RESUMEN
Two major chaperones, calreticulin (CRT) and binding immunoglobulin protein (GRP78/BiP) dependent on their location, have immunoregulatory or anti-inflammatory functions respectively. CRT induces pro-inflammatory cytokines, dendritic cell (DC) maturation and activates cytotoxic T cells against tumours. By contrast, GRP78/BiP induces anti-inflammatory cytokines, inhibits DC maturation and heightens T-regulatory cell responses. These latter functions rebalance immune homeostasis in inflammatory diseases, such as rheumatoid arthritis. Both chaperones are therapeutically relevant agents acting primarily on monocytes/DCs. Endogenous exposure of CRT on cancer cell surfaces acts as an 'eat-me' signal and facilitates improved elimination of stressed and dying tumour cells by DCs. Therefore, therapeutics that promote endogenous CRT translocation to the cell surface can improve the removal of cancer cells. However, infused recombinant CRT dampens this cancer cell eradication by binding directly to the DCs. Low levels of endogenous BiP appear as a surface biomarker of endoplasmic reticulum (ER) stress in some types of tumour cells, a reflection of cells undergoing proliferation, in which resulting hypoxia and nutrient deprivation perturb ER homeostasis triggering the unfolded protein response, leading to increased expression of GRP78/BiP and altered cellular location. Conversely, infusion of an analogue of GRP78/BiP (IRL201805) can lead to long-term immune resetting and restoration of immune homeostasis. The therapeutic potential of both chaperones relies on them being relocated from their intracellular ER environment. Ongoing clinical trials are employing therapeutic interventions to either enhance endogenous cell surface CRT or infuse IRL201805, thereby triggering several disease-relevant immune responses leading to a beneficial clinical outcome.
Asunto(s)
Chaperón BiP del Retículo Endoplásmico , Proteínas de Choque Térmico , Humanos , Proteínas de Choque Térmico/metabolismo , Chaperonas Moleculares/metabolismo , Proteínas Portadoras/metabolismo , Citocinas/metabolismo , AntiinflamatoriosRESUMEN
Janus kinases (JAKs) have a key role in regulating the expression and function of relevant inflammatory cytokines involved in asthma and chronic obstructive pulmonary disease. Herein are described the design, synthesis, and pharmacological evaluation of a series of novel purinone JAK inhibitors with profiles suitable for inhaled administration. Replacement of the imidazopyridine hinge binding motif present in the initial compounds of this series with a pyridone ring resulted in the mitigation of cell cytotoxicity. Further systematic structure-activity relationship (SAR) efforts driven by structural biology studies led to the discovery of pyridone 34, a potent pan-JAK inhibitor with good selectivity, long lung retention time, low oral bioavailability, and proven efficacy in the lipopolysaccharide-induced rat model of airway inflammation by the inhaled route.
Asunto(s)
Imidazoles/química , Inhibidores de las Cinasas Janus/farmacología , Quinasas Janus/antagonistas & inhibidores , Piridinas/química , Piridonas/química , Enfermedades Respiratorias/tratamiento farmacológico , Administración por Inhalación , Animales , Humanos , Inhibidores de las Cinasas Janus/administración & dosificación , Inhibidores de las Cinasas Janus/química , Inhibidores de las Cinasas Janus/uso terapéutico , Enfermedad Pulmonar Obstructiva Crónica/tratamiento farmacológico , Ratas , Relación Estructura-ActividadRESUMEN
AZD8871 is a novel muscarinic antagonist and ß 2-adrenoceptor agonist in development for chronic obstructive pulmonary disease. This study describes the pharmacological profile of AZD8871 in in vitro and in vivo assays. AZD8871 is potent at the human M3 receptor (pIC50 in binding assays: 9.5) and shows kinetic selectivity for the M3 (half-life: 4.97 hours) over the M2 receptor (half-life: 0.46 hour). It is selective for the ß 2-adrenoceptor over the ß 1 and ß 3 subtypes (3- and 6-fold, respectively) and shows dual antimuscarinic and ß 2-adrenoceptor functional activity in isolated guinea pig tissue (pIC50 in electrically stimulated trachea: 8.6; pEC50 in spontaneous tone isolated trachea: 8.8, respectively), which are sustained over time. AZD8871 exhibits a higher muscarinic component than batefenterol in human bronchi, with a shift in potency under propranolol blockade of 2- and 6-fold, respectively, together with a persisting relaxation (5.3% recovery at 8 hours). Nebulized AZD8871 prevents acetylcholine-induced bronchoconstriction in both guinea pig and dog with minimal effects on salivation and heart rate at doses with bronchoprotective activity. Moreover, AZD8871 shows long-lasting effects in dog, with a bronchoprotective half-life longer than 24 hours. In conclusion, these studies demonstrate that AZD8871 is a dual-acting molecule with a high muscarinic component and a long residence time at the M3 receptor; moreover, its preclinical profile in animal models suggests a once-daily dosing in humans and a favorable safety profile. Thus, AZD8871 has the potential to be a next generation of inhaled bronchodilators in respiratory diseases.
