Preclinical Assessment Addressing Intravenous Administration of a [68Ga]Ga-PSMA-617 Microemulsion: Acute In Vivo Toxicity, Tolerability, PET Imaging, and Biodistribution.

It has been herein presented that a microemulsion, known to be an effective and safe drug delivery system following intravenous administration, can be loaded with traces of [68Ga]Ga-PSMA-617 without losing its properties or causing toxicity. Following tolerated IV injections the capability of the microemulsion in altering [68Ga]Ga-PSMA-617 distribution was presented at 120 min post injection based on its ex vivo biodistribution results.

Spray-Dried, Nanoencapsulated, Multi-Drug Anti-Tuberculosis Therapy Aimed at Once Weekly Administration for the Duration of Treatment.

Aiming to improve the treatment outcomes of current daily tuberculosis (TB) chemotherapy over several months, we investigated whether nanoencapsulation of existing drugs would allow decreasing the treatment frequency to weekly, thereby ultimately improving patient compliance. Nanoencapsulation of three first-line anti-TB drugs was achieved by a unique, scalable spray-drying technology forming free-flowing powders in the nanometer range with encapsulation efficiencies of 82, 75, and 62% respectively for rifampicin, pyrazinamide, and isoniazid. In a pre-clinical study on TB infected mice, we demonstrate that the encapsulated drugs, administered once weekly for nine weeks, showed comparable efficacy to daily treatment with free drugs over the same experimental period. Both treatment approaches had equivalent outcomes for resolution of inflammation associated with the infection of lungs and spleens. These results demonstrate how scalable technology could be used to manufacture nanoencapsulated drugs. The formulations may be used to reduce the oral dose frequency from daily to once weekly in order to treat uncomplicated TB.

Preclinical assessment of 68 Ga-PSMA-617 entrapped in a microemulsion delivery system for applications in prostate cancer PET/CT imaging.

It has in recent years been reported that microemulsion (ME) delivery systems provide an opportunity to improve the efficacy of a therapeutic agent whilst minimising side effects and also offer the advantage of favourable treatment regimens. The prostate-specific membrane antigen (PSMA) targeting agents PSMA-11 and PSMA-617, which accumulate in prostate tumours, allow for [68 Ga]Ga3+ -radiolabelling and positron emission tomography/computed tomography (PET) imaging of PSMA expression in vivo. We herein report the formulation of [68 Ga]Ga-PSMA-617 into a ME ≤40 nm including its evaluation for improved cellular toxicity and in vivo biodistribution. The [68 Ga]Ga-PSMA-617-ME was tested in vitro for its cytotoxicity to HEK293 and PC3 cells. [68 Ga]Ga-PSMA-617-ME was administered intravenously in BALB/c mice followed by microPET/computed tomography (CT) imaging and ex vivo biodistribution determination. [68 Ga]Ga-PSMA-617-ME indicated negligible cellular toxicity at different concentrations. A statistically higher tolerance towards the [68 Ga]Ga-PSMA-617-ME occurred at 0.125 mg/mL by HEK293 cells compared with PC3 cells. The biodistribution in wild-type BALB/C mice showed the highest amounts of radioactivity (%ID/g) presented in the kidneys (31%) followed by the small intestine (10%) and stomach (9%); the lowest uptake was seen in the brain (0.5%). The incorporation of [68 Ga]Ga-PSMA-617 into ME was successfully demonstrated and resulted in a stable nontoxic formulation as evaluated by in vitro and in vivo means.

99mTc-MDP as an imaging tool to evaluate the in vivo biodistribution of solid lipid nanoparticles.

