RESUMEN
PURPOSE: Loss of vaccine potency due to extreme temperature exposure during storage and transport remains a significant obstacle to the success of many vaccines, including the Bacille Calmette-Guérin (BCG) vaccine, the only vaccine available against Mycobacterium tuberculosis. BCG is a live, attenuated vaccine requiring refrigerated storage for viability. In this study, we formulated a temperature-stable BCG dry powder using the spray drying technique. METHODS: We employed a factorial design to optimize our formulation of stabilizing excipients that included L-leucine, bovine serum albumin, polyvinylpyrrolidone, mannitol, and trehalose. Powders were characterized for their particle size, yield, water retention and uptake, glass transition temperature, and aerosol performance. Three optimal powder carrier mixtures were selected from the factorial design for BCG incorporation based on their stability-promoting and powder flow characteristics. Vaccine powders were also assessed for BCG viability and in vivo immunogenicity after long-term storage. RESULTS: Live BCG was successfully spray-dried using the optimized carriers. Dry powder BCG showed no loss in viability (25°C, up to 60% relative humidity; RH) and ~2-log loss in viability (40°C, 75% RH) after one year of storage. The aerodynamic size of the powders was in the respirable range. Further, when healthy mice were immunized intradermally with reconstituted BCG powders (storage for 2 years), the vaccine retained its immunogenicity. CONCLUSION: We developed a spray-dried BCG vaccine that was viable and antigenic after long-term storage. To our knowledge, this is a first study to show room temperature stability of live BCG vaccine without any loss in viability for 12 months.
Asunto(s)
Vacuna BCG/química , Vacuna BCG/farmacología , Composición de Medicamentos/métodos , Excipientes/química , Polvos/química , Aerosoles/química , Animales , Línea Celular , Supervivencia Celular , Desecación/métodos , Estabilidad de Medicamentos , Almacenaje de Medicamentos , Femenino , Humanos , Leucina/química , Manitol/química , Ratones Endogámicos C57BL , Mycobacterium bovis/citología , Povidona/química , Albúmina Sérica Bovina/química , Temperatura , Distribución Tisular , Trehalosa/químicaRESUMEN
Bacille Calmette-Guérin (BCG) is currently the only approved vaccine against tuberculosis (TB) and is administered in over 150 countries worldwide. Despite its widespread use, the vaccine has a variable protective efficacy of 0-80%, with the lowest efficacy rates in tropical regions where TB is most prevalent. This variability is partially due to ubiquitous environmental mycobacteria (EM) found in soil and water sources, with high EM prevalence coinciding with areas of poor vaccine efficacy. In an effort to elucidate the mechanisms underlying EM interference with BCG vaccine efficacy, we exposed mice chronically to Mycobacterium avium (M. avium), a specific EM, by two different routes, the oral and intradermal route, to mimic human exposure. After intradermal BCG immunization in mice exposed to oral M. avium, we saw a significant decrease in the pro-inflammatory cytokine IFN-γ, and an increase in T regulatory cells and the immunosuppressive cytokine IL-10 compared to naïve BCG-vaccinated animals. To circumvent the immunosuppressive effect of oral M. avium exposure, we vaccinated mice by the pulmonary route with BCG. Inhaled BCG immunization rescued IFN-γ levels and increased CD4 and CD8 T cell recruitment into airways in M. avium-presensitized mice. In contrast, intradermal BCG vaccination was ineffective at T cell recruitment into the airway. Pulmonary BCG vaccination proved protective against Mtb infection regardless of previous oral M. avium exposure, compared to intradermal BCG immunization. In conclusion, our data indicate that vaccination against TB by the pulmonary route increases BCG vaccine efficacy by avoiding the immunosuppressive interference generated by chronic oral exposure to EM. This has implications in TB-burdened countries where drug resistance is on the rise and health care options are limited due to economic considerations. A successful vaccine against TB is necessary in these areas as it is both effective and economical.
Asunto(s)
Vacuna BCG/administración & dosificación , Exposición a Riesgos Ambientales/efectos adversos , Tolerancia Inmunológica/inmunología , Mycobacterium avium/inmunología , Tuberculosis Pulmonar/inmunología , Animales , Vacuna BCG/inmunología , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Femenino , Citometría de Flujo , Inmunofenotipificación , Ratones , Ratones Endogámicos C57BLRESUMEN
This brief communication evaluates the cytotoxicity and targeting capability of a dry powder chemotherapeutic. Nano-in-microparticles (NIMs) are a dry powder drug delivery vehicle containing superparamagnetic iron oxide nanoparticles (SPIONs) and either doxorubicin (w/w solids) or fluorescent nanospheres (w/v during formulation; as a drug surrogate) in a lactose matrix. In vitro cytotoxicity was evaluated in A549 adenocarcinoma cells using MTS and LDH assays to assess viability and toxicity after 48 h of NIMs exposure. In vivo magnetic-field-dependent targeting of inhaled NIMs was evaluated in a healthy mouse model. Mice were endotracheally administered fluorescently labeled NIMs either as a dry powder or a liquid aerosol in the presence of an external magnet placed over the left lung. Quantification of fluorescence and iron showed a significant increase in both fluorescence intensity and iron content to the left magnetized lung. In comparison, we observed decreased targeting of fluorescent nanospheres to the left lung from an aerosolized liquid suspension, due to the dissociation of SPIONs and nanoparticles during pulmonary administration. We conclude that dry powder NIMs maintain the therapeutic cytotoxicity of doxorubicin and can be better targeted to specific regions of the lung in the presence of a magnetic field, compared to a liquid suspension.
