RESUMEN
Interruptin B has been isolated from Cyclosorus terminans, however, its pharamcological effect has not been fully identified. In the present study, the effects of interruptin B, from C. terminans, on brown adipocyte differentiation and glucose uptake in adiposederived stem cells (ASCs) were investigated. The results revealed that interruptin B dosedependently enhanced the adipogenic differentiation of ASCs, with an induction in the mRNA expression levels of peroxisome proliferatoractivated receptor (PPAR)α and PPARγ. In addition, interruptin B efficiently increased the number and the membrane potential of mitochondria and upregulated the mRNA expression levels of uncoupling protein (UCP)1 and cyclooxygenase (COX)2, which are all predominantly expressed in brown adipocytes. Interruptin B increased glucose consumption in differentiated ASCs, accompanied by the upregulation in the mRNA expression levels of glucose transporter (GLUT)1 and GLUT4. The computational analysis of molecular docking, a luciferase reporter assay and surface plasmon resonance confirmed the marked binding affinity of interruptin B to PPARα and PPARγ (K(D) values of 5.32 and 0.10 µm, respectively). To the best of our knowledge, the present study is the first report to show the stimulatory effects of interruptin B on brown adipocyte differentiation and glucose uptake in ASCs, through its role as a dual PPARα and PPARγ ligand. Therefore, interruptin B could be further developed as a therapeutic agent for the treatment of diabetes.
Asunto(s)
Adipocitos Marrones/citología , Tejido Adiposo/citología , Diferenciación Celular/efectos de los fármacos , Chalconas/farmacología , Cumarinas/farmacología , Glucosa/metabolismo , Células Madre/citología , Células Madre/metabolismo , Chalconas/química , Simulación por Computador , Cumarinas/química , Células Hep G2 , Humanos , Ligandos , Simulación del Acoplamiento Molecular , PPAR alfa/antagonistas & inhibidores , PPAR alfa/metabolismo , PPAR gamma/antagonistas & inhibidores , PPAR gamma/metabolismo , Células Madre/efectos de los fármacos , Resonancia por Plasmón de SuperficieRESUMEN
It has been shown that some chalcones are able to inhibit tubulin polymerization, giving cytotoxicity and destruction of tumoral vasculature. A library of 180 novel chalcone analogs has been synthesized via click chemistry and screened for their cytotoxicity and tubulin assembly inhibition. 10 out 180 click chalcones displayed low micromolar cytotoxicity but only compound Nf depicted antitubulin activity. While Nf displayed only micromolar potency this result shows click-chalcones may be anti-tubulin agents and validate this strategy to search for novel active chemical entities.
Asunto(s)
Chalconas/síntesis química , Moduladores de Tubulina/síntesis química , Tubulina (Proteína)/metabolismo , Línea Celular Tumoral , Chalconas/química , Chalconas/farmacología , Química Clic , Humanos , Tubulina (Proteína)/química , Moduladores de Tubulina/química , Moduladores de Tubulina/farmacologíaRESUMEN
AIMS: The aims of the present study were to investigate the effect of crebanine on memory and cognition impairment in mice and to elucidate the underlying molecular mechanisms. MAIN METHODS: The memory-enhancing effects of crebanine were assessed with a water maze test using scopolamine-induced amnesic mice. The molecular mechanism was explored in silico by docking crebanine against acetylcholine binding proteins (AChBPs) and in vitro with a radioligand competition assay using (±)-[(3)H]-epibatidine. The pharmacological behavior was assessed by observing changes to the functional activity of α7-nAChRs expressed in Xenopus oocytes and by fluorescent assays on recombinant ligand gated ion channel (LGIC) receptors expressed in mammalian cells. KEY FINDINGS: The administration of crebanine significantly improved the cognitive deficits induced by scopolamine, as measured by the water maze test. The docking results demonstrated that crebanine bound to the active binding site of the AChBP template with a good docking energy. Crebanine significantly inhibited the binding of (±)-[(3)H]-epibatidine to AChBPs with K(i) values of 179 nM and 538 nM for Ls and Ac, respectively. Further functional assays performed using two separate protocols indicated that crebanine is an antagonist of the α7-nAChR with an IC(50) of 19.1µM. SIGNIFICANCE: The observed actions of crebanine against amnesia and its effect on α7-nAChRs will be beneficial for target-based drug design; crebanine or its scaffold can be used as the starting point to develop a drug for Alzheimer's disease. The cognition-enhancing effects of crebanine and the underlying mechanism based on α7-nAChRs are consistent with its traditional use. These findings demonstrate the potential utility of crebanine in the development of neurodegenerative therapy.