Asunto(s)
Agonistas de Receptores Adrenérgicos beta 2/efectos adversos , Agonistas de Receptores Adrenérgicos beta 2/farmacología , Antagonistas Muscarínicos/efectos adversos , Antagonistas Muscarínicos/farmacología , Quinolinas/efectos adversos , Quinolinas/farmacología , Receptor Muscarínico M3/antagonistas & inhibidores , Receptores Adrenérgicos beta 2/metabolismo , Seguridad , Triazoles/efectos adversos , Triazoles/farmacología , Administración por Inhalación , Agonistas de Receptores Adrenérgicos beta 2/administración & dosificación , Agonistas de Receptores Adrenérgicos beta 2/farmacocinética , Animales , Bronquios/efectos de los fármacos , Bronquios/fisiología , Sistema Cardiovascular/efectos de los fármacos , Perros , Cobayas , Humanos , Masculino , Antagonistas Muscarínicos/administración & dosificación , Antagonistas Muscarínicos/farmacocinética , Quinolinas/administración & dosificación , Quinolinas/farmacocinética , Receptor Muscarínico M2/metabolismo , Distribución Tisular , Tráquea/efectos de los fármacos , Tráquea/fisiología , Triazoles/administración & dosificación , Triazoles/farmacocinéticaRESUMEN
The Janus-activated kinase (JAK) family together with signal transducer and activator of transcription (STAT) signaling pathway has a key role in regulating the expression and function of many inflammatory cytokines. This has led to the discovery of JAK inhibitors for the treatment of inflammatory diseases, some of them already in the market. Considering the adverse effects associated with JAK inhibition by oral route, we wanted to explore whether JAK inhibition by inhaled route is enough to inhibit airway inflammation. The aim of this study was to characterize the enzymatic and cellular potency and the selectivity of LAS194046, a novel JAK inhibitor, compared with the reference compounds ruxolitinib and tofacitinib. The efficacy of this new JAK inhibitor is described in a model of ovalbumin (OVA)-induced airway inflammation in Brown Norway rats by inhaled administration. As potential markers of target engagement, we assessed the effect of LAS194046 on the STAT activation state. LAS194046 is a selective inhaled pan-JAK inhibitor that reduces allergen-induced airway inflammation, late asthmatic response, and phosphor-STAT activation in the rat OVA model. Our results show that topical inhibition of JAK in the lung, without relevant systemic exposure, is sufficient to reduce lung inflammation and improve lung function in a rat asthma model. In summary, JAK-STAT pathway inhibition by inhaled route constitutes a promising therapeutic option for lung inflammatory diseases.
Asunto(s)
Alérgenos/inmunología , Asma/tratamiento farmacológico , Asma/inmunología , Inhibidores de las Cinasas Janus/farmacología , Quinasas Janus/antagonistas & inhibidores , Nitrilos/farmacología , Fosfoproteínas/metabolismo , Piperidinas/farmacología , Pirimidinas/farmacología , Factores de Transcripción STAT/metabolismo , Administración por Inhalación , Animales , Asma/metabolismo , Asma/patología , Inflamación/tratamiento farmacológico , Isoenzimas/antagonistas & inhibidores , Inhibidores de las Cinasas Janus/administración & dosificación , Inhibidores de las Cinasas Janus/farmacocinética , Inhibidores de las Cinasas Janus/uso terapéutico , Masculino , Nitrilos/administración & dosificación , Nitrilos/farmacocinética , Nitrilos/uso terapéutico , Piperidinas/administración & dosificación , Piperidinas/farmacocinética , Piperidinas/uso terapéutico , Pirimidinas/administración & dosificación , Pirimidinas/farmacocinética , Pirimidinas/uso terapéutico , Ratas , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos , Factores de TiempoRESUMEN
BACKGROUND AND PURPOSE: The Janus Kinase (JAK) family mediates the cytokine receptor-induced signalling pathways involved in inflammatory processes. The activation of the signal transducers and activators of transcription (STATs) by JAK kinases is a key point in these pathways. Four JAK proteins, JAK1, JAK2, JAK3 and tyrosine kinase 2 (Tyk2) associate with the intracellular domains of surface cytokine receptors are phosphorylating STATs and modulating gene expression. The aim of this study was to explore the role of JAK inhibition in an acute model of inhaled lipopolysaccharide (LPS)-induced airway inflammation in rats through evaluating the effects of tofacitinib, a marketed pan-JAK inhibitor. Specifically, some pulmonary inflammation parameters were studied and the lung STAT3 phosphorylation was assessed as a target engagement marker of JAK inhibition in the model. EXPERIMENTAL APPROACH: Rats were exposed to an aerosol of LPS (0.1 mg/ml) or phosphate-buffered saline (PBS) during 40 min. Bronchoalveolar lavage fluid (BALF) and lung samples were collected 4 h after PBS or LPS exposure. Neutrophils in BALF were counted and a panel of cytokines were measured in BALF. Phosphorylation of STAT3 was studied in lung homogenates by ELISA and localization of phospho-STAT3 (pSTAT3) in lung tissue was also evaluated by immunohistochemistry. In order to assess the effect of JAK inhibition, tofacitinib was administered 1 h before challenge at doses of 3, 10 and 30 mg/kg p.o. KEY RESULTS: Inhaled LPS challenge induced an augment of neutrophils and cytokines in the BALF as well as an increase in pSTAT3 expression in the lungs. Tofacitinib by oral route inhibited the LPS-induced airway neutrophilia, the levels of some cytokines in the BALF and the phosphorylation of STAT3 in the lung tissue. CONCLUSIONS AND IMPLICATIONS: In summary, this study shows that JAK inhibition ameliorates inhaled LPS-induced airway inflammation in rats, suggesting that at least JAK/STAT3 signalling is involved in the establishment of the pulmonary neutrophilia induced by LPS. JAKs inhibitors should be further investigated as a potential therapy for respiratory inflammatory diseases.