Asunto(s)
Doxorrubicina/administración & dosificación , Portadores de Fármacos/química , Compuestos Férricos/química , Nanopartículas de Magnetita/química , Células A549 , Aerosoles , Animales , Humanos , Pulmón/efectos de los fármacos , Pulmón/metabolismo , Campos Magnéticos , Masculino , Ratones , Ratones Endogámicos BALB C , Modelos Animales , Nanosferas/química , Polvos , Tráquea/efectos de los fármacos , Tráquea/metabolismoRESUMEN
Existing vaccines against human papillomavirus (HPV) require continuous cold-chain storage. Previously, we developed a bacteriophage virus-like particle (VLP)-based vaccine for HPV infection, which elicits broadly neutralizing antibodies against diverse HPV types. Here, we formulated these VLPs into a thermostable dry powder using a multicomponent excipient system and by optimizing the spray-drying parameters using a half-factorial design approach. Dry-powder VLPs were stable after spray drying and after long-term storage at elevated temperatures. Immunization of mice with a single dose of reconstituted dry-powder VLPs that were stored at 37 °C for more than a year elicited high anti-L2 IgG antibody titers. Spray-dried thermostable, broadly protective L2 bacteriophage VLPs vaccine could be accessible to remote regions of the world (where â¼84% of cervical cancer patients reside) by eliminating the cold-chain requirement during transportation and storage.
Asunto(s)
Papillomaviridae/inmunología , Infecciones por Papillomavirus/inmunología , Vacunas de Partículas Similares a Virus/química , Vacunas de Partículas Similares a Virus/inmunología , Animales , Anticuerpos Neutralizantes/inmunología , Química Farmacéutica/métodos , Humanos , Inmunización/métodos , Inmunoglobulina G/inmunología , Ratones , Ratones Endogámicos BALB C , Polvos/administración & dosificación , Polvos/química , Temperatura , Vacunación/métodosRESUMEN
Conventional spray drying using a 2-fluid nozzle forms matrix microparticles, where drug is distributed throughout the particle and may not effectively mask taste. In contrast, spray drying using a 3-fluid nozzle has been reported to encapsulate material. The objective of this study was to spray dry Eudragit® E-PO (EE) with acetaminophen (APAP), a water-soluble model drug with a bitter taste, using 2- and 3-fluid nozzles for taste masking. Spray drying EE with APAP, however, resulted in yields of ≤ 13 %, irrespective of nozzle configuration. Yields improved when Eudragit® L 100-55 (EL) or Methocel® E6 (HPMC) was used in the inner fluid stream of the 3-fluid nozzle or in place of EE for the 2-fluid nozzle. Drug release from microparticles prepared with the 2-fluid nozzle was relatively rapid. Using EE in the outer fluid stream of the 3-fluid nozzle resulted in comparatively slower drug release, although drug release was observed, indicating that encapsulation was incomplete. Results from these studies also show that miscible polymers used in the two fluid streams mix during the spray drying process. In addition, findings from this study indicate that the polymer used in the inner fluid stream can impact drug release.
