Double Emulsion Solvent Evaporation Method Research Articles

Luminescent/magnetic PLGA-based hybrid nanocomposites: a smart nanocarrier system for targeted codelivery and dual-modality imaging in cancer theranostics.

Cancer diagnosis and treatment represent an urgent medical need given the rising cancer incidence over the past few decades. Cancer theranostics, namely, the combination of diagnostics and therapeutics within a single agent, are being developed using various anticancer drug-, siRNA-, or inorganic materials-loaded nanocarriers. Herein, we demonstrate a strategy of encapsulating quantum dots, superparamagnetic Fe3O4 nanocrystals, and doxorubicin (DOX) into biodegradable poly(d,l-lactic-co-glycolic acid) (PLGA) polymeric nanocomposites using the double emulsion solvent evaporation method, followed by coupling to the amine group of polyethyleneimine premodified with polyethylene glycol-folic acid (PEI-PEG-FA [PPF]) segments and adsorption of vascular endothelial growth factor (VEGF)-targeted small hairpin RNA (shRNA). VEGF is important for tumor growth, progression, and metastasis. These drug-loaded luminescent/magnetic PLGA-based hybrid nanocomposites (LDM-PLGA/PPF/VEGF shRNA) were fabricated for tumor-specific targeting, drug/gene delivery, and cancer imaging. The data showed that LDM-PLGA/PPF/VEGF shRNA nanocomposites can codeliver DOX and VEGF shRNA into tumor cells and effectively suppress VEGF expression, exhibiting remarkable synergistic antitumor effects both in vitro and in vivo. The cell viability waŝ14% when treated with LDM-PLGA/PPF/VEGF shRNA nanocomposites ([DOX] =25 μg/mL), and in vivo tumor growth data showed that the tumor volume decreased by 81% compared with the saline group at 21 days postinjection. Magnetic resonance and fluorescence imaging data revealed that the luminescent/magnetic hybrid nanocomposites may also be used as an efficient nanoprobe for enhanced T2-weighted magnetic resonance and fluorescence imaging in vitro and in vivo. The present work validates the great potential of the developed multifunctional LDM-PLGA/PPF/VEGF shRNA nanocomposites as effective theranostic agents through the codelivery of drugs/genes and dual-modality imaging in cancer treatment.

Nano-particle engineered atorvastatin delivery to support mesenchymal stem cell survival in infarcted myocardium.

Atorvastatin (ATV) may support mesenchymal stem cells (MSC) survival in post-infarct myocardium (MI) as inflammatory reactions, oxidative stress and hypoxia condition get started in such tissues after damage. However, limited aqueous insolubility and rapid first-pass metabolism reduce the systemic availability of ATV. The aim of the present investigation was to develop ATV loaded nanoparticles (ATVNPs) which might ensure the maximum availability of ATV in systemic circulation for longer duration and to strengthen the support to MSC survival. ATVNPs were synthesized using double emulsion solvent evaporation method and characterized as spherical shape, positive charged, nanoparticles of uniform size distribution and higher entrapment efficiency. ATVNPs were non-cytotoxic and showed sustained release (up to 28 days). Assessment of cardiac function (in terms of echocardiographic and left heart catheterization parameters) and cytokines estimation revealed efficient improvement in post-infarct myocardium condition of rat. In conclusion, ATVNP was developed successfully that may ensure safe, cost effective, and efficacious treatment of post-infarct myo-cardium when compared with that of MSC alone and MSC supplemented with ATV solution.

Mn2+-coordinated PDA@DOX/PLGA nanoparticles as a smart theranostic agent for synergistic chemo-photothermal tumor therapy.