Asunto(s)
Aporfinas/farmacología , Trastornos del Conocimiento/tratamiento farmacológico , Memoria/efectos de los fármacos , Receptores Nicotínicos/metabolismo , Animales , Unión Competitiva , Línea Celular , Transferencia Resonante de Energía de Fluorescencia , Colorantes Fluorescentes/farmacología , Humanos , Ligandos , Ratones , Ratones Endogámicos ICR , Conformación Molecular , Oocitos/citología , Ensayo de Unión Radioligante/métodos , Proteínas Recombinantes/metabolismo , Xenopus laevis , Receptor Nicotínico de Acetilcolina alfa 7RESUMEN
Acetylcholinesterase (AChE) inhibitory activity is one of the proposed targets for indole analogs. Simple indoles with substitution of methoxy, carboxy or hydroxy at the benzene ring showed a low percent of inhibitory activity in eel-AChE. Adding a side chain at the pyrrole ring, such as serotonin, ß-carbolines and quinolines (the bioisostere of indole), improved the inhibitory activity significantly. However, proper substitution and conformation of the ring were required for good binding. The result of inhibition in human-AChE of serotonin, ß-carbolines and quinolines showed similar profile as eel-AChE with lower magnitude. The data from molecular docking showed that they shared the same binding site as galantamine.
Asunto(s)
Acetilcolinesterasa , Inhibidores de la Colinesterasa/síntesis química , Inhibidores de la Colinesterasa/farmacología , Indoles/síntesis química , Acetilcolinesterasa/metabolismo , Sitios de Unión , Carbolinas/química , Carbolinas/farmacología , Inhibidores de la Colinesterasa/química , Activación Enzimática/efectos de los fármacos , Humanos , Indoles/química , Indoles/farmacología , Conformación Molecular , Quinolinas/química , Quinolinas/farmacologíaRESUMEN
The nicotinic acetylcholine receptors (nAChRs) are a member of the ligand-gated ion channel family and play a key role in the transfer of information across neurological networks. The X-ray crystal structure of agonist-bound α(7) acetylcholine binding protein (AChBP) has been recognized as the most appropriate template to model the ligand-binding domain of nAChR for studying the molecular mechanism of the receptor-ligand interactions. Virtual screening of the National Cancer Institute diversity set, a library of 1990 compounds with nonredundant pharmacophore profiles, using AutoDock against AChBPs revealed 51 potential candidates. In vitro radioligand competition assays using [(3)H] epibatidine against the AChBPs from the freshwater snails, Lymnaea stagnalis, and from the marine species, Aplysia californica and the mutant (AcY55W), revealed seven compounds from the list of candidates that had micromolar to nanomolar affinities for the AChBPs. Further investigation on α(7)nAChR expressing in Xenopus oocytes and on the recombinant receptors with fluorescence resonance energy transfer (FRET)-based calcium sensor expressing in HEK cells showed that seven compounds were antagonists of α(7)nAChR, only one compound (NSC34352) demonstrated partial agonistic effect at low dose (10 µM), and two compounds (NSC36369 and NSC34352) were selective antagonists on α(7)nAchR with moderate potency. These hits serve as novel templates/scaffolds for development of more potent and specific in the AChR systems.
