RESUMEN
Treatment of the autoimmune demyelinating disease multiple sclerosis (MS) requires therapies that both limit and repair damage. While several immunomodulatory treatments exist to limit damage there are currently no treatments that promote the regenerative process of remyelination. A rapid way of screening potential pro-remyelination compounds is therefore required. The use of larval zebrafish in a drug reprofiling screen allows rapid in vivo screening and has been used successfully in the past as an efficient way of identifying new indications for existing drugs. A novel screening platform for potential pro-myelination compounds was developed using zebrafish larvae. Two percent of compounds screened from reprofiling libraries altered oligodendrocyte lineage cell recruitment and/or proliferation, as measured by the numbers of dorsally migrated spinal cord olig2(+) cells. Selective screening identified three compounds that altered levels of myelination, as measured by whole larvae myelin basic protein (mbp) transcript levels; the src family kinase inhibitor PP2, a biogenic amine and a thioxanthene. As well as many previously unrecognised compounds, identified compounds included those with previously known effects on myelin and/or the oligodendrocyte lineage, such as a PPAR agonist, steroid hormones and src family kinase inhibitors. As well as providing methods for further assessment of potentially beneficial compounds, this screen has highlighted 25 targets that are able to alter oligodendrocyte lineage cell recruitment or proliferation and/or mbp transcript levels in vivo and are worthy of further investigation for their potential effects on remyelination.
Asunto(s)
Diferenciación Celular/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Modelos Animales , Proteína Básica de Mielina/metabolismo , Oligodendroglía/metabolismo , Análisis de Varianza , Animales , Animales Modificados Genéticamente , Antiinflamatorios no Esteroideos/farmacología , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Recuento de Células/métodos , Diferenciación Celular/fisiología , Evaluación Preclínica de Medicamentos , Embrión no Mamífero , Regulación del Desarrollo de la Expresión Génica/fisiología , Proteínas Fluorescentes Verdes/genética , Proteína Básica de Mielina/genética , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Factor de Transcripción 2 de los Oligodendrocitos , Oligodendroglía/efectos de los fármacos , Piroxicam/análogos & derivados , Piroxicam/farmacología , Médula Espinal/citología , Pez Cebra , Proteínas de Pez Cebra/genética , Proteínas de Pez Cebra/metabolismoRESUMEN
Habituation, where a response is reduced when exposed to a continuous stimulus is one of the simplest forms of non-associative learning and has been shown in a number of organisms from sea slugs to rodents. However, very little has been reported in the zebrafish, a model that is gaining popularity for high-throughput compound screens. Furthermore, since most of the studies involving learning and memory in zebrafish have been conducted in adults, we sought to determine if zebrafish larvae could display non-associative learning and whether it could be modulated by compounds identified in previous rodent studies. We demonstrated that zebrafish larvae (7 days post fertilization) exhibit iterative reduction in a startle response to a series of acoustic stimuli. Furthermore, this reduction satisfied criteria for habituation: spontaneous recovery, more rapid reductions in startle to shorter intertrial intervals and dishabituation. We then investigated the pathways mediating this behavior using established compounds in learning and memory. Administration of rolipram (PDE4 inhibitor), donepezil (acetylcholinesterase inhibitor), and memantine (N-methyl-D-aspartic acid (NMDA) receptor antagonist) all increased the acoustic startle response and decreased habituation in the larvae, similar to previous rodent studies. Further studies demonstrated that NMDA blocked the memantine response and the effect of donepezil was blocked by mecamylamine but not atropine suggesting that the donepezil response was mediated by nicotinic rather than muscarinic receptors. Zebrafish larvae possess numerous advantages for medium to high-throughput screening; the model described herein therefore offers the potential to screen for additional compounds for further study on cognition function.
Asunto(s)
Habituación Psicofisiológica , Larva/fisiología , Reflejo de Sobresalto/fisiología , Estimulación Acústica/métodos , Factores de Edad , Análisis de Varianza , Animales , Área Bajo la Curva , Conducta Animal/fisiología , Relación Dosis-Respuesta a Droga , Relación Dosis-Respuesta en la Radiación , Inhibidores Enzimáticos/administración & dosificación , Agonistas de Aminoácidos Excitadores/farmacología , Antagonistas de Aminoácidos Excitadores/farmacología , Inhibición Psicológica , Memantina/farmacología , N-Metilaspartato/farmacología , Reflejo de Sobresalto/efectos de los fármacos , Pez CebraRESUMEN
Optimisation of ADS100380, a sub-micromolar HDAC inhibitor identified using a virtual screening approach, led to a series of substituted 5-(1H-pyrazol-3-yl)-thiophene-2-hydroxamic acids (6a-i), that possessed significant HDAC inhibitory activity. Subsequent functionalisation of the pendent phenyl group of compounds 6f and 6g provided analogues 6j-w with further enhanced enzyme and anti-proliferative activity. Compound 6j demonstrated efficacy in a mouse xenograft experiment.