The aim of this study was to establish the in vivo uptake and tissue distribution of 99mTc-MDP-encapsulated Solid Lipid Nanoparticles (SLNs) post administration. Radioactive 99mTc-MDP encapsulated into SLNs was administered to rats to trace their biodistribution through imaging and ex vivo studies. As expected IV injected 99mTc-MDP exhibited predominant visual bone uptake and a high localisation of particles in the kidneys (3.87%ID/g) followed by bone (2.66%ID/g). IV administered 99mTc-MDP encapsulated by SLN showed similar uptake than 99mTc-MDP. Orally administrated 99mTc-MDP showed no uptake in any organs except the GI-tract while orally administered 99mTc-MDP-SLN showed distinct transfer of 99mTc-MDP from the GI tract with measurable levels in the kidneys and bone.

Supercritical antisolvent co-precipitation of rifampicin and ethyl cellulose.

Rifampicin-loaded submicron-sized particles were prepared through supercritical anti-solvent process using ethyl cellulose as polymeric encapsulating excipient. Ethyl acetate and a mixture of ethyl acetate/dimethyl sulfoxide (70/30 and 85/15) were used as solvents for both drug and polymeric excipient. When ethyl acetate was used, rifampicin was crystallized separately without being embedded within the ethyl cellulose matrix while by using the ethyl acetate/dimethyl sulfoxide mixture, reduced crystallinity of the active ingredient was observed and a simultaneous precipitation of ethyl cellulose and drug was achieved. The effect of solvent/CO2 molar ratio and polymer/drug mass ratio on the co-precipitates morphology and drug loading was investigated. Using the solvent mixture, co-precipitates with particle sizes ranging between 190 and 230nm were obtained with drug loading and drug precipitation yield from respectively 8.5 to 38.5 and 42.4 to 77.2% when decreasing the ethyl cellulose/rifampicin ratio. Results show that the solvent nature and the initial drug concentrations affect morphology and drug precipitation yield of the formulations. In vitro dissolution studies revealed that the release profile of rifampicin was sustained when co-precipitation was carried out with the solvent mixture. It was demonstrated that the drug to polymer ratio influenced amorphous content of the SAS co-precipitates. Differential scanning calorimetry thermograms and infrared spectra revealed that there is neither interaction between rifampicin and the polymer nor degradation of rifampicin during co-precipitation. In addition, stability stress tests on SAS co-precipitates were carried out at 75% relative humidity and room temperature in order to evaluate their physical stability. SAS co-precipitates were X-ray amorphous and remained stable after 6months of storage. The SAS co-precipitation process using a mixture of ethyl acetate/dimethyl sulfoxide demonstrates that this strategy can be successful for controlling rifampicin delivery.

Development, characterization and antimalarial efficacy of dihydroartemisinin loaded solid lipid nanoparticles.

Effective use of dihydroartemisinin (DHA) is limited by poor water-solubility, poor pharmacokinetic profile and unsatisfactory clinical outcome especially in monotherapy. To reduce such limitations, we reformulated DHA into solid lipid nanoparticles (SLNs) as a nanomedicine drug delivery system. DHA-SLNs were characterized for physical parameters and evaluated for in vitro and in vivo antimalarial efficacy. DHA-SLNs showed desirable particle characteristics including particle size (240.7 nm), particle surface charge (+17.0 mV), drug loadings (13.9 wt %), encapsulation efficacy (62.3%), polydispersity index (0.16) and a spherical appearance. Storage stability up to 90 days and sustained release of drug over 20 h was achieved. Enhanced in vitro (IC50 0.25 ng/ml) and in vivo (97.24% chemosuppression at 2mg/kg/day) antimalarial activity was observed. Enhancement in efficacy was 24% when compared to free DHA. These encouraging results show potential of using the described formulation for DHA drug delivery for clinical application. FROM THE CLINICAL EDITOR: Malaria still poses a significant problem worldwide. One of the current drugs, artemisinin has been shown to be effective, but has poor water-solubility. The authors here described their formulation of making dihydroartemisinin (DHA) into solid lipid nanoparticles, with subsequent enhancement in efficacy. These results would have massive potential in the clinical setting.