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Acetaminofén , Liberación de Fármacos , Ácidos Polimetacrílicos , Gusto , Acetaminofén/química , Acetaminofén/administración & dosificación , Ácidos Polimetacrílicos/química , Secado por Pulverización , Composición de Medicamentos/métodos , Derivados de la Hipromelosa/química , Tamaño de la Partícula , Solubilidad , Desecación/métodos , Resinas AcrílicasRESUMEN
We propose the use of novel inhalable nano-in-microparticles (NIMs) for site-specific pulmonary drug delivery. Conventional lung cancer therapy has failed to achieve therapeutic drug concentrations at tumor sites without causing adverse effects in healthy tissue. To increase targeted drug delivery near lung tumors, we have prepared and characterized a magnetically responsive dry powder vehicle containing doxorubicin. A suspension of lactose, doxorubicin and Fe3O4 superparamagnetic iron oxide nanoparticles (SPIONs) were spray dried. NIMs were characterized for their size and morphological properties by various techniques: dynamic light scattering (DLS) and laser diffraction (LS) to determine hydrodynamic size of the SPIONs and the NIMs, respectively; next generation cascade impactor (NGI) to determine the aerodynamic diameter and fine particle fraction (FPF); scanning (SEM) and transmission (TEM) electron microscopy to analyze particle surface morphology; electron dispersive X-ray spectroscopy (EDS) to determine iron loading in NIMs; inductively coupled plasma atomic emission spectroscopy (ICP-AES) and superconducting quantum interference device (SQUID) to determine Fe3O4 content in the microparticles; and high performance liquid chromatography (HPLC) to determine doxorubicin loading in the vehicle. NIMs deposition and retention near a magnetic field was performed using a proof-of-concept cylindrical tube to mimic the conducting airway deposition. The hydrodynamic size and zeta potential of SPIONs were 56 nm and -49 mV, respectively. The hydrodynamic and aerodynamic NIM diameters were 1.6 µm and 3.27±1.69 µm, respectively. SEM micrographs reveal spherical particles with rough surface morphology. TEM and focused ion beam-SEM micrographs corroborate the porous nature of NIMs, and surface localization of SPIONs. An in vitro tracheal mimic study demonstrates more than twice the spatial deposition and retention of NIMs, compared to a liquid suspension, in regions under the influence of a strong magnetic gradient. We report the novel formulation of an inhaled and magnetically responsive NIM drug delivery vehicle. This vehicle is capable of being loaded with one or more chemotherapeutic agents, with future translational ability to be targeted to lung tumors using an external magnetic field.
Asunto(s)
Sistemas de Liberación de Medicamentos/métodos , Compuestos Férricos/química , Magnetismo , Nanopartículas/química , Administración por Inhalación , Pulmón/metabolismo , Nanopartículas/ultraestructura , Tamaño de la Partícula , Tráquea/metabolismoRESUMEN
Nisin ZP is an antimicrobial peptide (AMP) produced by the bacterium Lactococcus lactis, and we have previously demonstrated anticancer activity in NSCLC (A549) cells. In this study, we formulated a nisin ZP dry powder (NZSD) using a spray dryer to facilitate inhaled delivery for the treatment of NSCLC. Nisin ZP was spray-dried with mannitol, l-leucine, and trehalose in a ratio of 75:15:10 using Büchi mini spray-dryer B-290 in different drug loadings (10, 20, and 30% w/w). NZSD powder revealed a good powder yield of >55% w/w with ≤3 % w/w moisture content and high nisin ZP drug loading for all the peptide ratios. The NZSD powder particles were irregularly shaped with corrugated morphology. The presence of an endothermic peak in DSC thermograms and attenuated crystalline peaks in PXRD diffractograms confirmed the semi-crystalline powder nature of NZSD. The anticancer activity of nisin ZP was maintained after fabricating it into NZSD powder and showed a similar inhibitory concentration to free nisin ZP. Stability studies indicated that NZSD powders were stable for three months at 4 and 25 â with more than 90% drug content and semi-crystalline nature, as confirmed by DSC and PXRD. Aerosolization studies performed using NGI indicated an aerodynamic diameter (MMAD) within the desired range (1-5 µm) and a high fine particle fraction (FPF > 75%) for all peptide ratios, suggesting powder deposition in the lung's respiratory airways. In conclusion, a dry powder of nisin ZP was formulated using a spray dryer with enhanced storage stability and suitable for inhaled delivery.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Nisina , Humanos , Administración por Inhalación , Péptidos Antimicrobianos , Polvos/química , Aerosoles y Gotitas Respiratorias , Pulmón , Tamaño de la Partícula , Inhaladores de Polvo SecoRESUMEN
The global control of tuberculosis (TB) is at risk by the spread of multidrug-resistant TB (MDR TB). Treatment of MDR TB is lengthy and involves injected drugs, such as capreomycin, that have severe side effects. It was previously reported that a single daily dose of inhaled capreomycin had a positive effect on the bacterial burden of TB-infected guinea pigs. The modest effect observed was possibly due to a dose that resulted in insufficient time of exposure to therapeutic systemic and local levels of the drug. In order to determine the length of time that systemic and local drug concentrations are above therapeutic levels during the treatment period, the present study investigated the disposition of capreomycin powders after sequential pulmonary administration of doses of 20 mg/kg of body weight. Capreomycin concentrations in bronchoalveolar lavage fluid and lung tissue of animals receiving a series of one, two, or three doses of capreomycin inhalable powder were significantly higher (50- to 100-fold) at all time points than plasma concentrations at the same time points or those observed in animals receiving capreomycin solution by intramuscular (i.m.) injection (10- to 100-fold higher). Notably, at the end of each dosing period, capreomycin concentrations in the lungs were approximately 100-fold higher than those in plasma and severalfold higher than the MIC, suggesting that sufficient capreomycin remains in the lung environment to kill Mycobacterium tuberculosis. No accumulation of capreomycin powder was detected in the lungs after 3 pulmonary doses. These results indicate that the systemic disposition of capreomycin after inhalation is the same as when injected i.m. with the advantage that higher drug concentrations are present at all times in the lungs, the primary site of infection.
