Murine airway histology and intracellular uptake of inhaled amorphous itraconazole.

Aerosolization of amorphous itraconazole may be a safe and effective method of pulmonary delivery. Our objective was to evaluate the histologic effects, immunogenic potential, and cellular uptake of aerosolized amorphous itraconazole. Mice received amorphous itraconazole (30mg/kg), excipient placebo, or saline control by nebulization every 12h for up to 12 days. Broncho-alveolar lavage (BAL) and formalin fixation of both lungs were conducted. BAL supernatant was assayed for IL-12 by ELISA, and cellular components were analyzed by high performance liquid chromatography-mass spectroscopy. Coronal sections of the entire lung were stained, viewed by light microscopy, and the Cimolai histopathologic inflammatory score was obtained for each lobe. No evidence of bronchiolar, peribronchiolar or perivascular inflammation was found in any treatment group, nor were epithelial ulceration or repair observed. The Cimolai histopathologic scores for amorphous itraconazole, excipient, and saline control on days 3 and 8 did not differ between groups. ELISA analysis showed no cytokine induction of IL-12. Itraconazole was detected within cells collected from BAL fluid on days 1, 3, 8 and 12. Aerosolized administration of amorphous itraconazole or excipients does not cause inflammation or changes in pulmonary histology and are not associated with pro-inflammatory cytokine production.

Single dose and multiple dose studies of itraconazole nanoparticles.

The objective of this study was to determine and compare the lung and serum concentrations in mice following oral and pulmonary dosing of amorphous nanoparticulate itraconazole (ITZ) compositions as well as the Sporanox oral solution (itraconazole/Janssen). Second, the steady state partitioning of ITZ in lung tissue and circulatory compartments following repeated oral and pulmonary dosing was determined. The pulmonary formulation (ITZ-pulmonary) consisted of ITZ, polysorbate 80, and poloxamer 407 in a 1:0.75:0.75 ratio and the oral formulation (ITZ-oral) consisted of ITZ, PEG 8000, poloxamer 188, and sorbitan monooleate 80 in a 1:1:2:1 ratio. Mice were dosed every 12 h by nebulization with ITZ-pulmonary, or by oral gavage with ITZ-oral or Sporanox oral solution (n = 12 per study arm). ITZ-pulmonary achieved significantly greater (>10-fold) lung tissue concentrations compared to the Sporanox oral solution and ITZ-oral. There were no statistical differences between the two oral formulations. ITZ-pulmonary achieved significantly greater lung levels per unit serum concentration compared to the orally dosed ITZ compositions. High and sustained lung tissue concentrations were achieved via inhalation of an amorphous nanoparticulate ITZ-pulmonary composition while maintaining serum levels which are above the minimum lethal concentration (MLC) of Aspergillus fumigatus.

Cryogenic liquids, nanoparticles, and microencapsulation.

The biopharmaceutical classification system (BCS) is used to group pharmaceutical actives depending upon the solubility and permeability characteristics of the drug. BCS class II compounds are poorly soluble but highly permeable, exhibiting bioavailability that is limited by dissolution. The dissolution rate of BCS class II drug substances may be accelerated by enhancing the wetting of the bulk powder and by reducing the primary particle size of the drug to increase the surface area. These goals may be achieved by nucleating drug particles from solution in the presence of stabilizing excipients. In the spray freezing into liquid (SFL) process, a drug containing solution is atomized and frozen rapidly to engineer porous amorphous drug/excipient particles with high surface areas and dissolution rates. Aqueous suspensions of nanostructured particles may be produced from organic solutions by evaporative precipitation into aqueous solution (EPAS). The suspensions may be dried by lyophilization. The particle size and morphology may be controlled by the type and level of stabilizers. In vivo studies have shown increased bioavailability of a wide variety of drugs particles formed by SFL or EPAS. For both processes, increased serum levels of danazol (DAN) were observed in mice relative to bulk DAN and the commercial product, Danocrine. Orally dosed itraconazole (ITZ) compositions, formed by SFL, produce higher serum levels of the drug compared to the commercial product, Sporanox oral solution. Additionally, nebulized SFL processed ITZ particles suspended in normal saline have been dosed via the pulmonary route and led to extended survival times for mice inoculated with Aspergillis flavus. SFL and EPAS processes produce amorphous drug particles with increased wetting and dissolution rates, which will subsequently supersaturate biological fluids in vivo, resulting in increased drug bioavailability and efficacy.

Comparison of powder produced by evaporative precipitation into aqueous solution (EPAS) and spray freezing into liquid (SFL) technologies using novel Z-contrast STEM and complimentary techniques.

The objective of this study was to compare the properties of particles formed by nucleation and polymer stabilization (e.g. evaporative precipitation into aqueous solution (EPAS)) versus rapid freezing (e.g. spray freezing into liquid (SFL)). Powders formed by EPAS and SFL, composed of danazol and PVP K-15 in a 1:1 ratio, were characterized using X-ray powder diffraction, modulated differential scanning calorimetry (MDSC), contact angle determination, dissolution, scanning electron microscopy (SEM), environmental scanning electron microscopy (ESEM), BET specific surface area, and Z-contrast scanning transmission electron microscopy (STEM). Large differences in particle morphologies and properties were observed and explained in terms of the particle formation mechanisms. Both techniques produced amorphous powders with high T(g) and low contact angle values. However, STEM analysis showed highly porous bicontinuous nanostructured 30nm particles connected by narrow bridges for SFL versus aggregated 500 nm primary particles for EPAS. The combination of STEM and other characterization techniques indicates solid solutions were formed for the SFL powders consistent with rapid freezing. In contrast, the EPAS particle cores are enriched in hydrophobic API and the outer surface is enriched in the hydrophilic polymer, with less miscibility than in the SFL powders. Consequently, dissolution rates are faster for the SFL particles, although both techniques enhanced dissolution rates of the API.