Micronization, characterization and in-vitro dissolution of shellac from PGSS supercritical CO2 technique.

The purpose of this investigation was to determine whether shellac, a naturally occurring material with enteric properties, could be processed in supercritical CO2 (sc-CO2) using the particles from gas saturated solution (PGSS) process and how process parameters affect the physico-chemical properties of shellac. In-situ attenuated total reflection fourier transform infra-red (ATR-FTIR) spectroscopy showed that CO2 dissolves in shellac with solubility reaching a maximum of 13% (w/w) at 300 bar pressure and 40 °C and maximum swelling of 28%. The solubility of sc-CO2 in shellac allowed for the formation of porous shellac structures of which the average pore diameter and pore density could be controlled by adjustment of operating pressure and temperature. In addition, it was possible to produce shellac microparticles ranging in average diameter from 180 to 300 µm. It was also shown that processing shellac in sc-CO2 resulted in accelerated esterification reactions, potentially limiting the extent of post-processing "ageing" and thus greater stability. Due to additional hydrolysis reactions enhanced by the presence of sc-CO2, the solubility of shellac at pH 7.5 was increased by between 4 and 7 times, while dissolution rates were also increased. It was also shown that the in-vitro dissolution profiles of shellac could be modified by slight adjustment in operating temperatures.

Mycolic acids, a promising mycobacterial ligand for targeting of nanoencapsulated drugs in tuberculosis.

The appearance of drug-resistant strains of Mycobacterium tuberculosis (Mtb) poses a great challenge to the development of novel treatment programmes to combat tuberculosis. Since innovative nanotechnologies might alleviate the limitations of current therapies, we have designed a new nanoformulation for use as an anti-TB drug delivery system. It consists of incorporating mycobacterial cell wall mycolic acids (MA) as targeting ligands into a drug-encapsulating Poly dl-lactic-co-glycolic acid polymer (PLGA), via a double emulsion solvent evaporation technique. Bone marrow-derived mouse macrophages, either uninfected or infected with different mycobacterial strains (Mycobacterium avium, Mycobacterium bovis BCG or Mtb), were exposed to encapsulated isoniazid-PLGA nanoparticles (NPs) using MA as a targeting ligand. The fate of the NPs was monitored by electron microscopy. Our study showed that i) the inclusion of MA in the nanoformulations resulted in their expression on the outer surface and a significant increase in phagocytic uptake of the NPs; ii) nanoparticle-containing phagosomes were rapidly processed into phagolysosomes, whether MA had been included or not; and iii) nanoparticle-containing phagolysosomes did not fuse with non-matured mycobacterium-containing phagosomes, but fusion events with mycobacterium-containing phagolysosomes were clearly observed.

Oral lipid-based nanoformulation of tafenoquine enhanced bioavailability and blood stage antimalarial efficacy and led to a reduction in human red blood cell loss in mice.

Tafenoquine (TQ), a new synthetic analog of primaquine, has relatively poor bioavailability and associated toxicity in glucose-6-phosphate dehydrogenase (G6PD)-deficient individuals. A microemulsion formulation of TQ (MTQ) with sizes <20 nm improved the solubility of TQ and enhanced the oral bioavailability from 55% to 99% in healthy mice (area under the curve 0 to infinity: 11,368±1,232 and 23,842±872 min·µmol/L) for reference TQ and MTQ, respectively. Average parasitemia in Plasmodium berghei-infected mice was four- to tenfold lower in the MTQ-treated group. In vitro antiplasmodial activities against chloroquine-sensitive and chloroquine-resistant strains of Plasmodium falciparum indicated no change in half maximal inhibitory concentration, suggesting that the microemulsion did not affect the inherent activity of TQ. In a humanized mouse model of G6PD deficiency, we observed reduction in toxicity of TQ as delivered by MTQ at low but efficacious concentrations of TQ. We hereby report an enhancement in the solubility, bioavailibility, and efficacy of TQ against blood stages of Plasmodium parasites without a corresponding increase in toxicity.