Nanoparticle drug delivery carriers, which can implement high performances of multi-functions, are of great interest, especially for improving cancer therapy. Herein, we reported a new approach to construct Mn2+-coordinated doxorubicin (DOX)-loaded poly(lactic-co-glycolic acid) (PLGA) nanoparticles as a platform for synergistic chemo-photothermal tumor therapy. DOX-loaded PLGA (DOX/PLGA) nanoparticles were first synthesized through a double emulsion-solvent evaporation method, and then modified with polydopamine (PDA) through self-polymerization of dopamine, leading to the formation of PDA@DOX/PLGA nanoparticles. Mn2+ ions were then coordinated on the surfaces of PDA@DOX/PLGA to obtain Mn2+-PDA@DOX/PLGA nanoparticles. In our system, Mn2+-PDA@DOX/PLGA nanoparticles could destroy tumors in a mouse model directly, by thermal energy deposition, and could also simulate the chemotherapy by thermal-responsive delivery of DOX to enhance tumor therapy. Furthermore, the coordination of Mn2+ could afford the high magnetic resonance (MR) imaging capability with sensitivity to temperature and pH. The results demonstrated that Mn2+-PDA@ DOX/PLGA nanoparticles had a great potential as a smart theranostic agent due to their imaging and tumor-growth-inhibition properties.

Design, characterization and optimization of lamivudine-loaded amphiphilic HA-g-ECL nanoparticles

This investigation sought to engineer and optimize a novel Lamivudine (LMV)-loaded nanoformulation through grafting of hyaluronic acid (HA) and epsilon-caprolactam (ECL) (HA-g-ECL) for potentially enhanced treatment of Hepatitis B Virus (HBV) infection. The HA-g-ECL copolymer and subsequently its nanoparticles were prepared by anionic graft copolymerization, and a double emulsion solvent-evaporation method, respectively. Verification of grafting of the novel amphiphilic copolymer was via analysis of the vibrational, thermal and rheological transitions of HA-g-ECL, as well as demonstration of a critical micelle concentration (CMC). A Box-Behnken experimental design was applied to generate 15 nanoparticulate formulations, which were evaluated for their physicochemical properties, including mean particle size, morphology, drug entrapment efficiency (DEE), and drug release characteristics (MDT). There was a significant interaction effect on the MDT between the level of copolymer and surfactant (p = 0.008). Approximately 65% LMV was released in the first 15 h for design formulations comprising lower levels of HA-g-ECL, while ∼20% LMV was released with higher levels of HA-g-ECL. Varying the quantities of copolymer, surfactant and solvent thus had notable implications in controlling the drug entrapment efficiency and release from the copolymeric system. The optimized nanoparticulate system demonstrates potential for the controlled delivery of LMV for treatment of HBV infection.

Inhaled sildenafil as an alternative to oral sildenafil in the treatment of pulmonary arterial hypertension (PAH)

The practice of treating PAH patients with oral or intravenous sildenafil suffers from the limitations of short dosing intervals, peripheral vasodilation, unwanted side effects, and restricted use in pediatric patients. In this study, we sought to test the hypothesis that inhalable poly(lactic-co-glycolic acid) (PLGA) particles of sildenafil prolong the release of the drug, produce pulmonary specific vasodilation, reduce the systemic exposure of the drug, and may be used as an alternative to oral sildenafil in the treatment of PAH. Thus, we prepared porous PLGA particles of sildenafil using a water-in-oil-in-water double emulsion solvent evaporation method with polyethyleneimine (PEI) as a porosigen and characterized the formulations for surface morphology, respirability, in-vitro drug release, and evaluated for in vivo absorption, alveolar macrophage uptake, and safety. PEI increased the particle porosity, drug entrapment, and produced drug release for 36h. Fluorescent particles showed reduced uptake by alveolar macrophages. The polymeric particles were safe to rat pulmonary arterial smooth muscle cell and to the lungs, as evidenced by the cytotoxicity assay and analyses of the injury markers in the bronchoalveolar lavage fluid, respectively. Intratracheally administered sildenafil particles elicited more pulmonary specific and sustained vasodilation in SUGEN-5416/hypoxia-induced PAH rats than oral, intravenous, or intratracheal plain sildenafil did, when administered at the same dose. Overall, true to the hypothesis, this study shows that inhaled PLGA particles of sildenafil can be administered, as a substitute for oral form of sildenafil, at a reduced dose and longer dosing interval.

Core‑shell lipid polymer nanoparticles for combined chemoand gene therapy of childhood head and neck cancers.