Asunto(s)
Inflamación/tratamiento farmacológico , Neutrófilos/metabolismo , Piperidinas/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Pirimidinas/farmacología , Pirroles/farmacología , Animales , Líquido del Lavado Bronquioalveolar/química , Citocinas/metabolismo , Modelos Animales de Enfermedad , Inflamación/patología , Quinasas Janus/antagonistas & inhibidores , Lipopolisacáridos/administración & dosificación , Pulmón/metabolismo , Masculino , Fosforilación/efectos de los fármacos , Neumonía/tratamiento farmacológico , Neumonía/patología , Ratas , Ratas Sprague-Dawley , Factor de Transcripción STAT3/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
Abediterol is a novel long-acting ß2-adrenoceptor agonist (LABA) currently in development for once-daily combination maintenance therapy of asthma and COPD. This study investigated the preclinical profile of abediterol in terms of affinity, potency, selectivity, duration of action and cardiac effects in comparison to the marketed once-daily LABAs indacaterol, olodaterol and vilanterol. Abediterol was the compound with the highest in vitro potency for dog, guinea pig and human ß2-adrenoceptors. In electrical field stimulated guinea pig trachea, abediterol demonstrated 5-, 44- and 77-fold greater potency than olodaterol, indacaterol and vilanterol, respectively. In anaesthetised guinea pigs, inhaled abediterol was also the most potent compound, with 5-20 times higher bronchoprotective potency than other once-daily LABAs against acetylcholine. The bronchoprotective half-life of abediterol in guinea pigs was 36h compared with 51h for indacaterol, 47h for olodaterol, and 18h for vilanterol. In anaesthetised dogs, abediterol also inhibited acetylcholine-induced bronchoconstriction, with higher potency than olodaterol and vilanterol [ID40 (dose inhibiting bronchoconstriction by 40%) of 0.059µg/kg, 0.180µg/kg and 2.870µg/kg, respectively]. In parallel, effects on heart rate in dogs were also measured. Abediterol showed greater safety index (defined as the ratio of the maximal dose without effect on heart rate and the ID40) than olodaterol and vilanterol (10.5 versus 4.9 and 2.4, respectively). Taken together, these data suggest that abediterol offers potent bronchodilation and a sustained duration of action suited to once-daily dosing, plus a reduced potential for class-related cardiac side effects.
Asunto(s)
Agonistas de Receptores Adrenérgicos beta 2/administración & dosificación , Agonistas de Receptores Adrenérgicos beta 2/farmacología , Benzoxazinas/farmacología , Alcoholes Bencílicos/farmacología , Clorobencenos/farmacología , Indanos/farmacología , Quinolonas/administración & dosificación , Quinolonas/farmacología , Administración por Inhalación , Agonistas de Receptores Adrenérgicos beta 2/efectos adversos , Animales , Perros , Cobayas , Masculino , Quinolonas/efectos adversos , Receptores Adrenérgicos beta 1/metabolismo , Receptores Adrenérgicos beta 2/metabolismo , Seguridad , Factores de TiempoRESUMEN
Asthma is increasing globally and current treatments only manage a proportion of patients. There is an urgent need to develop new therapies. Lymphocytes are thought to play a central role in the pathophysiology of asthma through the production of inflammatory mediators. This is thought to be via the transcription factor NFAT which in turn can be activated through Ca(2+) release-activated Ca(2+) (CRAC) channels. The aim of this work was to investigate the role of CRAC in clinical and pre-clinical models of allergic asthma. Initial data demonstrated that the NFAT pathway is increased in stimulated lymphocytes from asthmatics. To confirm a role for the channel we showed that a selective inhibitor, Synta 66, blocked mediator production from lymphocytes. Synta 66 inhibited CD2/3/28 induced IL-2, IL-7, IL-13 & IFNΥ in a concentration-dependent manner in healthy and severe asthma donors, with over 60% inhibition observed for all cytokines. NFAT pathway was also increased in a pre-clinical asthma model. In this model we have demonstrated that CRAC played a central role in the airway inflammation and late asthmatic response (LAR). In conclusion, our data provides evidence that suggests targeting CRAC channels could be of therapeutic benefit for asthma sufferers.
