RESUMEN
The incidence of fungal pulmonary infections is known to be on the increase, and yet there is an alarming gap in terms of marketed antifungal therapies that are available for pulmonary administration. Amphotericin B (AmB) is a highly efficient broad-spectrum antifungal only marketed as an intravenous formulation. Based on the lack of effective antifungal and antiparasitic pulmonary treatments, the aim of this study was to develop a carbohydrate-based AmB dry powder inhaler (DPI) formulation, prepared by spray drying. Amorphous AmB microparticles were developed by combining 39.7 % AmB with 39.7 % γ-cyclodextrin, 8.1 % mannose and 12.5 % leucine. An increase in the mannose concentration from 8.1 to 29.8 %, led to partial drug crystallisation. Both formulations showed good in vitro lung deposition characteristics (80 % FPF < 5 µm and MMAD < 3 µm) at different air flow rates (60 and 30 L/min) when used with a DPI, but also during nebulisation upon reconstitution in water.
Asunto(s)
Anfotericina B , Neumonía , Humanos , Anfotericina B/farmacología , Antifúngicos/farmacología , Inhaladores de Polvo Seco , Manosa , Pulmón , Macrófagos Alveolares , Administración por Inhalación , Tamaño de la Partícula , Polvos/farmacología , Aerosoles y Gotitas RespiratoriasRESUMEN
Pulmonary delivery of fluoroquinolones (FQs) is an interesting approach to treat lung infections as it may lead to high local concentrations while minimizing systemic exposure. However, FQs have a rapid diffusion through the lung epithelium giving the pulmonary route no advantage compared to the oral route. Interactions between FQs and metal cations form complexes which limit the diffusion through the epithelial barrier and would reduce the absorption of FQs and maintain high concentrations in the lung. The effects of this complexation depend on the FQ and the metal cations and optimum partners should be selected through in vitro experiments prior to aerosol drug formulation. In this study, CIP was chosen as a representative FQ and 5 cations (Ca2+, Mg2+, Zn2+, Al3+, Cu2+) were selected to study the complexation and its effects on permeability, antimicrobial efficacy and cell toxicity. The results showed that the apparent association constants between CIP and cations ranked with the descending order: Cu2+>Al3+>Zn2+>Mg2+>Ca2+. When a target of 80% complexation was reached with the adequate concentrations of cations, the CIP permeability through the Calu-3 lung epithelial cells was decreased of 50%. Toxicity of the CIP on the Calu-3 cells, with an EC50 evaluated at 7µM, was not significantly affected by the presence of the cations. The minimum inhibitory concentration of CIP for Pseudomonas aeruginosa was not affected or slightly increased in the range of cation concentrations tested, except for Mg2+. In conclusion, permeability was the main parameter that was affected by the metal cation complexation while cell toxicity and antimicrobial activity were not or slightly modified. Cu2+, with the highest apparent constant of association and with no effect on cell toxicity and antimicrobial activity of the CIP, appeared as a promising cation for the development of a controlled-permeability formulation of CIP for lung treatment.
Asunto(s)
Antibacterianos/química , Ciprofloxacina/química , Complejos de Coordinación/química , Mucosa Respiratoria/efectos de los fármacos , Antibacterianos/administración & dosificación , Cationes , Línea Celular , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Ciprofloxacina/administración & dosificación , Complejos de Coordinación/administración & dosificación , Relación Dosis-Respuesta a Droga , Evaluación Preclínica de Medicamentos/métodos , Humanos , Pseudomonas aeruginosa/efectos de los fármacos , Pseudomonas aeruginosa/fisiología , Mucosa Respiratoria/fisiologíaRESUMEN
Pulmonary administration of protein and peptide drugs using inhaled dry powder particles is an interesting alternative to parenteral delivery. The stabilisation of these molecules is essential to the maintenance of biological activity in such inhalation formulations. Here salmon calcitonin (sCT) was co-spray dried with linear or branched PEG (L-PEG and B-PEG) and PVP in order to formulate aerosolisable particles of the bioactive peptide. Co-spray drying L-PEG and PVP resulted in porous particles, with minimal D(50) (median volume diameter) and MMAD (mass median aerodynamic diameter) values obtained for a PEG/PVP w/w ratio of 1. For particles based on both L-PEG and B-PEG, an increase in acetone, a poor solvent for the PVP, up to 70wt% of the spray dried solution led to a decrease in D(50) and MMAD. Crystallinity of PEG in the particles ranged between 90 and 97% when the PVP content varied between 15 and 70wt%, indicating a low degree of interaction between PVP and PEG. Additionally, dynamic vapour sorption analysis showed that an increase in PVP content increased the particle surface hygroscopicity. Hence, particle properties were adjusted by altering the water/acetone and PEG/PVP ratio in the spray dried solutions. PVP present at the particles surface protects them from melting during the spray drying process but also increases their hygroscopicity, adversely affecting their aerodynamic properties. Targeting a 5wt% of sCT loading resulted in a loading efficiency of 77.9 and 83.6% with L-PEG and B-PEG-based particles, respectively. Loading of sCT in L-PEG or B-PEG-based particles modified particle roughness and D(50), leading to an increase in MMAD of the L-PEG-based particles. However, particles were still considered to be suitable for aerosolisation as their FPFs (fine particle fractions) were higher than 30%. These particles formulated with PVP and PEG allowed sCT biological activity to be maintained when evaluated by measuring cAMP production by T47D cells.