Pediatric head and neck cancers account for overall 12% of all pediatric cancers. Despite recent advances in therapeutic modalities, children with tumor metastasis have poor prognosis. Therefore, there is an unmet need for new and effective treatment modalities for pediatric head and neck cancers. The present study describes a simple and efficient method for fabrication of cationic lipid‑polymer hybrid nanoparticles(CLPNs) for co‑delivery of cisplatin(CDDP) and DNA(CDDP/DNA CLPNs) for the therapy of childhood head and neck cancers. CDDP/DNA CLPNs were prepared by the modified double emulsion solvent evaporation method with self‑assembly. CDDP‑loaded CLPNs(CDDP CLPNs), CDDP-loaded polymeric nanoparticles(PNPs)(CDDP PNPs), and DNA‑loaded Lipofectamine®2000(DNA LIPO) were also prepared for comparison. The results illustrated that the concentration of the cationic lipid has influence on the characteristics of CLPNs. Invitro anticancer effect, invitro transfection efficiency, invivo antitumor and gene delivery efficacy of CDDP/DNA CLPNs have advantages over other formulations tested. In conclusion, outstanding delivery ability of CLPNs for both CDDP and DNA could combine the therapeutic efficiency of both drug and gene for the treatment of pediatric rhabdomyosarcoma(RMS).

Low-Molecular-Weight Heparin-Coated and Montelukast-Filled Inhalable Particles: A Dual-Drug Delivery System for Combination Therapy in Asthma

Montelukast, a cysteinyl leukotriene type 1 receptor antagonist, exhibits secondary anti-inflammatory properties when used at higher concentrations. Low-molecular-weight heparin (LMWH) evokes pronounced anti-inflammatory effects by interrupting leukocyte adhesion and migration. We hypothesized that inhalable particles containing montelukast plus LMWH release both drugs in a sustained fashion and protect the lungs against allergen-induced inflammation. Large porous particles of montelukast and LMWH were prepared using a double-emulsion–solvent-evaporation method. Montelukast was first encapsulated in copolymer-based particles using polyethylenimine as a porosigen; the resulting particles were then coated with LMWH. The particles were evaluated for physicochemical properties, respirability, and release profiles. The anti-inflammatory effect of the optimized formulation was studied in ovalbumin-sensitized asthmatic Sprague Dawley rats. The optimized large porous particles had a diameter of 10.3 ± 0.7 μm, exhibited numerous surface indentations and pores, showed acceptable drug entrapment efficiency (66.8% ± 0.4% for montelukast; 91.7% ± 0.8% adsorption efficiency for LMWH), demonstrated biphasic release patterns, and escaped the uptake by the rat alveolar macrophages. The number of infiltrating inflammatory cells in asthmatic rat lungs, treated with dual-drug particles, was >74% fewer than in untreated asthmatic rat lungs. Similarly, the airway walls of asthmatic animals treated with dual-drug particles were 3-fold thinner than those of untreated asthmatic animals (p < 0.001). The optimized formulation protects lungs against methacholine-induced airway hyper-reactivity. Overall, this study demonstrates the feasibility of loading 2 drugs, montelukast and LMWH, into an inhalable particulate system and establishes that this novel combination therapy produces sustained drug release and elicits a robust anti-inflammatory response in the lungs.

Fabrication of PLA-PEG Nanoparticles as Delivery Systems for Improved Stability and Controlled Release of Catechin

The purpose of this study was to develop an oral delivery system for the controlled release of catechin and evaluate the antioxidant potential and stability of catechin loaded PLA/PEG nanoparticles (CATNP). Nanoparticles were synthesized using a double emulsion solvent evaporation method. The fabricated nanoparticles were relatively small with a hydrodynamic diameter of 300 nm and an encapsulation efficiency of 95%. SEM image analysis showed uniform sized and spherically shaped nanoparticles.In vitrorelease profiles indicated a slow and sustained release of catechin from the nanoparticle. Stability of the nanoparticle in simulated gastric and intestinal fluids is maintained due to the PEG coating on the nanoparticles, which effectively protected catechin against gastrointestinal enzyme activity. Enhanced inhibition action of free radicals and metal chelation potential was noted when catechin was encapsulated in these polymeric nanoparticles. The reports obtained from this study would provide an opportunity for designing an oral delivery system aimed at inhibiting oxidative stress in the human body.

