical applications is the next step in the development of this technology. Here, a novel biodegradable aAPC platform for active immunotherapy was developed by co-encapsulating IL-2 and anti-CTLA-4 inside cell-sized polylactic-co-glycolic acid microparticles (PLGA-MPs) while co-coupling an H-2Kb/TRP2-Ig dimer and anti-CD28 onto the surface. Cytokines (activating signal) and antibodies (anti-inhibition signal) were efficiently co-encapsulated in PLGA-MP-based aAPCs and co-released without interfering with each other. The targeted, sustained co-release of IL-2 and anti-CTLA-4 achieved markedly enhanced, synergistic effects in activating and expanding tumor antigen-specific T cells both in vitro and in vivo, as well as in inhibiting tumor growth in a mouse melanoma model, as compared with conventional two-signal aAPCs and IL-2 or anti-CTLA-4 single-released aAPCs. These data revealed the feasibility and importance of the paracrine release of multiple costimulatory molecules and cytokines from biodegradable aAPCs and thus provide a proof of principle for the future use of polymeric aAPCs for active immunotherapy of tumors and infectious diseases.

Effect of solution and apparatus parameters on the morphology and size of electrosprayed PLGA microparticles

Because of a number of facilities, the Electrospray (ES) method is gaining ever-increasing popularity among researchers for producing nano-to-micron-sized particles. Microparticles (MPs) of poly lactic-co-glycolic acid (PLGA) were prepared by using the ES technique. The influence of both solution and apparatus parameters on the morphology, size, size distribution, and uniformity of produced MPs were investigated. Results of SEM images and calculations revealed that polymer concentration is a critical parameter in the ES system. In a semi-dilute moderately entangled regime, chain entanglement can easily occur. Solution flow rate is a key factor among apparatus parameters. Vapour pressure is a key parameter affecting MP morphology. The size of the particles tended to reduce with an increase in voltage. The needle gauge did not have an important impact on particle size. The role of the electric field changed at different collecting distances. Using a saturated combination of EtOH/PVA is an acceptable collecting medium for PLGA MPs. It is possible to produce uniform and spherical MPs by using chloroform as a solvent. However, a reduction in particle size is achievable by using a solvent of chloroform/DMF (90/10 w/w).

Coaxial Electrospray of Ranibizumab-Loaded Microparticles for Sustained Release of Anti-VEGF Therapies.

Age-related macular degeneration (AMD) is the leading cause of vision loss and blindness in people over age 65 in industrialized nations. Intravitreous injection of anti-VEGF (vascular endothelial growth factor) therapies, such as ranibizumab (trade name: Lucentis), provides an effective treatment option for neovascular AMD. We have developed an improved coaxial electrospray (CES) process to encapsulate ranibizumab in poly(lactic-co-glycolic) acid (PLGA) microparticles (MPs) for intravitreous injection and sustained drug release. This microencapsulation process is advantageous for maintaining the stability of the coaxial cone-jet configurations and producing drug-loaded MPs with as high as 70% encapsulation rate and minimal loss of bioactivitiy. The utility of this emerging process in intravitreous drug delivery has been demonstrated in both benchtop and in vivo experiments. The benchtop test simulates ocular drug release using PLGA MPs encapsulating a model drug. The in vivo experiment evaluates the inflammation and retinal cell death after intravitreal injection of the MPs in a chick model. The experimental results show that the drug-load MPs are able to facilitate sustained drug release for longer than one month. No significant long term microglia reaction or cell death is observed after intravitreal injection of 200 μg MPs. The present study demonstrates the technical feasibility of using the improved CES process to encapsulate water-soluble drugs at a high concentration for sustained release of anti-VEGF therapy.