Asunto(s)
Pulmón/metabolismo , Polímeros/química , Rastreo Diferencial de Calorimetría , Tamaño de la Partícula , Polímeros/administración & dosificación , Solubilidad , Tensión Superficial , Termogravimetría , Difracción de Rayos XRESUMEN
This study describes how the control of doxorubicin (DOX) polarity allows to encapsulate it inside poly(lactide-co-glycolide) (PLGA) nanoparticles formulated either by a single oil-in-water (O/W) or a double water-in-oil-in-water (W/O/W) emulsification method (SE and DE, respectively). DOX is commercially available as a water soluble hydrochloride salt, which is useful for DE. The main difficulty related to DE approach is that the low affinity of hydrophilic drugs to the polymer limits entrapment efficiency. Compared to DE method, SE protocol is easier and should provide an additional gain in entrapment efficiency. To be encapsulated by SE technique, DOX should be used in a more lipophilic molecular form. We evaluated the lipophilicity of DOX in terms of apparent partition coefficient (P) and modulated it by adjusting the pH of the aqueous phase. The highest P values were obtained at pH ranging from 8.6 to 9, i. e. between two DOX pK(a) values (8.2 and 9.6). The conditions favorable for the drug lipophilicity were then used to formulate DOX-loaded PLGA nanoparticles by SE method. DOX encapsulation efficiency as well as release profiles were evaluated for these nanoparticles and compared to those with nanoparticles formulated by DE. Our results indicate that the encapsulation of DOX in nanoparticles formulated by SE provides an increased drug entrapment efficiency and decreases the burst effect.
Asunto(s)
Doxorrubicina/administración & dosificación , Emulsiones , Ácido Láctico/administración & dosificación , Nanopartículas , Ácido Poliglicólico/administración & dosificación , Polímeros/administración & dosificación , Tecnología Farmacéutica/métodos , Química Farmacéutica , Doxorrubicina/química , Copolímero de Ácido Poliláctico-Ácido Poliglicólico , SolubilidadRESUMEN
A "market map" comparison methodology for cigarette smoke chemistry yields is presented. Federal Trade Commission machine-method smoke chemistry was determined for a range of filtered cigarettes from the US marketplace. These data were used to develop illustrative market maps for each smoke constituent as analytical tools for comparing new or non-conventional cigarettes to a sampling of the broader range of marketplace cigarettes. Each market map contained best-estimate "market-means," showing the relationship between commercial cigarette constituent and tar yields, and yield "market ranges" defined by prediction intervals. These market map means and ranges are the basis for comparing new cigarette smoke yields to those of conventional cigarettes. The potential utility of market maps for evaluating differences in smoke chemistry was demonstrated with 1R4F and 2R4F Kentucky reference cigarettes, an Accord cigarette, and an Advance cigarette. Conventional cigarette tobacco nicotine, nitrate, soluble ammonia, and tobacco specific nitrosamine levels are reported. Differences among conventional cigarette constituent yields at similar tar levels were explained in part by the chemical composition range of those cigarette tobaccos. The study also included a comparison of smoke constituent yields and in vitro smoke cytotoxicity and mutagenicity assay results for the 1R4F Kentucky reference cigarette and its replacement 2R4F. Significant smoke yield differences were noted for lead, NNK, and NNN. The majority of their smoke constituent yields were within the market range developed from the sampled conventional cigarettes. Within the sensitivity and specificity of the in vitro bioassays used, smoke toxic activity differences for the two reference cigarettes were not statistically significant. These results add to the limited information available for the 2R4F reference cigarette.