Asunto(s)
Asma/metabolismo , Canales de Calcio/metabolismo , Adulto , Alérgenos/toxicidad , Animales , Asma/patología , Líquido del Lavado Bronquioalveolar/citología , Canales de Calcio/efectos de los fármacos , Células Cultivadas , Citocinas/biosíntesis , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Técnicas In Vitro , Pulmón/patología , Linfocitos/efectos de los fármacos , Linfocitos/metabolismo , Masculino , Persona de Mediana Edad , Factores de Transcripción NFATC/metabolismo , Neutrófilos/efectos de los fármacos , Neutrófilos/metabolismo , RatasRESUMEN
Fibrotic lung diseases, such as idiopathic pulmonary fibrosis, are associated with spontaneous dry cough and hypersensitivity to tussive agents. Understanding the pathophysiology driving enhanced cough may help us to define better therapies for patients. We hypothesized that lung fibrosis induced by intratracheal bleomycin would exacerbate the cough reflex induced by tussive agents in guinea pigs. Disease progression in the lungs was characterized at days 1, 7, 14, 21 and 28 after bleomycin administration. Inflammatory and fibrotic markers, as well as neurotrophin levels, were assessed in bronchoalveolar lavage fluid and/or lung tissue. Cough sensitivity to citric acid, capsaicin and allylisothiocyanate was evaluated in conscious animals at days 14 and 21 after bleomycin administration. Pulmonary lesions evolved from an early inflammatory phase (from day 1 to day 7) to a fibrotic stage (between days 14 and 28). Fibrosis was related to increased levels of matrix metalloproteinase-2 in bronchoalveolar lavage fluid (day 21: saline, 0.26 ng/ml; bleomycin, 0.49 ng/ml). At day 14, we also observed increased cough reflexes to citric acid (163%), capsaicin (125%) and allylisothiocyanate (178%). Cough exacerbation persisted, but at a lower extent, by day 21 for capsaicin (100%) and allylisothiocyanate (54%). Moreover, bronchoalveolar lavage fluid concentrations of brain-derived neurotrophic factor, suggested to induce nerve remodelling in chronic cough, were also enhanced (day 1: saline, 14.21 pg/ml; bleomycin, 30.09 pg/ml). In summary, our model of bleomycin-induced cough exacerbation may be a valuable tool to investigate cough hypersensitivity and develop antitussive therapies for fibrotic lung diseases.
Asunto(s)
Bleomicina , Tos/fisiopatología , Pulmón/inervación , Fibrosis Pulmonar/fisiopatología , Reflejo Anormal , Animales , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Líquido del Lavado Bronquioalveolar/química , Capsaicina , Ácido Cítrico , Tos/inducido químicamente , Tos/genética , Tos/metabolismo , Citocinas/genética , Citocinas/metabolismo , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Regulación de la Expresión Génica , Cobayas , Canales Iónicos/genética , Canales Iónicos/metabolismo , Isotiocianatos , Pulmón/metabolismo , Pulmón/patología , Masculino , Metaloproteinasa 2 de la Matriz/metabolismo , Fibrosis Pulmonar/inducido químicamente , Fibrosis Pulmonar/genética , Fibrosis Pulmonar/metabolismo , Factores de TiempoRESUMEN
RNA viruses are a major cause of respiratory infections and are known to exacerbate asthma and other respiratory diseases. Our aim was to test the ability of poly(I:C) (polyinosinic:polycytidylic acid), a viral surrogate, to elicit exacerbation in a model of severe asthma driven by HDM (house dust mite) in FCA (Freund's complete adjuvant). Poly(I:C) was administered intranasally around the HDM challenge in FCA-HDM-sensitized animals. Changes in AHR (airway hyperresponsiveness), BALF (bronchoalveolar lavage fluid) inflammatory infiltrate, HDM-specific immunoglobulins and cytokine/chemokine release were evaluated at different points after the challenge. The effect of oral dexamethasone was also assessed. Exacerbation was achieved when poly(I:C) was administered 24 h before the HDM challenge and was characterized by enhanced AHR and an increase in the numbers of neutrophils, macrophages and lymphocytes in the BALF. Th1, Th2 and Th17 cytokines were also elevated at different time points after the challenge. Peribronchial and alveolar inflammation in lung tissue were also augmented. AHR and inflammatory infiltration showed reduced sensitivity to dexamethasone treatment. We have set up a model that mimics key aspects of viral exacerbation in a corticosteroid-refractory asthmatic phenotype which could be used to evaluate new therapies for this condition.
Asunto(s)
Corticoesteroides/farmacología , Asma/inducido químicamente , Hiperreactividad Bronquial/inducido químicamente , Dexametasona/farmacología , Resistencia a Medicamentos , Pulmón/efectos de los fármacos , Poli I-C/toxicidad , Animales , Antígenos Dermatofagoides , Proteínas de Artrópodos , Asma/tratamiento farmacológico , Asma/inmunología , Asma/metabolismo , Asma/fisiopatología , Hiperreactividad Bronquial/inmunología , Hiperreactividad Bronquial/fisiopatología , Líquido del Lavado Bronquioalveolar/química , Líquido del Lavado Bronquioalveolar/citología , Broncoconstricción/efectos de los fármacos , Quimiotaxis de Leucocito/efectos de los fármacos , Cisteína Endopeptidasas , Citocinas/metabolismo , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Relación Dosis-Respuesta a Droga , Inmunoglobulinas/sangre , Mediadores de Inflamación/metabolismo , Pulmón/inmunología , Pulmón/metabolismo , Pulmón/fisiopatología , Masculino , Ratones Endogámicos BALB C , Infiltración Neutrófila/efectos de los fármacos , Fenotipo , Índice de Severidad de la Enfermedad , Factores de TiempoRESUMEN
Spleen tyrosine kinase (Syk) is essential for signal transduction of immunoreceptors. Inhibition of Syk abrogates mast cell degranulation and B cell responses. We hypothesized that Syk inhibition in the lung by inhaled route could block airway mast cells degranulation and the early asthmatic response without the need of systemic exposure. We discovered LAS189386, a novel Syk inhibitor with suitable properties for inhaled administration. The aim of this study was to characterize the in vitro and in vivo profile of LAS189386. The compound was profiled in Syk enzymatic assay, against a panel of selected kinases and in Syk-dependent cellular assays in mast cells and B cells. Pharmacokinetics and in vivo efficacy was assessed by intratracheal route. Airway resistance and mast cell degranulation after OVA challenge was evaluated in an ovalbumin-sensitized Brown Norway rat model. LAS189386 potently inhibits Syk enzymatic activity (IC50 7.2 nM), Syk phosphorylation (IC50 41 nM), LAD2 cells degranulation (IC50 56 nM), and B cell activation (IC50 22 nM). LAS189386 inhibits early asthmatic response and airway mast cell degranulation without affecting systemic mast cells. The present results support the hypothesis that topical inhibition of Syk in the lung, without systemic exposure, is sufficient to inhibit EAR in rats. Syk inhibition by inhaled route constitutes a promising therapeutic option for asthma.