Asunto(s)
Antituberculosos/farmacocinética , Capreomicina/farmacocinética , Pulmón/efectos de los fármacos , Mycobacterium tuberculosis/efectos de los fármacos , Tuberculosis Resistente a Múltiples Medicamentos/tratamiento farmacológico , Administración por Inhalación , Animales , Antituberculosos/sangre , Antituberculosos/uso terapéutico , Líquido del Lavado Bronquioalveolar/química , Capreomicina/sangre , Capreomicina/uso terapéutico , Modelos Animales de Enfermedad , Esquema de Medicación , Cobayas , Inyecciones Intramusculares , Pulmón/microbiología , Masculino , Mycobacterium tuberculosis/fisiología , Tamaño de la Partícula , Polvos , Tuberculosis Resistente a Múltiples Medicamentos/sangre , Tuberculosis Resistente a Múltiples Medicamentos/microbiologíaRESUMEN
By manufacturing a single-particle system in two particulate forms (i.e., micrometer size and nanometer size), we have designed a bacterial vaccine form that exhibits improved efficacy of immunization. Microstructural properties are adapted to alter dispersive and aerosol properties independently. Dried "nanomicroparticle" vaccines possess two axes of nanoscale dimensions and a third axis of micrometer dimension; the last one permits effective micrometer-like physical dispersion, and the former provides alignment of the principal nanodimension particle axes with the direction of airflow. Particles formed with this combination of nano- and micrometer-scale dimensions possess a greater ability to aerosolize than particles of standard spherical isotropic shape and of similar geometric diameter. Here, we demonstrate effective application of this biomaterial by using the live attenuated tuberculosis vaccine bacille Calmette-Guérin (BCG). Prepared as a spray-dried nanomicroparticle aerosol, BCG vaccine exhibited high-efficiency delivery and peripheral lung targeting capacity from a low-cost and technically simple delivery system. Aerosol delivery of the BCG nanomicroparticle to normal guinea pigs subsequently challenged with virulent Mycobacterium tuberculosis significantly reduced bacterial burden and lung pathology both relative to untreated animals and to control animals immunized with the standard parenteral BCG.
Asunto(s)
Aerosoles/administración & dosificación , Aerosoles/farmacología , Vacunas Bacterianas/inmunología , Inmunización/métodos , Animales , Vacuna BCG/inmunología , Vías de Administración de Medicamentos , Cobayas , Humedad , Hipersensibilidad Tardía/inmunología , Hipersensibilidad Tardía/microbiología , Leucina/administración & dosificación , Leucina/farmacología , Pulmón/efectos de los fármacos , Pulmón/microbiología , Pulmón/patología , Masculino , Viabilidad Microbiana/efectos de los fármacos , Mycobacterium smegmatis/inmunología , Mycobacterium smegmatis/ultraestructura , Bazo/efectos de los fármacos , Bazo/microbiología , Bazo/patología , TuberculinaRESUMEN
Three prophylactic vaccines are approved to protect against HPV infections. These vaccines are highly immunogenic. The most recent HPV vaccine, Gardasil-9, protects against HPV types associated with ~90% of cervical cancer (worldwide). Thus, ~10% of HPV-associated cancers are not protected by Gardasil-9. Although this is not a large percentage overall, the HPV types associated with 10% of cervical cancer not protected by the current vaccine are significantly important, especially in HIV/AIDS patients who are infected with multiple HPV types. To broaden the spectrum of protection against HPV infections, we developed mixed MS2-L2 VLPs (MS2-31L2/16L2 VLPs and MS2-consL2 (69-86) VLPs) in a previous study. Immunization with the VLPs neutralized/protected mice against infection with eleven high-risk HPV types associated with ~95% of cervical cancer and against one low-risk HPV type associated with ~36% of genital warts & up to 32% of recurrent respiratory papillomatosis. Here, we report that the mixed MS2-L2 VLPs can protect mice from three additional HPV types: HPV51, which is associated with ~0.8% of cervical cancer; HPV6, which is associated with up to 60% of genital warts; HPV5, which is associated with skin cancers in patients with epidermodysplasia verruciformis (EV). Overall, mixed MS2-L2 VLPs can protect against twelve HPV types associated with ~95.8% of cervical cancers and against two HPV types associated with ~90% of genital warts and >90% recurrent respiratory papillomatosis. Additionally, the VLPs protect against one of two HPV types associated with ~90% of HPV-associated skin cancers in patients with EV. More importantly, we observed that mixed MS2-L2 VLPs elicit protective antibodies that last over 9 months. Furthermore, a spray-freeze-dried formulation of the VLPs is stable, immunogenic, and protective at room temperature and 37 °C.