PLGA-PEG-PLGA triblock copolymeric micelles as oral drug delivery system: In vitro drug release and in vivo pharmacokinetics assessment

Poly (d,l-lactide-co-glycolide)-poly (ethylene glycol)-poly (d,l-lactide-co-glycolide) triblock copolymers (PLGA-PEG-PLGA) has been proven to be desirable for anti-cancer drug delivery by intravenous administration. But till now there is no report of developing this micelle as a sustained oral formulation for cancer therapy. 3β-acetoxy-urs-12-en-28-oic acid hexamethylenediamine (US597), a derivative of natural product ursolic acid has been developed as a novel cancer metastasis chemopreventive agent by us. Herein, we developed a new oral dosage formulation of PLGA-PEG-PLGA tri-block micelles loaded with US597 (US597@micelles). US597@micelles was prepared by a double emulsion solvent evaporation method, and characterized in regards to mean diameter (<100nm), drug loading (25.9–28.5%), zeta potential (5.76–10.65mV) and encapsulation efficiency (55.7–74.3%), respectively. In vitro, US597@micelles could ameliorate sustained drug release, inhibit cell proliferation by inducing apoptosis (46.6% of late apoptosis), and influence the integrity of nuclei and mitochondrial on HepG2. Moreover, in vivo pharmacokinetic study by UPLC/MS/MS method demonstrated better absorption, metabolism and elimination characters of US597@micelles as an oral dosage form (Cmax=53±49ng/mL, t1/2=8.716±7.033h) over free US597 (Cmax=14±11ng/mL, t1/2=16.433±8.821h). In conclusion, PLGA-PEG-PLGA micelles as a promising oral drug delivery system are able to improve the bioavailability and efficacy of US597 in cancer therapy.

Chitosan-coated PLGA nanoparticles of bevacizumab as novel drug delivery to target retina: optimization, characterization, and in vitro toxicity evaluation

In several ocular diseases, the vascular endothelial growth factor (VEGF) level has been found to be upregulated. Bevacizumab, an anti-VEGF drug, is the most commonly used off level drug for these conditions. Delivery of drug to the posterior site is desired for the effective management of these diseases. The present study was to develop and optimize the chitosan (CS)-coated poly(lactide-co-glycolic acid) (PLGA) nanoparticles (NPs) of bevacizumab for sustained and effective delivery to posterior ocular tissues. NPs were prepared by double emulsion solvent evaporation method and optimized for various variables (i.e., CS concentration, PLGA content, polyvinyl alcohol (PVA) concentration, and sonication time) by employing a 4-factor 3-level Box–Behnken statistical design. NPs were characterized for particle size, polydispersity index (PDI), entrapment efficiency (EE), and in vitro release. Transscleral flux was determined through goat sclera, and ocular tolerance assay was done by Hen’s Egg Test chorioallantoic membrane method. The particle size and PDI of the optimized NPs were 222.28 ± 7.45 nm and 0.19 ± 0.08, respectively. The developed NPs showed an EE of 69.26 ± 1.31% with an extended release profile. The flux was significantly higher that is, 0.3204 ± 0.026 μg/cm2/h for the NPs compared to drug solution. Thus, CS-coated PLGA NPs can be potentially useful as ocular drug carriers to target retina.