Asunto(s)
Asma/prevención & control , Compuestos de Azabiciclo/administración & dosificación , Indazoles/administración & dosificación , Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Mastocitos/efectos de los fármacos , Inhibidores de Proteínas Quinasas/administración & dosificación , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Administración por Inhalación , Animales , Asma/patología , Asma/fisiopatología , Compuestos de Azabiciclo/farmacocinética , Linfocitos B/efectos de los fármacos , Linfocitos B/fisiología , Degranulación de la Célula/efectos de los fármacos , Línea Celular , Modelos Animales de Enfermedad , Humanos , Indazoles/farmacocinética , Masculino , Mastocitos/fisiología , Inhibidores de Proteínas Quinasas/farmacocinética , Ratas , Ratas Endogámicas BN , Ratas Wistar , Quinasa SykRESUMEN
Synthesis and SAR of a series of 7-azaindoles as Orai channel inhibitors showing good potency inhibiting IL-2 production in Jurkat cells is described. Compound 14d displaying best pharmacokinetic properties was further characterized in a model of allergen induced asthma showing inhibition in the number of eosinophils in BALF. High lipophilicity remains as one of the main challenges for this class of compounds.
Asunto(s)
Bloqueadores de los Canales de Calcio/química , Canales de Calcio/química , Indoles/química , Piridinas/química , Pirroles/química , Animales , Asma , Compuestos Aza/química , Bloqueadores de los Canales de Calcio/farmacocinética , Bloqueadores de los Canales de Calcio/uso terapéutico , Canales de Calcio/metabolismo , Modelos Animales de Enfermedad , Evaluación Preclínica de Medicamentos , Semivida , Humanos , Hipersensibilidad/tratamiento farmacológico , Hipersensibilidad/inmunología , Indoles/farmacocinética , Indoles/uso terapéutico , Interleucina-2/sangre , Interleucina-2/metabolismo , Células Jurkat , Microsomas/metabolismo , Modelos Biológicos , Ovalbúmina/inmunología , Unión Proteica , Piridinas/farmacocinética , Piridinas/uso terapéutico , Pirroles/farmacocinética , Pirroles/uso terapéutico , Ratas , Relación Estructura-ActividadRESUMEN
Abediterol is a novel potent, long-acting inhaled ß(2)-adrenoceptor agonist in development for the treatment of asthma and chronic obstructive pulmonary disease. Abediterol shows subnanomolar affinity for the human ß(2)-adrenoceptor and a functional selectivity over ß(1)-adrenoceptors higher than that of formoterol and indacaterol in both a cellular model with overexpressed human receptors and isolated guinea pig tissue. Abediterol is a full agonist at the human ß(2)-adrenoceptor (E(max) = 91 ± 5% of the maximal effect of isoprenaline). The potency and onset of action that abediterol shows in isolated human bronchi (EC(50) = 1.9 ± 0.4 nM; t½ onset = 7-10 min) is not significantly different from that of formoterol, but its duration of action (t½ â¼ 690 min) is similar to that of indacaterol. Nebulized abediterol inhibits acetylcholine-induced bronchoconstriction in guinea pigs in a concentration-dependent manner, with higher potency and longer duration of action (t½ = 36 h) than salmeterol (t½ = 6 h) and formoterol (t½ = 4 h) and similar duration of action to indacaterol up to 48 h. In dogs, the bronchoprotective effect of abediterol is more sustained than that of salmeterol and indacaterol at doses without effects on heart rate, thus showing a greater safety margin (defined as the ratio of dose increasing heart rate by 5% and dose inhibiting bronchospasm by 50%) than salmeterol, formoterol, and indacaterol (5.6 versus 3.3, 2.2, and 0.3, respectively). In conclusion, our results suggest that abediterol has a preclinical profile for once-daily dosing in humans together with a fast onset of action and a favorable cardiovascular safety profile.