Asunto(s)
Anticuerpos Antivirales/sangre , Bacteriófagos/inmunología , Papillomaviridae/inmunología , Infecciones por Papillomavirus/prevención & control , Vacunas contra Papillomavirus/administración & dosificación , Vacunas de Partículas Similares a Virus/inmunología , Animales , Condiloma Acuminado/prevención & control , Protección Cruzada/inmunología , Femenino , Humanos , Inmunización , Ratones , Ratones Endogámicos BALB C , Papillomaviridae/clasificación , Papillomaviridae/patogenicidad , Vacunas contra Papillomavirus/inmunología , Neoplasias del Cuello Uterino/prevención & control , Vacunas de Partículas Similares a Virus/administración & dosificaciónRESUMEN
OBJECTIVES: Health care workers experience an uncertain risk of aerosol exposure during patient oxygenation. To improve our understanding of these risks, we sought to measure aerosol production during various approaches to oxygenation in healthy volunteers in an emergency department. METHODS: This was a prospective study conducted in an empty patient room in an academic ED. The room was 10 ft. long x 10 ft. wide x 9 ft. tall (total volume 900 ft3) with positive pressure airflow (1 complete turnover of air every 10 minutes). Five oxygenation conditions were used: humidified high-flow nasal cannula (HFNC) at 3 flow rates [15, 30, and 60 liters per minute (LPM)], non-rebreather mask (NRB) at 1 flow rate (15 LPM), and closed-circuit continuous positive airway pressure (CPAP) using the ED ventilator; in all cases a simple procedural mask was used. The NRB and HFNC at 30 LPM maneuvers were also repeated without the procedural mask, and CPAP was applied both with and without a filter. Each subject then sequentially underwent 8 total oxygenation conditions, always in the same order. Each oxygenation condition was performed with the participant on a standard ED bed. Particles were measured by laser aerosol spectrometer, with the detector sampling port positioned directly over the center of the bed, 0.35 meters away and at a 45-degree angle from the subject's mouth. Each approach to oxygenation was performed for 10 minutes, followed by a 20-minute room washout (≈ 2 complete room air turnovers). Particle counts were summated for 2 size ranges (150-300 nm and 0.5-2.0 µm) and compared before, during, and after each of the 8 oxygenation conditions. RESULTS: Eight adult subjects were enrolled (mean age 42 years, body mass index 25). All subjects completed 8 oxygenation procedures (64 total). Mean particle counts per minute across all oxygenation procedures was 379 ± 112 (mean ± SD) for smaller aerosols (150-300 nm) and 9.3 ± 4.6 for larger aerosols (0.5-2.0 µm). HFNC exhibited a flow-dependent increase in particulate matter (PM) generation-at 60 LPM, HFNC had a substantial generation of small (55% increase) and large particles (70% increase) compared to 15 LPM. CPAP was associated with lowered small and large particle generation (≈ 10-15% below baseline for both sizes of PM). A patient mask limited particle generation with the NRB, where it was associated with a reduction in small and large particulates (average 40% and 20% lower, respectively). CONCLUSION: Among 3 standard oxygenation procedures, higher flow rates generally were associated with greater production of both small and large aerosols. A patient mask lowered aerosol counts in the NRB only. Protocol development for oxygenation application should consider these factors to increase health care worker safety.
RESUMEN
We investigated the preclinical efficacy and safety/tolerability of biodegradable polymeric particles containing isoniazid (INH) and rifabutin (RFB) dry powder for inhalation (DPI) as an adjunct to oral first-line therapy. Mice and guinea pigs infected with Mycobacterium tuberculosis H37Rv (Mtb) were treated with â¼80 and â¼300 µg of the DPI, respectively, for 3-4 weeks starting 3, 10, and 30 days post-infection. Adjunct combination therapy eliminated culturable Mtb from the lungs and spleens of all but one of 52 animals that received the DPI. Relapse-free cure was not achieved in one mouse that received DPI + oral, human-equivalent doses (HED) of four drugs used in the Directly Observed Treatment, Short Course (DOTS), starting 30 days post-infection. Oral doses (20 mg/Kg/day, each) of INH + RFB reduced Mtb burden from â¼106 to â¼103â¯colony-forming units. Combining half the oral dose with DPI prevented relapse of infection four weeks after stopping the treatment. The DPI was safe in rodents, guinea pigs, and monkeys at 1, 10, and 100⯵g/day doses over 90 days. In conclusion, we show the efficacy and safety/tolerability of the DPI as an adjunct to oral chemotherapy in three different animal models of TB.
