RESUMEN
Tau is a microtubule (MT)-stabilizing protein that is altered in Alzheimer's disease (AD) and other tauopathies. It is hypothesized that the hyperphosphorylated, conformationally altered, and multimeric forms of tau lead to a disruption of MT stability; however, direct evidence is lacking in vivo. In this study, an in vivo stable isotope-mass spectrometric technique was used to measure the turnover, or dynamicity, of MTs in brains of living animals. We demonstrated an age-dependent increase in MT dynamics in two different tau transgenic mouse models, 3xTg and rTg4510. MT hyperdynamicity was dependent on tau expression, since a reduction of transgene expression with doxycycline reversed the MT changes. Treatment of rTg4510 mice with the epothilone, BMS-241027, also restored MT dynamics to baseline levels. In addition, MT stabilization with BMS-241027 had beneficial effects on Morris water maze deficits, tau pathology, and neurodegeneration. Interestingly, pathological and functional benefits of BMS-241027 were observed at doses that only partially reversed MT hyperdynamicity. Together, these data suggest that tau-mediated loss of MT stability may contribute to disease progression and that very low doses of BMS-241027 may be useful in the treatment of AD and other tauopathies.
Asunto(s)
Trastornos del Conocimiento/tratamiento farmacológico , Epotilonas/uso terapéutico , Microtúbulos/patología , Degeneración Nerviosa/tratamiento farmacológico , Tauopatías/tratamiento farmacológico , Moduladores de Tubulina/uso terapéutico , Proteínas tau/fisiología , Animales , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Encéfalo/patología , Trastornos del Conocimiento/complicaciones , Trastornos del Conocimiento/metabolismo , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Doxiciclina/farmacología , Evaluación Preclínica de Medicamentos/métodos , Evaluación Preclínica de Medicamentos/psicología , Epotilonas/farmacología , Femenino , Hipocampo/efectos de los fármacos , Hipocampo/patología , Masculino , Aprendizaje por Laberinto/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Microtúbulos/efectos de los fármacos , Tauopatías/complicaciones , Tauopatías/genética , Tauopatías/patología , Tauopatías/psicología , Moduladores de Tubulina/farmacología , Proteínas tau/antagonistas & inhibidores , Proteínas tau/biosíntesis , Proteínas tau/genéticaRESUMEN
Ghrelin, through action on its receptor, GH secretagogue receptor type 1a (GHS-R1a), exerts a variety of metabolic functions including stimulation of appetite and weight gain and suppression of insulin secretion. In the present study, we examined the effects of novel small-molecule GHS-R1a antagonists on insulin secretion, glucose tolerance, and weight loss. Ghrelin dose-dependently suppressed insulin secretion from dispersed rat islets. This effect was fully blocked by a GHS-R1a antagonist. Consistent with this observation, a single oral dose of a GHS-R1a antagonist improved glucose homeostasis in an ip glucose tolerance test in rat. Improvement in glucose tolerance was attributed to increased insulin secretion. Daily oral administration of a GHS-R1a antagonist to diet-induced obese mice led to reduced food intake and weight loss (up to 15%) due to selective loss of fat mass. Pair-feeding experiments indicated that weight loss was largely a consequence of reduced food intake. The impact of a GHS-R1a antagonist on gastric emptying was also examined. Although the GHS-R1a antagonist modestly delayed gastric emptying at the highest dose tested (10 mg/kg), delayed gastric emptying does not appear to be a requirement for weight loss because lower doses produced weight loss without an effect on gastric emptying. Consistent with the hypothesis that ghrelin regulates feeding centrally, the anorexigenic effects of potent GHS-R1a antagonists in mice appeared to correspond with their brain exposure. These observations demonstrate that GHS-R1a antagonists have the potential to improve the diabetic condition by promoting glucose-dependent insulin secretion and promoting weight loss.