Human food safety and environmental hazards associated with the use of methyltestosterone and other steroids in production of all-male tilapia.

In recent years, all-male cultures of Nile tilapia (Oreochromis niloticus) have been the most preferred mode of production in aquaculture industry. All-male individuals achieve higher somatic growth rate and shut high energy losses associated with gonadal development and reproduction. The economic advantages of culturing all-male tilapia have led to the development of procedures for producing unisex cultures, using 17α-methyltestosterone (MT). Despite widespread use of the MT in tilapia farming, the implications of hormone treatment in relation to human health and the environment have raised a number of concerns in the scientific community. In this review, the hormonal application processes, economic and ecological significance of MT, food safety and residual MT, comparative uses of steroids in aquaculture, animal husbandry, and medicine have been briefly reviewed for regulatory guidelines, and finally, future research perspectives have been addressed. The review can be used as policy-making guidelines in aquaculture framework development as can be emphasized in African continent, among others. The most important conclusion to draw is that the quantity of MT used in conventional practice is large compared to the actual dose required for sex reversal, fish produced are safe for human consumptions, and the environmental hazards should be further emphasized.

Effects of spray-drying on w/o/w multiple emulsions prepared from a stearic acid matrix.

The goal of this study was to explore the effects of spray-drying on w/o/w double emulsions of methyltestosterone (MT) loaded in a stearic acid matrix. MT-loaded nanoparticles were formulated by a water-in-oil-in-water emulsion technique using 50, 75, and 100 mg of stearic acid, 2% and 3% w/v polyvinyl alcohol, 5% w/v lactose, and 0.2% w/v chitosan. The emulsions were immediately spray-dried based on an optimized model of inlet temperature and pump rate, and characterized for optimized responses with regard to particle size, polydispersity index, and zeta potential, for both emulsion and powder samples. Dynamic light scattering analysis shown that the nanoparticles increased in size with increasing concentrations of polyvinyl alcohol and stearic acid. Scanning electron microscopy indicated that the MT-loaded nanoparticles were spherical in shape, had a smooth surface, and were in an amorphous state, which was confirmed by differential scanning calorimetry. These MT-loaded nanoparticles are a promising candidate carrier for the delivery of MT; however, further studies are needed in order to establish the stability of the system and the cargo release profile under normal conditions of use.

Preparation, characterization, and optimization of primaquine-loaded solid lipid nanoparticles.

Primaquine (PQ) is one of the most widely used antimalarial drugs and is the only available drug that combats the relapsing form of malaria. PQ use in higher doses is limited by severe tissue toxicity including hematological- and gastrointestinal-related side effects. Nanoformulation of drugs in an appropriate drug carrier system has been extensively studied and shown to have the potential to improve bioavailability, thereby enhancing activity, reducing dose frequency, and subsequently reducing toxicity. The aim of this work was to design, synthesize, and characterize PQ-loaded solid lipid nanoparticles (SLNs) (PQ-SLNs) as a potential drug-delivery system. SLNs were prepared by a modified solvent emulsification evaporation method based on a water-in-oil-in-water (w/o/w) double emulsion. The mean particle size, zeta potential, drug loading, and encapsulation efficiency of the PQ-SLNs were 236 nm, +23 mV, 14%, and 75%, respectively. The zeta potential of the SLNs changed dramatically, from -6.54 mV to +23.0 mV, by binding positively charged chitosan as surface modifier. A spherical morphology of PQ-SLNs was seen by scanning electron microscope. In vitro, release profile depicted a steady drug release over 72 hours. Differential scanning calorimeter thermograms demonstrated presence of drug in drug-loaded nanoparticles along with disappearance of decomposition exotherms, suggesting increased physical stability of drug in prepared formulations. Negligible changes in characteristic peaks of drug in Fourier transform infrared spectra indicated absence of any interaction among the various components entrapped in the nanoparticle formulation. The nanoformulated PQ was 20% more effective as compared with conventional oral dose when tested in Plasmodium berghei-infected Swiss albino mice. This study demonstrated an efficient method of forming a nanomedicine delivery system for antimalarial drugs.