Rapid dissolution of high-potency danazol particles produced by evaporative precipitation into aqueous solution.

High-potency danazol particles with high dissolution rates were produced by evaporative precipitation into aqueous solution (EPAS). Aqueous suspensions formed by EPAS were centrifuged to remove the nonadsorbed surfactant. The resulting surfactant-coated drug particles had extremely high drug-to-surfactant ratios greater than 5, corresponding to potencies (wt drug/wt drug + wt surfactant) as high as 93%. The mechanism of the high dissolution rates was characterized as a function of surfactant adsorption, particle size and surface area, drug crystallinity, and the contact angle for water on the drug surface. For danazol stabilized by polyvinyl pyrrolidone (PVP) alone or with sodium lauryl sulfate (SLS), small particle diameter and high surface area led to high dissolution rates with approximately 90% drug dissolved in 2 min. The crystallinity of the danazol was typically 80%. The properties of the particles and the dissolution rates were mostly unchanged under a 2-week thermal cycling stress test.

Supersaturation produces high bioavailability of amorphous danazol particles formed by evaporative precipitation into aqueous solution and spray freezing into liquid technologies.

The bioavailability of high surface area danazol formulations was evaluated in a mouse model to determine what effect high supersaturation, as measured in vitro, has on the absorption of a poorly water soluble drug. Danazol, a biopharmaceutics classification system II (BCS II) compound, was used as the model drug. Evaporative precipitation into aqueous solution (EPAS) and spray freezing into liquid (SFL) technologies were used to prepare powders of danazol/PVP K-15 in a 1:1 ratio. The evaporative precipitation into aqueous solution (EPAS) and SFL compositions, physical mixture and commercial product were dosed by oral gavage to 28 male Swiss/ICR mice for each arm of the study. Samples were taken at time points ranging from 0.5 to 24 h. Pooled mouse serum was analyzed for danazol by high performance liquid chromatography (HPLC). Powders were analyzed for their ability to form supersaturated solutions through dissolution at concentrations of 1 mg/mL which was the dose delivered to the mouse models. Spray freezing into liquid (SFL) and EPAS compositions displayed higher C(max) at 392.5 ng/mL and 430.1 ng/mL, respectively, compared to the physical mixture (204.4 ng/mL) and commercially available danazol (199.3 ng/mL). The T(max) for all compositions studied was near the 1 h time point. The area under the curve (AUC) for the SFL composition was 2558 ng.h/mL compared to EPAS composition at 1534 ng.h/mL. The area under the curve (AUC) for the physical mixture and commercially available danazol were 672 ng.h/mL and 1519 ng.h/mL, respectively. The elimination rate constants for the EPAS composition, SFL composition, and physical mixture were similar at approximately 0.15 h(-1) where as the commercially available danazol capsules displayed an elimination rate constant of 0.103 h(-1). The extent of danazol absorption in the mouse model was higher for SFL composition compared to the less amorphous EPAS composition, physical mixture, and commercially available danazol powders. Both EPAS and SFL compositions were able to form supersaturated solutions. However, the SFL composition displayed a supersaturation of 33% above control and was able to maintain supersaturation for 90 min compared to the EPAS composition (27% supersaturation above control for 60 min). Through the use of a testing method for supersaturation, it was found that EPAS and SFL compositions achieve higher apparent solubilities when compared to the physical mixture and commercially available danazol capsules. Because of the greater extent of dissolution of the SFL composition, the bioavailability was enhanced in a mouse model.

Targeted high lung concentrations of itraconazole using nebulized dispersions in a murine model.

PURPOSE: The purpose of this study was to investigate the delivery of itraconazole (ITZ) particles to a murine lung model by nebulization. METHODS: Three ITZ formulations were prepared and characterized in the dry state using contact angle, dissolution, X-ray powder diffraction, scanning electron microscopy, and Brunauer-Emmett-Teller surface area analysis. Aerodynamic particle size distributions and lung deposition studies in 14 outbred male ICR mice were performed using aqueous dispersions of all the formulations. A separate dosing uniformity study was also performed to qualify use of the chamber. RESULTS: All formulations had an aggregated particle size of approximately 30 microm in diameter. Two formulations showed that 80% of the drug dissolved in less than 5 min. The remaining ITZ formulation had a slower dissolution and the lowest total emitted dose from the nebulizer used. High concentrations of ITZ were shown to be present in the mouse lung during the lung deposition study, up to 16.8 +/- 0.13 microg/g (+/- SE) were achieved. Concentrations of up to 0.76 +/- 0.03 microg/g (+/- SE) could be maintained from the single nebulized dose for at least 24 h. CONCLUSION: An effective method of targeted delivery of ITZ to the deep lung is presented that may be useful for the treatment and prevention of acute fungal infections.