Asunto(s)
Depresores del Apetito/uso terapéutico , Apetito/efectos de los fármacos , Intolerancia a la Glucosa/tratamiento farmacológico , Receptores de Ghrelina/antagonistas & inhibidores , Pérdida de Peso/efectos de los fármacos , Animales , Depresores del Apetito/farmacología , Células Cultivadas , Evaluación Preclínica de Medicamentos , Ghrelina/antagonistas & inhibidores , Hipoglucemiantes/farmacología , Hipoglucemiantes/uso terapéutico , Masculino , Ratones , Ratones Endogámicos C57BL , Modelos Biológicos , Piperidinas/farmacología , Piperidinas/uso terapéutico , Quinazolinonas/farmacología , Quinazolinonas/uso terapéutico , Ratas , Ratas Sprague-Dawley , Ratas WistarRESUMEN
Selective activation of the NPY2 receptor to suppress appetite provides an approach to obesity management. Selective NPY2 PEGylated peptide agonists are described that consist of a peptide core corresponding to residues 25-36 of PYY and a nonpeptidic moiety at the peptide N-terminus that contributes to in vitro potency and in vivo efficacy and provides a PEGylation site. The lead peptide elicits a dose-dependent reduction of food intake in lean mice and of food intake, body weight, and fat mass in DIO mice.
Asunto(s)
Fármacos Antiobesidad/síntesis química , Peso Corporal/efectos de los fármacos , Ingestión de Alimentos/efectos de los fármacos , Oligopéptidos/síntesis química , Péptido YY/química , Polietilenglicoles/química , Receptores de Neuropéptido Y/agonistas , Animales , Fármacos Antiobesidad/química , Fármacos Antiobesidad/farmacología , AMP Cíclico/biosíntesis , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Oligopéptidos/química , Oligopéptidos/farmacología , Fragmentos de Péptidos/química , Fragmentos de Péptidos/farmacología , Péptido YY/farmacología , Ensayo de Unión Radioligante , Relación Estructura-ActividadRESUMEN
In Alzheimer's disease (AD), an extensive accumulation of extracellular amyloid plaques and intraneuronal tau tangles, along with neuronal loss, is evident in distinct brain regions. Staging of tau pathology by postmortem analysis of AD subjects suggests a sequence of initiation and subsequent spread of neurofibrillary tau tangles along defined brain anatomical pathways. Further, the severity of cognitive deficits correlates with the degree and extent of tau pathology. In this study, we demonstrate that phospho-tau (p-tau) antibodies, PHF6 and PHF13, can prevent the induction of tau pathology in primary neuron cultures. The impact of passive immunotherapy on the formation and spread of tau pathology, as well as functional deficits, was subsequently evaluated with these antibodies in two distinct transgenic mouse tauopathy models. The rTg4510 transgenic mouse is characterized by inducible over-expression of P301L mutant tau, and exhibits robust age-dependent brain tau pathology. Systemic treatment with PHF6 and PHF13 from 3 to 6 months of age led to a significant decline in brain and CSF p-tau levels. In a second model, injection of preformed tau fibrils (PFFs) comprised of recombinant tau protein encompassing the microtubule-repeat domains into the cortex and hippocampus of young P301S mutant tau over-expressing mice (PS19) led to robust tau pathology on the ipsilateral side with evidence of spread to distant sites, including the contralateral hippocampus and bilateral entorhinal cortex 4 weeks post-injection. Systemic treatment with PHF13 led to a significant decline in the spread of tau pathology in this model. The reduction in tau species after p-tau antibody treatment was associated with an improvement in novel-object recognition memory test in both models. These studies provide evidence supporting the use of tau immunotherapy as a potential treatment option for AD and other tauopathies.