Asunto(s)
Agonistas de Receptores Adrenérgicos beta 2/farmacología , Broncodilatadores/farmacología , Receptores Adrenérgicos beta 2/metabolismo , Albuterol/análogos & derivados , Albuterol/farmacología , Animales , Asma/tratamiento farmacológico , Asma/metabolismo , Bronquios/efectos de los fármacos , Bronquios/metabolismo , Broncoconstricción/efectos de los fármacos , Perros , Relación Dosis-Respuesta a Droga , Etanolaminas/farmacología , Fumarato de Formoterol , Cobayas , Humanos , Masculino , Monocitos/efectos de los fármacos , Monocitos/patología , Quinolonas/farmacología , Xinafoato de SalmeterolRESUMEN
BACKGROUND: In allergic asthma, exposure to relevant antigens leads to an early asthmatic response (EAR) followed, in certain subjects, by a late asthmatic response (LAR). Although many subjects with asthma consider LAR to be one of the defining symptoms of their disease, and despite its widespread use in the clinical assessment of new therapeutic entities, the mechanism underlying the LAR remains unclear. METHOD: A study was undertaken using ovalbumin-sensitised and challenged Brown Norway rat and C57BL/6J mouse models which recapitulate phenotypic features of allergic asthma including the LAR and its susceptibility to clinically effective agents. RESULTS: In conscious animals an EAR was followed by a LAR. The LAR was subjectively evidenced by audible (wheeze) and visual signs of respiratory distress associated with quantifiable changes in non-invasive lung function assessment. Treatments that attenuated the EAR failed to impact on the LAR and, while anaesthesia did not impact on EAR, it abolished LAR. A key role for airway sensory neuronal reflexes in the LAR was therefore hypothesised, which was confirmed by the blockade observed after administration of ruthenium red (non-selective cation channel blocker), HC-030031 (TRPA1 inhibitor) and tiotropium bromide (anticholinergic) but not JNJ-17203212 (TRPV1 inhibitor). CONCLUSION: These results suggest that LAR involves the following processes: allergen challenge triggering airway sensory nerves via the activation of TRPA1 channels which initiates a central reflex event leading to a parasympathetic cholinergic constrictor response. These data are supported by recent clinical trials suggesting that an anticholinergic agent improved symptoms and lung function in patients with asthma.
Asunto(s)
Asma/fisiopatología , Bronquios/inervación , Broncoconstricción/fisiología , Sistema Nervioso Parasimpático/fisiopatología , Células Receptoras Sensoriales/fisiología , Acetanilidas/farmacología , Animales , Asma/inducido químicamente , Bronquios/efectos de los fármacos , Bronquios/fisiopatología , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Masculino , Ratones , Ratones Endogámicos C57BL , Ovalbúmina/administración & dosificación , Ovalbúmina/inmunología , Ovalbúmina/toxicidad , Sistema Nervioso Parasimpático/efectos de los fármacos , Purinas/farmacología , Ratas , Ratas Endogámicas BN , Rojo de Rutenio/farmacología , Células Receptoras Sensoriales/efectos de los fármacos , Canales de Potencial de Receptor Transitorio/antagonistas & inhibidoresRESUMEN
The liver X receptors (LXRalpha/beta) are orphan nuclear receptors that are expressed in a large number of cell types and have been shown to have anti-inflammatory properties. Nuclear receptors have previously proved to be amenable targets for small molecular mass pharmacological agents in asthma, and so the effect of an LXR ligand was assessed in models of allergic airway inflammation. LXR agonist, GW 3965, was profiled in rat and mouse models of allergic asthma. In the Brown Norway rats, GW 3965 (3-30 mg/kg) was unable to reduce the bronchoalveolar lavage eosinophilia associated with this model and had no impact on inflammatory biomarkers (eotaxin and IL-1beta). The compound did significantly stimulate ABCA-1 (ATP-binding cassette A1) mRNA expression, indicating that there was adequate exposure/LXR activation. In the mouse model, the LXR ligand surprisingly increased airway reactivity, an effect that was apparent in both the Ag and nonchallenged groups. This increase was not associated with a change in lung tissue inflammation or number of mucus-containing cells. There was, however, a marked increase in airway smooth muscle thickness in both treated groups. We demonstrated an increase in contractile response to exogenous methacholine in isolated airways taken from LXR agonist-treated animals compared with the relevant control tissue. We corroborated these findings in a human system by demonstrating increased proliferation of cultured airway smooth muscle. This phenomenon, if evidenced in man, would indicate that LXR ligands may directly increase airway reactivity, which could be detrimental, especially in patients with existing respiratory disease and with already compromised lung function.