Enhanced oral bioavailability of the antiretroviral efavirenz encapsulated in poly(epsilon-caprolactone) nanoparticles by a spray-drying method.

AIM: To encapsulate efavirenz (EFV) within poly(epsilon-caprolactone) (PCL) nanoparticles (NPs) and compare the oral pharmacokinetics with that of EFV-loaded micelles and pure EFV NPs. MATERIALS & METHODS: EFV-loaded PCL NPs were produced by a double-emulsion/spray-drying method. RESULTS: NPs displayed a hydrodynamic diameter of 200-250 nm. The encapsulation efficiency was 86-93% and the mass recovery was above 60%. X-ray diffraction indicated that drug and PCL underwent amorphization during the spray-drying process. Encapsulation within NPs significantly increased the maximum concentration in plasma and the bioavailability. CONCLUSION: EFV-loaded PCL NPs represent a promising platform to develop scalable pharmaceuticals with improved (bio)pharmaceutic performance. Original submitted 2 May 2013; Revised submitted 4 September 2013.

State of the art and future directions in nanomedicine for tuberculosis.

INTRODUCTION: Tuberculosis (TB) ranks the second leading cause of death from an infectious disease worldwide. However, treatment of TB is affected by poor patient compliance due to the requirement for daily drug administration, for lengthy periods of time, often with severe drug-induced side effects. Nanomedicines have the potential to improve treatment outcomes by providing therapies with reduced drug doses, administered less frequently, under shortened treatment durations. AREAS COVERED: In this article, we present the pathophysiology of the disease, focusing on pulmonary TB and the characteristics of drugs used in treatment and discuss the application of nanomedicines within this scope. We also discuss new formulation approaches for TB nanomedicines and directions for future research. EXPERT OPINION: Nanomedicines have the potential to improve TB treatment outcomes. New approaches such as nanoparticle systems able to impact the immune response of macrophages and deliver drug intracellularly, as well as the use of polymer-drug conjugates for drug delivery, are likely to play an important role in TB nanomedicines in future. However, further research is required before TB nanomedicines can be translated to the clinic.

Permeation of PLGA nanoparticles across different in vitro models.

Many drug delivery systems have indicated improvement in delivery of various drug molecules and among these biodegradable and biocompatible polymers such as poly(D,L-lactide-co-glycolide) (PLGA) have been shown to enhance intracellular uptake of drug candidates when formulated as nanoparticles. PLGA nanoparticles were prepared by means of a double emulsion solvent evaporation technique and evaluated in terms of size, encapsulation efficiency, surface charge, isoniazid release and in vitro transport. The nanoparticles have an average size of 237 nm and were previously shown to be distributed in several tissues after oral administration without triggering an immune response. This study focussed on the in vitro permeation of the PLGA nanoparticles across different membranes and showed that although Rhodamine 6G-labelled nanoparticles are efficiently delivered across the intestinal epithelium, its epithelial permeability changes when a drug such as isoniazid is encapsulated. Future studies should focus on ways to optimise PLGA nanoparticle delivery when a drug such as isoniazid is encapsulated for instance by coating with polymers such as polyethylene glycol.

Effects of protein binding on the biodistribution of PEGylated PLGA nanoparticles post oral administration.