Asunto(s)
Enfermedad de Alzheimer/terapia , Anticuerpos Monoclonales/farmacología , Trastornos del Conocimiento/terapia , Inmunización Pasiva , Fosfoproteínas/farmacología , Proteínas tau/antagonistas & inhibidores , Enfermedad de Alzheimer/inducido químicamente , Enfermedad de Alzheimer/inmunología , Enfermedad de Alzheimer/patología , Animales , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/inmunología , Corteza Cerebral/patología , Trastornos del Conocimiento/inducido químicamente , Trastornos del Conocimiento/inmunología , Trastornos del Conocimiento/patología , Modelos Animales de Enfermedad , Conducta Exploratoria/efectos de los fármacos , Regulación de la Expresión Génica , Hipocampo/efectos de los fármacos , Hipocampo/inmunología , Hipocampo/patología , Masculino , Ratones , Ratones Transgénicos , Neuronas/efectos de los fármacos , Neuronas/inmunología , Neuronas/patología , Cultivo Primario de Células , Proteínas Recombinantes/administración & dosificación , Proteínas Recombinantes/efectos adversos , Transducción de Señal , Resultado del Tratamiento , Proteínas tau/genética , Proteínas tau/inmunologíaRESUMEN
Filamentous inclusions of the microtubule-associated protein, tau, define a variety of neurodegenerative diseases known as tauopathies, including Alzheimer's disease (AD). To better understand the role of tau-mediated effects on pathophysiology and global central nervous system function, we extensively characterized gene expression, pathology and behavior of the rTg4510 mouse model, which overexpresses a mutant form of human tau that causes Frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17). We found that the most predominantly altered gene expression pathways in rTg4510 mice were in inflammatory processes. These results closely matched the causal immune function and microglial gene-regulatory network recently identified in AD. We identified additional gene expression changes by laser microdissecting specific regions of the hippocampus, which highlighted alterations in neuronal network activity. Expression of inflammatory genes and markers of neuronal activity changed as a function of age in rTg4510 mice and coincided with behavioral deficits. Inflammatory changes were tau-dependent, as they were reversed by suppression of the tau transgene. Our results suggest that the alterations in microglial phenotypes that appear to contribute to the pathogenesis of Alzheimer's disease may be driven by tau dysfunction, in addition to the direct effects of beta-amyloid.
Asunto(s)
Enfermedad de Alzheimer/genética , Expresión Génica/genética , Redes Reguladoras de Genes/genética , Inflamación/genética , Proteínas tau/genética , Animales , Cromosomas Humanos Par 17/genética , Modelos Animales de Enfermedad , Femenino , Demencia Frontotemporal/genética , Hipocampo/metabolismo , Humanos , Ratones , Microglía/metabolismo , Proteínas Asociadas a Microtúbulos/genética , Enfermedades Neurodegenerativas/genética , Neuronas/metabolismo , Trastornos Parkinsonianos/genéticaRESUMEN
Levels of tau in cerebrospinal fluid (CSF) are elevated in Alzheimer's disease (AD) patients. It is believed this elevation is related to the tau pathology and neurodegeneration observed in AD, but not all tauopathies have increased CSF tau. There has been little pre-clinical work to investigate mechanisms of increased CSF tau due to the difficulty in collecting CSF samples from mice, the most commonly used pre-clinical models. We developed methods to collect CSF from mice without contamination from tau in brain tissue, which is approximately 50,000 fold more abundant in brain than CSF. Using these methods, we measured CSF tau from 3xTg, Tg4510, and Tau Alone transgenic mice. All three lines of mice showed age-dependent increases in CSF tau. They varied in phenotype from undetectable to severe tau pathology and neurodegeneration, suggesting that degenerating neurons are unlikely to be the only source of pathologic CSF tau. Overall, CSF tau levels mirrored expression levels and changes of tau in the brain, but they did not always correlate exactly. CSF tau was often more sensitive to changes in brain transgene expression and pathology. In addition, we also developed ELISA assays specific to different regions of the tau protein. We used these assays to provide evidence that CSF tau exists as fragments, with little intact C-terminus and partial loss of the N-terminus. Taken together, these assays and mouse models may be used to facilitate a deeper understanding of CSF tau in neurodegenerative disease.
Asunto(s)
Enfermedad de Alzheimer/líquido cefalorraquídeo , Proteínas tau/líquido cefalorraquídeo , Factores de Edad , Enfermedad de Alzheimer/patología , Precursor de Proteína beta-Amiloide/genética , Análisis de Varianza , Animales , Biomarcadores/líquido cefalorraquídeo , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Modelos Animales de Enfermedad , Doxiciclina/farmacología , Ensayo de Inmunoadsorción Enzimática , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/genética , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas de Neurofilamentos/metabolismo , Presenilina-1/genética , ARN Mensajero/metabolismo , Tubulina (Proteína)/metabolismo , Proteínas tau/genéticaRESUMEN
Activation of the NPY2 receptor to reduce appetite while avoiding activation of the NPY1 and NPY5 receptors that stimulate feeding provides a pharmaceutical approach to modulate food intake. The naturally occurring peptide and development candidate PYY(3-36) is a non-selective NPY1, NPY2, and NPY5 agonist of limited in vivo duration of action. N-terminal modification with 20 kDa PEG of a selective NPY2 receptor agonist peptide results in a long-acting agent that outperforms PYY(3-36) in reducing food intake in mice. The results suggest that PEGylated, selective NPY2 peptide agonists offer a significantly improved therapeutic benefit over PYY(3-36) for obesity management.