Nevirapine Loaded Core Shell Gold Nanoparticles by Double Emulsion Solvent Evaporation: In vitro and In vivo Evaluation

HIV/AIDS is a macrophage resident infection localized in the reticuloendothelial system and remote locations of brain and bone marrow. We present core shell nanoparticles of gold(AuNPs) and nevirapine(NVP) for targeted delivery to the multiple HIV reservoirs. The aim of the study was to design core shell NVP loaded AuNPs with high drug loading and to evaluate biodistribution of the nanoparticles in possible HIV reservoirs in vivo. A specific objective was to assess the possible synergy of AuNPs with NVP on anti-HIV activity in vitro. Core shell nanoparticles were prepared by double emulsion solvent evaporation method and characterized. Glyceryl monostearate-nevirapine-gold nanoparticles(GMS-NVP-AuNPs) revealed high entrapment efficiency (>70%), high loading (~40%), particle size <250 nm and zeta potential -35.9± 1.41mv and exhibited sustained release with good stability. Surface plasmon resonance indicated shell formation while SEM coupled EDAX confirmed the presence of Au. TEM confirmed formation of spherical core shell nanoparticles. GMS-NVP-AuNPs revealed low hemolysis (<10 %) and serum stability upto 6 h. GMS-NVP-AuNPs exhibited rapid, high and sustained accumulation in the possible HIV reservoir organs, including the major organs of liver, spleen, lymph nodes, thymus and also remote locations of brain, ovary and bone marrow. High cell viability and enhanced uptake in PBMC's and TZM-bl cells were observed. While uptake in PBMC's proposed monocytes/macrophages enabled brain delivery. GMS-NVP-AuNPs demonstrated synergistic anti-HIV activity. The superior anti-HIV activity in vitro coupled with extensive localization of the nanoparticles in multiple HIV reservoirs suggests great promise of the core shell GMS-NVP-AuNPs for improved therapy of HIV.

Synergistic effect of rSAG1 and rGRA2 antigens formulated in PLGA microspheres in eliciting immune protection against Toxoplasama gondii

There is still no human vaccine against Toxoplasma gondii (T. gondii), as one of the most successful parasites. In present study, we designed a subunit vaccine composed of recombinant SAG1 (rSAG1) and recombinant GRA2 (rGRA2) proteins. In order to improve the induced immune responses, rSAG1 and rGRA2 were adsorbed on Poly (DL-lactide-co-glycolide) (PLGA) microspheres (MS) prepared by double emulsion solvent evaporation method. BALB/c mice were subcutaneously vaccinated by rSAG1-adsorbed PLGA MS (rSAG1-PLGA), rGRA2-adsorbed PLGA MS (rGRA2-PLGA), and the mixture of both formulations (rSAG1/rGRA2-PLGA), twice with a 3-week interval. PLGA MS characteristics, protein release, cellular and humoral immune responses, and protection against acute toxoplasmosis were evaluated. All vaccinated mice induced significantly partial protection and longer survival times associated with higher IFN-γ/IL-10 ratio and higher amount of Toxoplasma-specific IgG antibodies compared to control groups. Interestingly, the synergistic effect of rSAG1 and rGRA2 in eliciting more potent cellular and humoral responses and consequently higher protection in comparison to single antigen was confirmed.This study introduces the mixture of rSAG1 and rGRA2 (derived from different stages of Toxoplasma life-cycle) formulated in PLGA MS as a promising candidate in vaccine development against T. gondii.

Characterizing release mechanisms of leuprolide acetate-loaded PLGA microspheres for IVIVC development I: In vitro evaluation

Release testing of parental controlled release microspheres is an essential step in controlling quality and predicting the duration of efficacy. In the first of a two-part study, we examined the effect of various incubation media on release from leuprolide-loaded PLGA microspheres to understand the influence of external pH, plasticization, and buffer type on mechanism of accelerated release. PLGA 50/50 microspheres encapsulating ~5% w/w leuprolide were prepared by the double emulsion-solvent evaporation method with or without gelatin or by the self-healing encapsulation method. The microspheres were incubated at 37°C up to 56days in various media: pH5.5, 6.5, and 7.4 phosphate buffered-saline (PBS) containing 0.02% Tween 80; pH7.4 PBS containing 1.0% triethyl citrate (PBStc); and pH7.4 HEPES buffered-saline containing 0.02% Tween 80 (all media contained 0.02% sodium azide). The recovered release media and microspheres were examined for released drug, polymer molecular weight (Mw), water uptake, mass loss, and BODIPY (green-fluorescent dye) diffusion coefficient in PLGA. After the initial burst release, release of leuprolide from acid-capped PLGA microspheres appeared to be controlled initially by erosion and then by a second mechanism after day 21, which likely consists of a combination of peptide desorption and/or water-mediated breakage of pore connections. PBStc and acidic buffers accelerated degradation of PLGA and pore-network development and increased BODIPY diffusion coefficient, resulting in faster release. Release of leuprolide from the end-capped PLGA showed similar trends as found with acid capped PLGA but with a longer lag time before release. These data provide a baseline mechanistic signature of in vitro release of leuprolide for future comparison with corresponding in vivo performance, and in turn could lead to future development of rational in vitro-in vivo correlations.