The surface of nanoparticles is often functionalised with polymeric surfactants, in order to increase systemic circulation time. This has been investigated mainly for intravenously administered nanoparticles. This study aims to elucidate the effect of surface coating with various concentrations of polymeric surfactants (PEG and Pluronics F127) on the in vitro protein binding as well as the tissue biodistribution, post oral administration, of PLGA nanoparticles. The in vitro protein binding varied depending on the polymeric surfactant used. However, in vivo, 1% PEG and 1% Pluronics F127 coated particles presented similar biodistribution profiles in various tissues over seven days. Furthermore, the percentage of PEG and Pluronics coated particles detected in plasma was higher than that of uncoated PLGA particles, indicating that systemic circulation time can also be increased with oral formulations. The difference in the in vitro protein binding as a result of the different poloxamers used versus similar in vivo profiles of these particles indicates that in vitro observations for nanoparticles cannot represent or be correlated to the in vivo behaviour of the nanoparticles. Our results therefore suggest that more studies have to be conducted for oral formulations to give a better understanding of the kinetics of the particles.

In vivo evaluation of the biodistribution and safety of PLGA nanoparticles as drug delivery systems.

The remarkable physicochemical properties of particles in the nanometer range have been proven to address many challenges in the field of science. However, the possible toxic effects of these particles have raised some concerns. The aim of this article is to evaluate the effects of poly(lactide-co-glycolide) (PLGA) nanoparticles in vitro and in vivo compared to industrial nanoparticles of a similar size range such as zinc oxide, ferrous oxide, and fumed silica. An in vitro cytotoxicity study was conducted to assess the cell viability following exposure to PLGA nanoparticles. Viability was determined by means of a WST assay, wherein cell viability of greater than 75% was observed for both PLGA and amorphous fumed silica particles and ferrous oxide, but was significantly reduced for zinc oxide particles. In vivo toxicity assays were performed via histopathological evaluation, and no specific anatomical pathological changes or tissue damage was observed in the tissues of Balb/C mice. The extent of tissue distribution and retention following oral administration of PLGA particles was analyzed for 7 days. After 7 days, the particles remained detectable in the brain, heart, kidney, liver, lungs, and spleen. The results show that a mean percentage (40.04%) of the particles were localized in the liver, 25.97% in the kidney, and 12.86% in the brain. The lowest percentage was observed in the spleen. Thus, based on these assays, it can be concluded that the toxic effects observed with various industrial nanoparticles will not be observed with particles made of synthetic polymers such as PLGA when applied in the field of nanomedicine. Furthermore, the biodistribution of the particles warrants surface modification of the particles to avoid higher particle localization in the liver. FROM THE CLINICAL EDITOR: The aim of this study was to evaluate the effects of poly(lactide-co-glycolide) (PLGA) nanoparticles in vitro and in vivo compared to industrial nanoparticles including zinc oxide, ferrous oxide, and fumed silica. The authors concluded that the toxic effects observed with various industrial nanoparticles is unlikely to be observed with particles made of PLGA. The biodistribution of these particles warrants surface modification to avoid particle accumulation in the liver.

Nanomedicine for respiratory diseases.

Treatment of respiratory diseases and infections has proved to be a challenging task, with the incidence of these ailments increasing worldwide. Nanotechnology-based drug and gene delivery systems offer a possible solution to some of the shortfalls of the current treatment regimen. Nanobased drug delivery systems have revolutionised the field of pharmacotherapy by presenting the ability to alter the pharmacokinetics of the conventional drugs to extend the drug retention time, reduce the toxicity and increase the half-life of the drugs. Delivery of exogenous genes to the airway epithelium in vivo has been limited by several physiological barriers, resulting in the low success rate of these systems. With the advent of nanotechnology, DNA compacted with cationic polymers to produce nanoparticles has exhibited a significant increase in the transfection efficiencies. With nanoparticulate drug/gene delivery systems, specific cells can be targeted by functionalising the polymeric nanoparticles with ligands that allow the particles to dock at a specific site of the cell. In addition, polymeric systems allow for the cargo to be released in a controlled and stimuli-responsive manner. The advantages that nanoparticulate delivery systems present in the treatment of respiratory diseases and infections are summarised in this review.