Asunto(s)
Nicotiana/química , Humo/efectos adversos , Humo/análisis , Animales , Línea Celular , Embrión de Mamíferos/efectos de los fármacos , Plomo/análisis , Mercadotecnía , Ratones , Pruebas de Mutagenicidad , Nitrosaminas/análisis , Valor Predictivo de las Pruebas , Estándares de Referencia , Valores de Referencia , Análisis de Regresión , Breas/química , Breas/normas , Nicotiana/clasificación , Nicotiana/toxicidad , Pruebas de Toxicidad , Estados UnidosRESUMEN
The mouse lymphoma thymidine kinase assay (MLA) has been optimized to quantitatively determine the in vitro mutagenicity of cigarette mainstream smoke particulate phase. To test whether the MLA is able to discriminate between different cigarette types, specially constructed cigarettes each containing a single tobacco type - Bright, Burley, or Oriental - were investigated. The mutagenic activity of the Burley cigarette was statistically significantly lower, up to approximately 40%, than that of the Bright and Oriental cigarettes. To determine the impact of two different sets of smoking conditions, American-blend cigarettes were smoked under US Federal Trade Commission/International Organisation for Standardisation conditions and under Massachusetts Department of Public Health (MDPH) conditions. Conventional cigarettes - eight from the US commercial market plus the Reference Cigarettes 1R4F and 2R4F - and an electrically heated cigarette smoking system (EHCSS) prototype were tested. There were no statistically significant differences between the two sets of smoking conditions on a per mg total particulate matter basis, although there was a consistent trend towards slightly lower mutagenic activity under MDPH conditions. The mutagenic activity of the EHCSS prototype was distinctly lower than that of the conventional cigarettes under both sets of smoking conditions. These results show that the MLA can be used to assess and compare the mutagenic activity of cigarette mainstream smoke particulate phase in the comprehensive toxicological assessment of cigarette smoke.
Asunto(s)
Bioensayo/métodos , Pruebas de Mutagenicidad/métodos , Mutágenos/toxicidad , Nicotiana/toxicidad , Timidina Quinasa/genética , Contaminación por Humo de Tabaco/efectos adversos , Animales , Técnicas de Cultivo de Célula , Supervivencia Celular/efectos de los fármacos , Seguridad de Productos para el Consumidor , Medios de Cultivo , Ratones , Ratas , Nicotiana/química , Células Tumorales CultivadasRESUMEN
The in vitro toxicity of cigarette mainstream smoke from an electrically heated cigarette (EHC) with controlled combustion was compared with that of the standard University of Kentucky Reference Cigarette 1R4F. In the Salmonella reverse mutation assay, strains TA98, TA100, TA102, TA1535 and TA1537 were used in the absence and presence of a metabolic promutagen activation system (S9) to determine the mutagenic potential of the total particulate matter (TPM), which was collected on a glass-fiber filter. In the neutral red uptake assay, mouse embryo BALB/c 3T3 cells were used to determine the cytotoxic potential of TPM as well as of the water-solubles in the gas/vapor phase trapped in phosphate-buffered saline. The TPM from the electrically heated cigarette was up to 90% lower in mutagenicity than that of the 1R4F calculated on an equal TPM basis. This reduction in mutagenicity is consistent with the significantly lower concentration of nearly all constituents analyzed in EHC smoke. With regard to cytotoxicity when calculated on an equal TPM basis, TPM from the electrically heated cigarette was 40% less active relative to the 1R4F. When calculated on a per cigarette basis, the cytotoxicity of both the TPM fraction and the water-solubles in the gas/vapor phase of smoke from the EHC was ca. 80% lower relative to the 1R4F.
Asunto(s)
Calefacción , Nicotiana/toxicidad , Humo/análisis , Animales , Células 3T3 BALB , Supervivencia Celular/efectos de los fármacos , Electricidad , Ratones , Pruebas de Mutagenicidad , Humo/efectos adversosRESUMEN
Cigarette mainstream smoke from blended cigarettes with and without the addition of ingredients was assayed for its cytotoxicity and genotoxicity. In total, 333 ingredients commonly used in cigarette manufacturing were assigned to three different groups. Each group of ingredients was added at a low and a high level to the test cigarettes. The mutagenicity of the particulate phase of the resulting cigarette smoke was assayed in the Salmonella plate incorporation (Ames) assay with tester strains TA98, TA100, TA102, TA1535 and TA1537. The cytotoxicity of the gas/vapor phase and the particulate phase was determined in the neutral red uptake assay with mouse embryo BALB/c 3T3 cells. Within the sensitivity and specificity of the test systems, the in vitro mutagenicity and cytotoxicity of the cigarette smoke were not increased by the addition of the ingredients.
Asunto(s)
Muerte Celular , Pruebas de Mutagenicidad , Nicotiana/química , Humo/efectos adversos , Humo/análisis , Células 3T3 , Animales , Embrión de Mamíferos , Ratones , Ratones Endogámicos BALB C , Rojo Neutro/metabolismoRESUMEN
Stimulation of the Interleukin-1 receptor type I (IL-1-RI) with IL-1 activates an associated serine/threonine kinase, IRAK, which phosphorylates downstream targets, resulting in NFkappaB activation. The signaling cascade is accompanied by oxidative processes and contains putative targets for redox regulation. Preincubation of the murine T cell line EL-4 and the human umbilical cord vein endothelial cell line ECV 304 with thiol modifying compounds like diamide, menadione or phenylarsine oxide inhibited the IL-1-induced phosphorylation of an endogenous substrate with a molecular mass of 60 kD. In the endothelial cell line, a second target of about 85 kD was phosphorylated after IL-1 stimulation, which was also inhibited by thiol modification. These data suggest that IL-1 signal transduction depends on free thiols which might be targets for redox regulation not only in lymphocytes, but also in endothelial cells.