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Asma/inmunología , Asma/metabolismo , Benzoatos/administración & dosificación , Bencilaminas/administración & dosificación , Hiperreactividad Bronquial/metabolismo , Proteínas de Unión al ADN/agonistas , Músculo Liso/crecimiento & desarrollo , Músculo Liso/metabolismo , Receptores Citoplasmáticos y Nucleares/agonistas , Regulación hacia Arriba/inmunología , Animales , Antiinflamatorios no Esteroideos/administración & dosificación , Asma/patología , Asma/fisiopatología , Hiperreactividad Bronquial/inmunología , Hiperreactividad Bronquial/fisiopatología , Proliferación Celular/efectos de los fármacos , Proteínas de Unión al ADN/fisiología , Relación Dosis-Respuesta Inmunológica , Humanos , Receptores X del Hígado , Masculino , Ratones , Ratones Endogámicos BALB C , Músculo Liso/inmunología , Receptores Nucleares Huérfanos , Ovalbúmina/administración & dosificación , Ovalbúmina/inmunología , Ratas , Ratas Endogámicas BN , Receptores Citoplasmáticos y Nucleares/fisiología , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Estrogen receptor (ER) beta agonists have been demonstrated to possess anti-inflammatory properties in inflammatory disease models. The objective of this study was to determine whether ERbeta agonists affect in vitro and in vivo preclinical models of asthma. mRNA expression assays were validated in human and rodent tissue panels. These assays were then used to measure expression in human cells and our characterized rat model of allergic asthma. ERB-041 [7-ethenyl-2-(3-fluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol], an ERbeta agonist, was profiled on cytokine release from interleukin-1beta-stimulated human airway smooth muscle (HASM) cells and in the rodent asthma model. Although ERbeta expression was demonstrated at the gene and protein level in HASM cells, the agonist failed to have an impact on the inflammatory response. Similarly, in vivo, we observed temporal modulation of ERbeta expression after antigen challenge. However, the agonist failed to have an impact on the model endpoints such as airway inflammation, even though plasma levels reflected linear compound exposure and was associated with an increase in receptor activation after drug administration. In these modeling systems of airway inflammation, an ERbeta agonist was ineffective. Although ERbeta agonists are anti-inflammatory in certain models, this novel study would suggest that they would not be clinically useful in the treatment of asthma.
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Receptor beta de Estrógeno/biosíntesis , Perfilación de la Expresión Génica , Mediadores de Inflamación/fisiología , Animales , Asma/tratamiento farmacológico , Asma/genética , Asma/metabolismo , Células Cultivadas , Receptor beta de Estrógeno/agonistas , Receptor beta de Estrógeno/genética , Humanos , Inflamación/metabolismo , Inflamación/prevención & control , Mediadores de Inflamación/metabolismo , Masculino , Oxazoles/farmacología , Oxazoles/uso terapéutico , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Ratas , Ratas Endogámicas BNRESUMEN
Since the discovery of the first matrix metalloproteinase (MMP), this ever-growing family of proteinases has been the subject of intense research. Although it was initially believed that MMPs were solely involved in matrix turnover and degradation, there are now data suggesting MMPs are actively involved in the inflammatory process. In previous studies, we have demonstrated an increase in MMP expression in human cell-based assays and in preclinical rat models of airway inflammation. Therefore, the aim of this study was to characterize the role of MMPs in these models by profiling the impact of a broad-spectrum MMP inhibitor. In lipopolysaccharide (LPS)-stimulated THP-1 cells and primary human lung tissue macrophages, the MMP inhibitor had no significant effect on the release of tumor necrosis factor-alpha, interleukin (IL)-8, IL-1 beta, growth-regulated oncogene-alpha, macrophage inflammatory protein-1 alpha, or IL-6 whereas dexamethasone has a significant impact on all cytokines from both cell types. Similarly, in the more biologically complex LPS-driven rat model of airway inflammation, the MMP inhibitor did not have an impact on mediator release and cellular burden. The compound did, however, significantly reduce levels of lung MMP-9. Furthermore, in a "disease" model, the compound did not affect cellular inflammation but did significantly reduce elastase-induced experimental emphysema. In summary, these data demonstrate for the first time that MMPs do not play a role in the increase in inflammatory mediators or cellular burden observed in these preclinical models. However, they do appear to be involved in the elastase-driven breakdown of airway structure, which is not due to a direct effect of the stimulus.
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Inflamación/enzimología , Metaloproteinasas de la Matriz/fisiología , Enfermedades Respiratorias/enzimología , Animales , Biomarcadores/metabolismo , Líquido del Lavado Bronquioalveolar/citología , Línea Celular , Citocinas/metabolismo , Humanos , Inflamación/inducido químicamente , Inflamación/patología , Lipopolisacáridos , Pulmón/citología , Pulmón/enzimología , Pulmón/metabolismo , Macrófagos/efectos de los fármacos , Macrófagos/enzimología , Macrófagos/metabolismo , Masculino , Inhibidores de la Metaloproteinasa de la Matriz , Páncreas/enzimología , Elastasa Pancreática , Inhibidores de Proteasas/farmacología , Ratas , Ratas Wistar , Enfermedades Respiratorias/inducido químicamente , Enfermedades Respiratorias/patología , PorcinosRESUMEN
Clinically, inflammatory pain is far more persistent than that typically modelled pre-clinically, with the majority of animal models focussing on short-term effects of the inflammatory pain response. The large attrition rate of compounds in the clinic which show pre-clinical efficacy suggests the need for novel models of, or approaches to, chronic inflammatory pain if novel mechanisms are to make it to the market. A model in which a more chronic inflammatory hypersensitivity phenotype is profiled may allow for a more clinically predictive tool. The aims of these studies were to characterise and validate a chronic model of inflammatory pain. We have shown that injection of a large volume of adjuvant to the intra-articular space of the rat knee results in a prolonged inflammatory pain response, compared to the response in an acute adjuvant model. Additionally, this model also results in a hypersensitive state in the presence and absence of inflammation. A range of clinically effective analgesics demonstrate activity in this chronic model, including morphine (3mg/kg, t.i.d.), dexamethasone (1mg/kg, b.i.d.), ibuprofen (30mg/kg, t.i.d.), etoricoxib (5mg/kg, b.i.d.) and rofecoxib (0.3-10mg/kg, b.i.d.). A further aim was to exemplify the utility of this chronic model over the more acute intra-plantar adjuvant model using two novel therapeutic approaches; NR2B selective NMDA receptor antagonism and iNOS inhibition. Our data shows that different effects were observed with these therapies when comparing the acute model with the model of chronic inflammatory joint pain. These data suggest that the chronic model may be more relevant to identifying mechanisms for the treatment of chronic inflammatory pain states in the clinic.