Asunto(s)
Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Inhibidores de Histona Desacetilasas , Ácidos Hidroxámicos/síntesis química , Ácidos Hidroxámicos/farmacología , Pirazoles/síntesis química , Pirazoles/farmacología , Animales , Antineoplásicos/síntesis química , Antineoplásicos/farmacología , Disponibilidad Biológica , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Simulación por Computador , Evaluación Preclínica de Medicamentos , Inhibidores Enzimáticos/farmacocinética , Humanos , Indicadores y Reactivos , Ratones , Trasplante de Neoplasias , Neoplasias Experimentales/tratamiento farmacológico , Ratas , Relación Estructura-Actividad , Trasplante HeterólogoRESUMEN
Further investigation of a series of thienyl-based hydroxamic acids that included ADS100380 and ADS102550 led to the identification of the 5-pyridin-2-yl-thiophene-2-hydroxamic acid 3c, which possessed modest HDAC inhibitory activity. Substitution at the 5- and 6-positions of the pyridyl ring of compound 3c provided compounds 5a-g, 7a, b, 9, and 13a. Compound 5b demonstrated improved potency, in vitro DMPK profile, and rat oral bioavailability, compared to ADS102550. Functionalisation of the pendent phenyl group of compounds 5b, 5e and 13a provided analogues that possessed excellent enzyme inhibition and anti-proliferative activity.
Asunto(s)
Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Inhibidores de Histona Desacetilasas , Ácidos Hidroxámicos/síntesis química , Ácidos Hidroxámicos/farmacología , Tiofenos/síntesis química , Tiofenos/farmacología , Animales , Antineoplásicos/síntesis química , Antineoplásicos/farmacología , Disponibilidad Biológica , Células CACO-2 , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Citocromo P-450 CYP3A , Inhibidores Enzimáticos del Citocromo P-450 , Inhibidores Enzimáticos/farmacocinética , Humanos , Indicadores y Reactivos , Inyecciones Intravenosas , Ratones , Microsomas Hepáticos/efectos de los fármacos , Microsomas Hepáticos/enzimología , Microsomas Hepáticos/metabolismo , Ratas , Relación Estructura-ActividadRESUMEN
We have modified the previously reported 2-aminoquinoline 1 to provide two novel series of MCH-1R antagonists. Representative compounds from the quinazoline and benzimidazole series have been shown to be potent and selective, with promising in vitro eADME profiles.
Asunto(s)
Bencimidazoles/síntesis química , Quinazolinas/síntesis química , Receptores de Somatostatina/antagonistas & inhibidores , Bencimidazoles/farmacología , Humanos , Concentración 50 Inhibidora , Unión Proteica , Quinazolinas/farmacología , Relación Estructura-ActividadRESUMEN
Melanin-concentrating hormone (MCH) has been known to be an appetite-stimulating peptide for a number of years. However, it is only recently that MCH has been discovered to be the natural ligand for a previously "orphan" G-protein-coupled receptor, now designated MCH-1R. This receptor has been shown to mediate the effects of MCH on appetite and body weight, and consequently, drug discovery programs have begun to exploit this information in the search for MCH-1R antagonists for the treatment of obesity. In this paper, we report the rapid discovery of multiple, structurally distinct series of MCH-1R antagonists using a variety of virtual screening techniques. The most potent of these compounds (12) demonstrated an IC(50) value of 55 nM in the primary screen and exhibited antagonist properties in a functional cellular assay measuring Ca(2+) release. More potent compounds were identified by follow-up searches around the initial hit. A proposed binding mode for compound 12 in a homology model of the MCH-1R is also presented.
Asunto(s)
Piperazinas/química , Quinolinas/química , Receptores de Somatostatina/antagonistas & inhibidores , Secuencia de Aminoácidos , Animales , Células CHO , Calcio/metabolismo , Cricetinae , Bases de Datos Factuales , Humanos , Modelos Moleculares , Datos de Secuencia Molecular , Piperazinas/farmacología , Quinolinas/farmacología , Receptores de Somatostatina/química , Receptores de Somatostatina/metabolismo , Relación Estructura-ActividadRESUMEN
A new series of 2-aminoquinolines has been identified as antagonists of the melanin concentrating hormone receptor (MCH-1R). Syntheses and structure-activity relationships are described leading to a compound having low nanomolar activity against the receptor and demonstrating functional antagonism. Studies also showed that some of the compounds were selective against a range of other G protein-coupled receptors.