Asunto(s)
Química Farmacéutica/métodos , Ingestión de Alimentos/efectos de los fármacos , Conducta Alimentaria/efectos de los fármacos , Péptido YY/síntesis química , Péptido YY/farmacología , Polietilenglicoles/química , Receptores de Neuropéptido Y/química , Animales , Relación Dosis-Respuesta a Droga , Diseño de Fármacos , Humanos , Cinética , Ratones , Conformación Molecular , Fragmentos de Péptidos , Péptidos/química , Unión ProteicaRESUMEN
Selective activation of the neuropeptide Y (NPY)2 receptor to suppress appetite provides a promising approach to obesity management. A selective NPY2 polyethylene glycol-conjugated (PEGylated) peptide agonist is described that consists of a peptide core corresponding to residues 13 to 36 of human peptide YY (PYY) and a nonpeptidic moiety (2-mercaptonicotinic acid) at the peptide N terminus that is derivatized with 20-kDa monomethoxypolyethylene glycol. The PEGylated peptide elicits a dose-dependent reduction in food intake in lean C57BL/6 mice and Wistar rats that persists for 72 and 48 h, respectively. The effect on food intake in lean C57BL/6 mice is blocked by the selective NPY2 antagonist BIIE0246 (N-[(1S)-4-[(aminoiminomethyl)amino]-1-[[[2-(3,5-dioxo-1,2-diphenyl-1,2,4-triazolidin-4-yl)ethyl]amino]carbonyl]butyl]-1-[2-[4-(6,11-dihydro-6-oxo-5H-dibenz[b,e]azepin-11-yl)-1-piperazinyl]-2-oxoethyl]-cyclopentaneacetamide formate). A dose-dependent reduction in body weight in diet-induced obese (DIO) mice is seen following daily dosing for 14 days. The reduction in body weight is sustained following dosing for 40 days, and it is accompanied by an increase in plasma adiponectin. Improvements in glucose disposal and in plasma insulin and glucose levels that are risk factors for type II diabetes are observed following once-daily subcutaneous dosing in DIO mice. The results provide evidence from two animal species that the long-acting selective NPY2 peptide agonist has potential for obesity management.
Asunto(s)
Depresores del Apetito/farmacología , Peso Corporal/efectos de los fármacos , Ingestión de Alimentos/efectos de los fármacos , Glucosa/metabolismo , Fragmentos de Péptidos/farmacología , Péptido YY/farmacología , Polietilenglicoles/farmacología , Receptores de Neuropéptido Y/agonistas , Adiponectina/sangre , Animales , Arginina/análogos & derivados , Arginina/farmacología , Benzazepinas/farmacología , Relación Dosis-Respuesta a Droga , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratas , Ratas Wistar , Relación Estructura-ActividadRESUMEN
Activation of the NPY2 receptor to reduce appetite while avoiding stimulation of the NPY1 and NPY5 receptors that induce feeding provides a pharmaceutical approach to modulate food intake. The naturally occurring peptide PYY(3-36) is a nonselective NPY1, NPY2, and NPY5 agonist. N-terminal truncation of PYY to abrogate affinity for the NPY1 and NPY5 receptors and subsequent N-terminal modification with aminobenzoic analogs to restore NPY2 receptor potency results in a series of highly selective NPY2 receptor peptide agonists.
Asunto(s)
Ingestión de Alimentos/efectos de los fármacos , Péptido YY/química , Péptido YY/farmacología , Receptores de Neuropéptido Y/agonistas , Unión Competitiva/efectos de los fármacos , Línea Celular , Cromatografía Líquida de Alta Presión , Guanosina 5'-O-(3-Tiotrifosfato)/farmacología , Humanos , Indicadores y Reactivos , Fragmentos de Péptidos , Receptores de Neuropéptido Y/efectos de los fármacosRESUMEN
Modulation of cAMP levels has been linked to insulin secretion in preclinical animal models and in humans. The high expression of PDE-10A in pancreatic islets suggested that inhibition of this enzyme may provide the necessary modulation to elicit increased insulin secretion. Using an HTS approach, we have identified quinoline-based PDE-10A inhibitors as insulin secretagogues in vitro. Optimized compounds were evaluated in vivo where improvements in glucose tolerance and increases in insulin secretion were measured.