Asunto(s)
Antituberculosos/uso terapéutico , Isoniazida/uso terapéutico , Rifabutina/uso terapéutico , Tuberculosis/tratamiento farmacológico , Administración por Inhalación , Animales , Quimioterapia Combinada , Femenino , Cobayas , Isoniazida/administración & dosificación , Macaca mulatta , Masculino , Ratones , Mycobacterium tuberculosis , Recurrencia , Rifabutina/administración & dosificaciónRESUMEN
Novel treatments for multidrug-resistant tuberculosis (MDR-TB), extensively drug-resistant tuberculosis (XDR-TB), or latent TB are needed urgently. Recently, we reported the formulation and characterization of the nitroimidazo-oxazine PA-824 for efficient aerosol delivery as dry powder porous particles and the subsequent disposition in guinea pigs after pulmonary administration. The objective of the present study was to evaluate the effects of these PA-824 therapeutic aerosols on the extent of TB infection in the low-inoculum aerosol infection guinea pig model. Four weeks after infection by the pulmonary route, animals received daily treatment for 4 weeks of either a high or a low dose of PA-824 dry powder aerosol. Animals received PA-824 cyclodextrin/lecithin suspensions orally as positive controls, and those receiving placebo particles or no treatment were negative controls. The lungs and spleens of animals receiving the high dose of inhaled PA-824 particles exhibited a lower degree of inflammation (indicated by wet tissue weights), bacterial burden, and tissue damage (indicated by histopathology) than those of untreated or placebo animals. Treatment with oral PA-824 cyclodextrin/lecithin suspension resulted in a more significant reduction in the bacterial burden of lungs and spleen, consistent with a dose that was larger than inhaled doses (eight times the inhaled low dose and four times the inhaled high dose). However, histopathological analysis revealed that the extent of tissue damage was comparable in groups receiving the oral or either inhaled dose. The present studies indicate the potential use of PA-824 dry powder aerosols in the treatment of TB.
Asunto(s)
Antituberculosos/administración & dosificación , Nitroimidazoles/administración & dosificación , Tuberculosis Pulmonar/tratamiento farmacológico , Administración por Inhalación , Administración Oral , Aerosoles , Animales , Antituberculosos/sangre , Química Farmacéutica/métodos , Recuento de Colonia Microbiana , Modelos Animales de Enfermedad , Tuberculosis Extensivamente Resistente a Drogas/tratamiento farmacológico , Cobayas , Humanos , Tuberculosis Latente/tratamiento farmacológico , Pulmón/microbiología , Pulmón/patología , Masculino , Nitroimidazoles/sangre , Tamaño de la Partícula , Polvos , Bazo/microbiología , Bazo/patología , Tuberculosis Resistente a Múltiples Medicamentos/tratamiento farmacológico , Tuberculosis Pulmonar/sangre , Tuberculosis Pulmonar/microbiología , Tuberculosis Pulmonar/patologíaRESUMEN
Lactate dehydrogenase (LDH) catalyzes the conversion of pyruvate to lactate, with concomitant oxidation of reduced nicotinamide adenine dinucleotide as the final step in the glycolytic pathway. Glycolysis plays an important role in the metabolic plasticity of cancer cells and has long been recognized as a potential therapeutic target. Thus, potent, selective inhibitors of LDH represent an attractive therapeutic approach. However, to date, pharmacological agents have failed to achieve significant target engagement in vivo, possibly because the protein is present in cells at very high concentrations. We report herein a lead optimization campaign focused on a pyrazole-based series of compounds, using structure-based design concepts, coupled with optimization of cellular potency, in vitro drug-target residence times, and in vivo PK properties, to identify first-in-class inhibitors that demonstrate LDH inhibition in vivo. The lead compounds, named NCATS-SM1440 (43) and NCATS-SM1441 (52), possess desirable attributes for further studying the effect of in vivo LDH inhibition.
Asunto(s)
Inhibidores Enzimáticos/farmacología , L-Lactato Deshidrogenasa/antagonistas & inhibidores , Pirazoles/farmacología , Animales , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacocinética , Semivida , Humanos , Ratones , Relación Estructura-Actividad , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
One third of the world population is infected with tuberculosis (TB), and new infections occur at a rate of one per second. The recent increase in the emergence of drug-resistant strains of Mycobacterium tuberculosis and the dearth of anti-TB drugs is threatening the future containment of TB. New drugs or delivery systems that will stop the spread of TB and slow down or prevent the development of drug-resistant strains are urgently required. One of the reasons for the emergence of drug-resistant strains is the exposure of mycobacteria to sub-therapeutic levels of one or more antibiotics. Lung lesions containing large numbers of bacteria are poorly vascularized and are fortified with thick fibrous tissue; conventional therapy by the oral and parenteral routes may provide sub-therapeutic levels of anti-TB drugs to these highly sequestered organisms. Administering drugs by the pulmonary route to the lungs allows higher drug concentrations in the vicinity of these lesions. Supplementing conventional therapy with inhaled anti-TB therapy may allow therapeutic concentrations of drug to penetrate effectively into lung lesions and treat the resident mycobacteria.