Asunto(s)
Endotelio Vascular/fisiología , Interleucina-1/fisiología , Receptores de Interleucina-1/fisiología , Transducción de Señal/fisiología , Compuestos de Sulfhidrilo/farmacología , Animales , Arsenicales/farmacología , Células Cultivadas , Diamida/farmacología , Endotelio Vascular/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Humanos , Interleucina-1/farmacología , Ratones , Peso Molecular , FN-kappa B/metabolismo , Oxidación-Reducción , Fosforilación , Receptores de Interleucina-1/efectos de los fármacos , Receptores Tipo I de Interleucina-1 , Transducción de Señal/efectos de los fármacos , Timoma , Neoplasias del Timo , Células Tumorales Cultivadas , Venas Umbilicales , Vitamina K/farmacologíaRESUMEN
The interleukin-1 receptor type I (IL-1RI) is associated with other proteins thus forming a complex system by which IL-1 exerts its various signals. The initiating event is still uncertain, but activation of a recently described receptor-associated protein kinase is one of the earliest events detectable (Martin et al., Eur. J. Immunol. 1994. 24: 1566). IL-1 signaling is commonly accompanied by oxidative processes and is thought to be subject to redox regulation. We therefore investigated whether the activation of the IL-1RI-associated protein kinase could be a target for redox regulation and whether an altered activity of the kinase could influence IL-1-mediated NF-kappa B activation. A murine T cell line, EL4, was stimulated with IL-1 with and without pretreatment with different compounds known to influence the cellular redox status. Thiol modifying agents like diamide, menadione, pyrrolidine dithiocarbamate (PDTC), diethyl dithiocarbamate or phenylarsine oxide inhibited the IL-1-induced activation of the IL-1RI-associated protein kinase. N-Acetylcysteine, alpha,alpha'-dipyridyl, aminotriazole or nitrofurantoin did not show any effect. The inhibition by PDTC was reversible unless glutathione synthesis was blocked by buthionine sulfoximine. The described conditions which inhibited or prevented the activation of the IL-1RI-associated kinase similarly impaired the activation of NF-kappa B in EL4 cells. From these observations we conclude that free thiols in the IL-1RI complex are essential for the activation of the IL-1RI-associated protein kinase and that this process is mandatory for IL-1 signaling leading to NF-kappa B activation.
Asunto(s)
Interleucina-1/antagonistas & inhibidores , Interleucina-1/farmacología , FN-kappa B/metabolismo , Proteínas Quinasas/metabolismo , Receptores de Interleucina-1/metabolismo , Compuestos de Sulfhidrilo/metabolismo , Animales , Ditiocarba/farmacología , Activación Enzimática/efectos de los fármacos , Activación Enzimática/inmunología , Glutatión/farmacología , Humanos , Quinasas Asociadas a Receptores de Interleucina-1 , Ratones , FN-kappa B/antagonistas & inhibidores , Oxidación-Reducción/efectos de los fármacos , Inhibidores de Proteínas Quinasas , Pirrolidinas/antagonistas & inhibidores , Pirrolidinas/farmacología , Receptores de Interleucina-1/antagonistas & inhibidores , Linfocitos T/efectos de los fármacos , Linfocitos T/enzimología , Linfocitos T/metabolismo , Tiocarbamatos/antagonistas & inhibidores , Tiocarbamatos/farmacología , Timoma , Neoplasias del Timo , Células Tumorales Cultivadas , Vitamina K/farmacologíaRESUMEN
To study the possible association between drinking tea and lung cancer, epidemiological and experimental materials were collected from the Chinese population in Hong Kong. In a retrospective study of 200 female lung cancer patients and 200 matched controls, all subjects were interviewed concerning their eating habits, smoking histories, and lifetime exposures to environmental pollutants. Analysis of the data demonstrated an adjusted and statistically significant increased lung cancer risk of 2.7 among those who drank green tea. Several brands of tea commonly drank in Hong Kong were assayed for mutagenicity using Ames' assay. Significantly elevated levels of mutagenic activity were found to be present after metabolic activation using cell-free extracts of cecal bacteria from rats. These data suggest that further research is needed to understand the possible consequences to human health of ingestants taken at low doses but long duration over the normal lifespan, and the possible interactive effect between mutagens in tea and other ingestants and inhalants in human cancer etiology.