Inhalable microspheres with hierarchical pore size for tuning the release of biotherapeutics in lungs

A series of customized inhalable mono-dispersed poly-L-lactide microspheres (PLA-MS) having hierarchical pore structures with either open or closed porosity were synthesized by double emulsion solvent evaporation method using various molar concentrations of porogens. We have presented a simple method for large scale, reproducible synthesis of hierarchically porous PLA-MS, controlling 3D morphologies with well-ordered porosity ranging from ∼2.5 to 850 nm. The prepared PLA-MS were characterized in terms of size, morphology, porosity, cellular uptake using particle size analysis, scanning electron microscopy, Brunauer-Emmett-Teller (BET) and confocal microscopy respectively. We explored, how tailoring the pore size and surface properties of micro-matrix would improve the aerodynamic properties, cellular uptake and release of the bio-therapeutic molecules of variable size. Porous PLA-MS with geometric size 5–10 μm and lower Mass Median Aerodynamic diameter (MMAD) (≤3 μm) shown to aerosolize well, evade macrophages phagocytosis and deposit in deep lungs. These results demonstrated that the type of porogens rather the concentration was more important in generating MS of variable porosity. The in-vitro release profiles of four model-biotherapeutics of significantly different molecular size, from MS of different porosities were investigated up to ∼6 days, their release patterns were found to be significantly different on a chronological basis. It is noteworthy observation that in-vitro cellular phagocytosis of MS is also governed by variable porosity of MS. Our findings shed light on the possible influence of size and porosity of MS on the lung deposition, cellular uptake and sustained delivery of a size-range of biotherapeutics. This approach of tuning porous structures of PLA-MS could be suitable to develop promising inhalable delivery systems.

Efficacy of Poly-Lactic-Co-Glycolic Acid Micro- and Nanoparticles of Ciprofloxacin Against Bacterial Biofilms

Bacterial biofilms are associated with a number of recurring infectious diseases and are a major cause for antibiotic resistance. Despite the broad use of polymeric microparticles and nanoparticles in biomedical research, it is not clear which particle size is more effective against biofilms. The purpose of this study was to evaluate the efficacy of sustained release poly-lactic-co-glycolic acid (PLGA) micro- and nanoparticles containing ciprofloxacin against biofilms of Staphylococcus aureus and Pseudomonas aeruginosa. The PLGA particles were prepared by the double emulsion solvent evaporation method. The resulting microparticles (12 μm) and nanoparticles (300 nm) contained drug loads of 7.3% and 4.5% (wt/wt) ciprofloxacin, respectively. Drug release was complete within 1 week following comparable release profiles for both particle sizes. Micro- and nanoparticles demonstrated a similar in vitro antibiofilm performance against mature P aeruginosa and S aureus with marked differences between the 2 strains. The sustained release of ciprofloxacin from micro- and nanoparticles over 6 days was equally effective as the continuous treatment with ciprofloxacin solution over the same period resulting in the eradication of culturable S aureus suggesting that reformulation of ciprofloxacin as sustained release PLGA micro- and nanoparticles might be valuable formulation approaches for the treatment of biofilms.