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Antituberculosos/uso terapéutico , Mycobacterium tuberculosis/fisiología , Tuberculosis/tratamiento farmacológico , Administración por Inhalación , Antituberculosos/administración & dosificación , Humanos , Tuberculosis/fisiopatologíaRESUMEN
Macrophage responses to infection with Mycobacterium tuberculosis (MTB) and treatment with soluble isoniazid (INH) plus rifabutin (RFB) versus microparticles containing equivalent amounts of drugs were compared. It was investigated whether macrophages driven to alternative activation upon infection with MTB could be rescued to display the classical activation phenotype. It was established that microparticles sustain high levels of drugs in cytosol of macrophages for longer period as compared to soluble drugs. Microparticles co-localized with intracellular bacteria, and induced a variety of innate bactericidal responses, including induction of free radicals, alteration of mitochondrial membrane potential and apoptosis. The data strongly suggest that additional benefit may be derived from the nature of the drug delivery system, which fulfils Koch's dictum 'stimulate the phagocyte' for curing tuberculosis.
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Antituberculosos , Isoniazida , Macrófagos/efectos de los fármacos , Fagocitos/efectos de los fármacos , Rifabutina , Administración por Inhalación , Antituberculosos/administración & dosificación , Antituberculosos/farmacología , Fragmentación del ADN , Humanos , Isoniazida/administración & dosificación , Isoniazida/farmacología , Activación de Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Mycobacterium tuberculosis/inmunología , Tamaño de la Partícula , Fagocitos/metabolismo , Rifabutina/administración & dosificación , Rifabutina/farmacología , Tuberculosis/inmunologíaRESUMEN
High-risk human papillomavirus (HPV) types are responsible for nearly all cases of cervical cancers. Cervarix® and Gardasil® 9 are the current prophylactic vaccines available that protect against the majority of HPVs associated with cancer. Although these vaccines are highly effective, HPV vaccine implementation has been slow, particularly in low-and-middle income countries. Major barriers to the widespread availability of the HPV vaccines is its cost and the requirement for continuous refrigeration (2-8°C). Here, we used spray drying along with stabilizing excipients to formulate a thermostable Gardasil® 9 vaccine. We evaluated the immunogenicity and protective efficacy of the vaccine in mice immediately after spray drying and following storage for three months at 4°C, 25°C, and 40°C. The immunogenicity studies were performed using Gardasil® 9 as a whole antigen, and not individual HPV types, for ELISA. At the dose tested, the spray dried vaccine conferred protection against HPV following storage at temperatures up to 40°C. In addition to the spray-dried vaccine, our studies revealed that the Gardasil® 9 vaccine, as currently marketed, may be stored and transported at elevated temperatures for up to 3 months without losing efficacy, especially against HPV16. This study is critical, as a thermostable vaccine will decrease vaccine cost associated with cold-chain maintenance and could increase vaccine access and coverage, especially in remote regions of the world.
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Anticuerpos Antivirales/sangre , Vacunas contra Papillomavirus/química , Vacunas contra Papillomavirus/inmunología , Temperatura , Animales , Química Farmacéutica , Femenino , Higroscópicos , Inyecciones Intramusculares , Ratones , Ratones Endogámicos BALB C , Polvos , Refrigeración , VacunaciónRESUMEN
Human papillomaviruses (HPVs) are the most common sexually transmitted infections. HPVs are transmitted through anogenital sex or oral sex. Anogenital transmission/infection is associated with anogenital cancers and genital warts while oral transmission/infection is associated with head and neck cancers (HNCs) including recurrent respiratory papillomatosis. Current HPV vaccines protect against HPV types associated with â¼90% of cervical cancers and are expected to protect against a percentage of HNCs. However, only a few studies have assessed the efficacy of current vaccines against oral HPV infections. We had previously developed a mixed MS2-L2 candidate HPV vaccine based on bacteriophage MS2 virus-like particles (VLPs). The mixed MS2-L2 VLPs consisted of a mixture of two MS2-L2 VLPs displaying: i) a concatemer of L2 peptide (epitope 20-31) from HPV31 & L2 peptide (epitope 17-31) from HPV16 and ii) a consensus L2 peptide representing epitope 69-86. The mixed MS2-L2 VLPs neutralized/protected mice against six HPV types associated with â¼87% of cervical cancer. Here, we show that the mixed MS2-L2 VLPs can protect mice against additional HPV types; at the genital region, the VLPs protect against HPV53, 56, 11 and at the oral region, the VLPs protect against HPV16, 35, 39, 52, and 58. Thus, mixed MS2-L2 VLPs protect against eleven oncogenic HPV types associated with â¼95% of cervical cancer. The VLPs also have the potential to protect, orally, against the same oncogenic HPVs, associated with â¼99% of HNCs, including HPV11, which is associated with up to 32% of recurrent respiratory papillomatosis. Moreover, mixed MS2-L2 VLPs are thermostable at room temperature for up to 60 days after spray-freeze drying and they are protective against oral HPV infection.