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Artritis/tratamiento farmacológico , Artritis/patología , Modelos Animales de Enfermedad , Dolor/tratamiento farmacológico , Dolor/patología , Animales , Artritis/inducido químicamente , Enfermedad Crónica , Inhibidores de la Ciclooxigenasa 2/uso terapéutico , Adyuvante de Freund , Lactonas/uso terapéutico , Masculino , Óxido Nítrico Sintasa de Tipo II/antagonistas & inhibidores , Dolor/inducido químicamente , Fenoles/uso terapéutico , Piperidinas/uso terapéutico , Ratas , Receptores de N-Metil-D-Aspartato/antagonistas & inhibidores , Rodilla de Cuadrúpedos/fisiología , Sulfuros/uso terapéutico , Sulfonas/uso terapéutico , Factores de Tiempo , Soporte de PesoRESUMEN
Nitric oxide (NO), synthesised by different isoforms of nitric oxide synthase (NOS), has been linked with the development and maintenance of nociception. We studied the role of the inducible isoform, iNOS, in two different rat pain models with an inflammatory component. iNOS was immunohistochemically detected locally in the paw 6h after Freund's Complete Adjuvant (FCA) injection, showing a plateau at 24-72 h and falling slowly in the following weeks. This correlated with the late phase of the hypersensitivity to pain revealed in the behavioural tests. A highly selective iNOS inhibitor GW274150 (1-30 mg/kg orally, 24h after FCA) suppressed the accumulation of nitrite in the inflamed paw indicating substantial iNOS inhibition. At the same time it partially reversed FCA-induced hypersensitivity to pain and edema in a dose-dependent manner. After Chronic Constriction Injury (CCI) surgery to the sciatic nerve, iNOS presence was only detected locally in the region of the nerve (inflammatory cells). GW274150 (3-30 mg/kg orally, 21 days after surgery) also reversed significantly the CCI-associated hypersensitivity to pain. No iNOS was detectable in dorsal root ganglia, spinal cord or brain in either model. This study demonstrates a role for peripherally-expressed iNOS in pain conditions with an inflammatory component and the potential value of iNOS inhibitors in such conditions.
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Hiperalgesia/diagnóstico , Hiperalgesia/enzimología , Inflamación/tratamiento farmacológico , Inflamación/enzimología , Óxido Nítrico Sintasa de Tipo II/antagonistas & inhibidores , Óxido Nítrico Sintasa de Tipo II/metabolismo , Sulfuros/administración & dosificación , Analgésicos/administración & dosificación , Animales , Relación Dosis-Respuesta a Droga , Adyuvante de Freund , Hiperalgesia/inducido químicamente , Inflamación/inducido químicamente , Inflamación/diagnóstico , Masculino , Óxido Nítrico Sintasa/administración & dosificación , Ratas , Resultado del TratamientoRESUMEN
RATIONALE: Nuclear factor (NF)-kappaB is a transcription factor known to regulate the expression of many inflammatory genes, including cytokines, chemokines, and adhesion molecules. NF-kappaB is held inactive in the cytoplasm, bound to I-kappaB. The removal of I-kappaB, via the actions of inhibitor of kappaB (I-kappaB) kinase-2 (IKK-2), allows NF-kappaB to enter the nucleus. OBJECTIVES: To determine the impact of inhibiting IKK-2 on in vitro and in vivo models of airway inflammation. METHODS: The effect of inhibiting IKK-2 was assessed in stimulated, cultured, primary human airway smooth muscle cells and an antigen-driven rat model of lung inflammation. MEASUREMENTS: The release of cytokines from cultured cells and inflammatory cytokine expression and cellular burden in the lung were determined. MAIN RESULTS: Two structurally distinct molecules and dominant negative technology demonstrated that inhibition of IKK-2 activity completely blocked cytokine release from cultured cells, whereas the two glucocorticoid comparators had limited impact on granulocyte colony-stimulating factor, interleukin 8, and eotaxin release. In addition, in an in vivo antigen-driven model of airway inflammation, the IKK-2 inhibitor blocked NF-kappaB nuclear translocation, which was associated with a reduction in inflammatory cytokine gene and protein expression, airway eosinophilia, and late asthmatic reaction, similar in magnitude to that obtained with budesonide. CONCLUSION: This study demonstrates that inhibiting IKK-2 results in a general reduction of the inflammatory response in vitro and in vivo. Compounds of this class could have therapeutic utility in the treatment of asthma and may, in certain respects, possess a beneficial efficacy profile compared with that of a steroid.