Intracellular delivery of peptide cargos using polyhydroxybutyrate based biodegradable nanoparticles: Studies on antitumor efficacy of BCL-2 converting peptide, NuBCP-9

Faster biodegradation, immunogenicity and lack of cell penetrative capabilities are hurdles in development of peptidyl drugs for cancer therapy. Polymeric carriers can be used to overcome these problems. The present study is focused on the use of polyhydroxybutyrate as a potential nanovehicle for the delivery of anticancer peptides. PHB (72kDa) was produced by thermal treatment of high molecular weight PHB (300kDa) under melt conditions and then conjugated with PEG (4kDa) by Steglich esterification reaction. Anticancer peptide NuBCP-9 (FSRSLHSLL) encapsulated PHB72K-PEG4K NPs were prepared by double emulsion solvent evaporation method. PHB72K-PEG4K NPs showed encapsulation efficiency of 61% and exhibited sustained release of peptide over a period of 26days at physiological pH. NuBCP-9 loaded PHB72K-PEG4K NPs showed an IC50 value of 2.2μM & 1.6μM in MCF-7 cells in 48h and 72h respectively. Confocal laser microscopy confirmed efficient cellular uptake and induction of apoptosis by peptide loaded NPs in a time dependent manner. In vivo intraperitonial administration of 20mg/kg NuBCP-9/NPs twice a week for three weeks triggered 90% tumor regression in Ehrlich syngeneic mouse model. Our results illustrated the potential of PHB72K-PEG4K based nanoformulation as a tool for targeting intracellular proteins.

Polymer nanoparticles for cross-presentation of exogenous antigens and enhanced cytotoxic T-lymphocyte immune response.

Effective induction of an antigen-specific cytotoxic T lymphocyte (CTL) immune response is one of the key goals of cancer immunotherapy. We report the design and fabrication of polyethylenimine (PEI)-coated polymer nanoparticles (NPs) as efficient antigen-delivery carriers that can induce antigen cross-presentation and a strong CTL response. After synthesis of poly(d,l-lactide-co-glycolide) (PLGA) NPs containing ovalbumin (OVA) by the double-emulsion solvent-evaporation method, cationic-charged PLGA NPs were generated by coating them with PEI. In a methyl tetrazolium salt assay, no discernible cytotoxic effect of PEI-coated PLGA (OVA) NPs was observed. The capacity and mechanism of PEI-coated PLGA (OVA) NPs for antigen delivery and cross-presentation on dendritic cells (DCs) were determined by fluorescence microscopy and flow cytometry. PEI-coated PLGA (OVA) NPs were internalized efficiently via phagocytosis or macropinocytosis in DCs and induced efficient cross-presentation of the antigen on MHC class I molecules via both endosome escape and a lysosomal processing mechanism. The DCs treated with PEI-coated PLGA (OVA) NPs induced a release of IL-2 cytokine from OVA-specific CD8-OVA1.3 T cells more efficiently than DCs treated with PLGA (OVA) NPs. Therefore, the PEI-coated PLGA (OVA) NPs can induce antigen cross-presentation and are expected to be used for induction of a strong CTL immune response and for efficient anticancer immunotherapy.

In vitro IFN-α release from IFN-α- and pegylated IFN-α-loaded poly(lactic-co-glycolic acid) and pegylated poly(lactic-co-glycolic acid) nanoparticles.

Interferon alpha (IFN-α) controlled release of nanoparticles was investigated under in vitro conditions. IFN-α and pegylated IFN-α (PEG-IFN-α) were encapsulated by poly(lactic-co-glycolic acid) (PLGA) and pegylated PLGA (PEG-PLGA) copolymers using double emulsion solvent evaporation method. The size of resulting four nanoparticles (IFN-α in poly(lactic-co-glycolic acids), IFN-α in poly(lactic-co-glycolic acid)-polyethylene glycol, PEG-IFN-α in poly(lactic-co-glycolic acids) and PEG-IFN-α in poly(lactic-co-glycolic acid)-polyethylene glycol) was below 130 nm diameter. IFN-α encapsulation efficiency of the nanoparticles was between 78 and 91%. The in vitro drug release studies conducted in phosphate-buffered saline and human plasma highlighted the role of incubation medium on the IFN release from the nanoparticles. The PEG-IFN-α in poly(lactic-co-glycolic acid)-polyethylene glycol was the most promising nanoparticle among the four formulations because of its remarkably constant release in both phosphate-buffered saline and plasma.