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Protección Cruzada , Papillomaviridae/inmunología , Infecciones por Papillomavirus/prevención & control , Vacunas de Partículas Similares a Virus/inmunología , Animales , Anticuerpos Neutralizantes , Anticuerpos Antivirales/inmunología , Proteínas de la Cápside/inmunología , Protección Cruzada/inmunología , Epítopos/inmunología , Femenino , Neoplasias de Cabeza y Cuello/etiología , Neoplasias de Cabeza y Cuello/prevención & control , Neoplasias de Cabeza y Cuello/virología , Humanos , Inmunización/métodos , Levivirus/inmunología , Ratones , Pruebas de Neutralización , Proteínas Oncogénicas Virales/inmunología , Vacunas contra Papillomavirus/inmunología , Infecciones del Sistema Respiratorio/prevención & control , Neoplasias del Cuello Uterino/etiología , Neoplasias del Cuello Uterino/prevención & control , Neoplasias del Cuello Uterino/virología , Vacunación/métodosRESUMEN
Ambient particulate matter (PM) is associated with increased mortality and morbidity, an effect influenced by the metal components of the PM. We characterized five sediment samples obtained near a tungsten-molybdenum ore-processing complex in Zakamensk, Russia for elemental composition and PM toxicity with regard to pulmonary, vascular, and neurological outcomes. Elemental and trace metals analysis of complete sediment and PM10 (the respirable fraction, < 10 µm mass mean aerodynamic diameter) were performed using inductively coupled plasma optical emission spectrometry (ICP-OES) and mass spectrometry (ICP-MS). Sediment samples and PM10 consisted largely of silicon and iron and silicon and sodium, respectively. Trace metals including manganese and uranium in the complete sediment, as well as copper and lead in the PM10 were observed. Notably, metal concentrations were approximately 10 × higher in the PM10 than in the sediment. Exposure to 100 µg of PM10 via oropharyngeal aspiration in C56BL/6 mice resulted in pulmonary inflammation across all groups. In addition, mice exposed to three of the five PM10 samples exhibited impaired endothelial-dependent relaxation, and correlative analysis revealed associations between pulmonary inflammation and levels of lead and cadmium. A tendency for elevated cortical ccl2 and Tnf-α mRNA expression was induced by all samples and significant upregulation was noted following exposure to PM10 samples Z3 and Z4, respectively. Cortical Nqo1 mRNA levels were significantly upregulated in mice exposed to PM10 Z2. In conclusion, pulmonary exposure to PM samples from the Zakamensk region sediments induced varied pulmonary and systemic effects that may be influenced by elemental PM composition. Further investigation is needed to pinpoint putative drivers of neurological outcomes.
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Contaminantes Atmosféricos/toxicidad , Aorta Torácica/efectos de los fármacos , Cadmio/toxicidad , Corteza Cerebral/efectos de los fármacos , Polvo , Plomo/toxicidad , Minería , Material Particulado/toxicidad , Neumonía/inducido químicamente , Animales , Aorta Torácica/fisiopatología , Corteza Cerebral/metabolismo , Quimiocina CCL2/genética , Quimiocina CCL2/metabolismo , Exposición por Inhalación , Masculino , Ratones Endogámicos C57BL , NAD(P)H Deshidrogenasa (Quinona)/genética , NAD(P)H Deshidrogenasa (Quinona)/metabolismo , Estrés Oxidativo/efectos de los fármacos , Tamaño de la Partícula , Neumonía/genética , Neumonía/metabolismo , Medición de Riesgo , Siberia , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo , Vasodilatación/efectos de los fármacosRESUMEN
Microparticles containing isoniazid and rifabutin were aerosolised using a simple apparatus fabricated from a 15-ml centrifuge tube. The dose available for inhalation by rodents was determined by collecting microparticles emitted at the delivery port. The dose available for inhalation was proportional to durations of exposure ranging from 10 to 90 s (10.5-13.5 CV%) and the weight of powder taken for fluidization (10-50 mg, r2=0.982). The apparatus was then used to administer inhalations of microparticles to mice. Other groups of mice received free rifabutin orally, or by i.v. injection. Rifabutin was estimated in serum and tissues of dosed mice by HPLC. When approximately 20 mg of microparticles were loaded in the apparatus, approximately 2.5 mg were collected at the delivery port in 30 s of operation. Mice inhaled approximately 300 microg of the 2.5 mg emitted at the delivery port. Airway and lung macrophages of mice receiving inhalations for 30 s accumulated 0.38 microg of rifabutin, while the amount in blood serum of these mice was 0.62 microg. In mice receiving 83 microg rifabutin i.v. or orally, the intracellular amounts were 0.06 and 0.07 microg respectively, while the amounts in serum were 1.02 and 0.80 microg. These observations confirmed that inhalation of microparticles targeted airway and lung macrophages.