Abstract 1366: Fabrication of minocycline loaded PLGA microparticles for the treatment of intracranial tumors

Abstract Minocycline is a tetracycline derivative that was originally used clinically as an antibiotic and is currently being investigated as an anti-angiogenic factor for the treatment of different cancers including intracranial tumors such as glioblastomas. Due to its lipophilic nature, it has difficulty dissolving completely in organic or aqueous solvents and thus, presents a challenge for the fabrication of these particles via the emulsion-solvent evaporation method. The aim of this study is to investigate the effects of different methods of fabricating minocycline loaded microparticles on particle properties, such as the diameter, drug loading and release kinetics of the microparticles. Microparticles loaded with drug during the fabrication process were produced via two different methods: 1) an oil-in-water (O/W) single emulsion-solvent evaporation method where the drug was dissolved in the oil phase (denoted as “O/W particles”) and 2) a water-in-oil-in-water (W/O/W) double emulsion-solvent evaporation method where the drug was dissolved in the first water phase (denoted as “W/O/W particles”). Empty microparticles were also fabricated by a W/O/W method and loaded with drug after fabrication by dripping a drug solution onto freeze dried particles which were then left overnight at 4°C to allow the drug to adsorb onto and absorb into the particles (denoted as “prefabricated particles”). Light microscopy images of the particles were obtained and used to measure the diameters of the particles with the ImageJ software. The drug loading and release kinetics were determined by measuring the absorbance of minocycline at 350 nm with a microplate reader. The prefabricated particles resulted in larger diameters compared to the O/W and W/O/W particles which were loaded during the fabrication process. This could have resulted from the use of a vortex instead of a homogenizer/sonicator for the prefabricated scaffold which produces an emulsion with a lower speed compared to a homogenizer/sonicator. The prefabricated particles had a higher amount of drug loaded compared to O/W and W/O/W particles which were loaded during the fabrication process. By loading the drug after fabricating the particles, almost all of the drug can be adsorbed onto and/or absorbed into the particles without resulting in much loss of the drug. Alternatively, by loading the particles during the fabrication process, a lot of drug is lost into the water and/or oil phase and thus reducing the loading efficiencies of the particles. However, the particles loaded during the fabrication process were able to prolong the release of the loaded drug compared to the prefabricated scaffold which lost almost all of its drug by Day 1. In conclusion, although the prefabricated particles allow for the complete loading of the drug, the particles loaded during the fabrication process are more promising as controlled release vehicles as they are able to better sustain the release of the loaded drug. Citation Format: Sue Anne Chew, Marco Arriaga, Jesus R. Franco, Daniela Barbosa, Victor A. Hinojosa, Jose-Carlos Martinez, Paul Lenz. Fabrication of minocycline loaded PLGA microparticles for the treatment of intracranial tumors. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1366.

Preparation and characterization of Chinese yam polysaccharide PLGA nanoparticles and their immunological activity

This paper first provides that Chinese yam polysaccharide (CYP) is encapsulated by PLGA using a double emulsion solvent evaporation method and aims to screen the optimal preparation of CYP-PLGA nanoparticles (CYPP) using response surface methodology (RSM). The volume ratio of the internal water phase to the organic phase (W1:O), the volume ratio of the primary emulsion to the external water phase (PE:W2) and the concentration of Poloxamer 188 (F68) are deemed key variables for the encapsulation efficiency of CYPP. The results demonstrated that the data were accurately fitted into the RSM model. According to the RSM, the optimal scheme was a volume ratio of W1:O of 1:9, a volume ratio of PE: W2 of 1:10 and a concentration of F68 (W/V) of 0.7%. TEM and SEM images demonstrated that the nanoparticles had a spherical shape and smooth surface. The CYP and CYPP in vitro release studies demonstrated that the CYPP showed a release rate 53.41% lower than the release rate of CYP after 48h. The result of pro-proliferation and flow cytometry emerged that the CYPP were more effective compared with the free CYP and blank PLGA nanoparticles in promoting lymphocyte proliferation and triggering the transformation of T lymphocytes into Th cells.