Asunto(s)
Proteínas Portadoras/agonistas , Proteínas Portadoras/antagonistas & inhibidores , Evaluación Preclínica de Medicamentos/métodos , Receptores Nicotínicos/metabolismo , Algoritmos , Animales , Aplysia , Sitios de Unión , Cristalografía por Rayos X , Transferencia Resonante de Energía de Fluorescencia , Células HEK293 , Humanos , Canales Iónicos Activados por Ligandos , Lymnaea , Oocitos , Conformación Proteica , Receptores Nicotínicos/química , Programas Informáticos , XenopusRESUMEN
Rediocides A and G, the principle components of Trigonostemon reidioides (Kurz) Craib, which is known as Lotthanong in Thai, were investigated for a detoxification mechanism against Naja kaouthia venom by in silico, in vitro, and in vivo methods. Molecular dockings of alpha-cobratoxin with rediocides A and G were performed, and the binding energies were found to be -14.17 and -14.14 kcal/mol, respectively. Rediocides bind to alpha-cobratoxin at the same location as alpha-cobratoxin binds to the nicotinic acetylcholine receptor (nAChR), i.e., at the Asp27, Phe29, Arg33, Gly34, Lys35, and Val37 residues. alpha-Cobratoxin cannot bind to nAChR, because some of its binding sites are occupied with rediocides. From in vitro SDS-PAGE, it was found that rediocides can diminish the bands of alpha-cobratoxin. In the presence of acetylcholine-binding protein (AChBP), it was apparent that rediocides can bind both alpha-cobratoxin and AChBP. From an in vivo test, it was found that injection of rediocides at 0.5 mg/kg immediately after an alpha-cobratoxin dose of three times LD(50) cannot prolong the survival time of mice. However, rediocide can prolong the survival time, if it is injected 30 min before the injection of alpha-cobratoxin. The in vitro SDS-PAGE and the in vivo results support the in silico detoxification mechanism of rediocides against cobra venom at a molecular level.
Asunto(s)
Antitoxinas/química , Proteínas Neurotóxicas de Elápidos/metabolismo , Diterpenos/química , Macrólidos/química , Animales , Antitoxinas/farmacología , Sitios de Unión , Proteínas Neurotóxicas de Elápidos/química , Simulación por Computador , Diterpenos/farmacología , Enlace de Hidrógeno , Macrólidos/farmacología , Ratones , Unión Proteica , Receptores Nicotínicos/química , Receptores Nicotínicos/metabolismoRESUMEN
alpha-Cobratoxin (Cbtx), the neurotoxin isolated from the venom of the Thai cobra Naja kaouthia , causes paralysis by preventing acetylcholine (ACh) binding to nicotinic acetylcholine receptors (nAChRs). In the current study, the region of the Cbtx molecule that is directly involved in binding to nAChRs is used as the target for anticobratoxin drug design. The crystal structure (1YI5) of Cbtx in complex with the acetylcholine binding protein (AChBP), a soluble homolog of the extracellular binding domain of nAChRs, was selected to prepare an alpha-cobratoxin active binding site for docking. The amino acid residues (Ser182-Tyr192) of the AChBP structure, the binding site of Cbtx, were used as the positive control to validate the prepared Cbtx active binding site (root mean square deviation < 1.2 A). Virtual screening of the National Cancer Institute diversity set, a library of 1990 compounds with nonredundant pharmacophore profiles, using AutoDock against the Cbtx active site, revealed 39 potential inhibitor candidates. The adapted in vitro radioligand competition assays using [(3)H]epibatidine and [(125)I]bungarotoxin against the AChBPs from the marine species, Aplysia californica (Ac), and from the freshwater snails, Lymnaea stagnalis (Ls) and Bolinus truncates (Bt), revealed 4 compounds from the list of inhibitor candidates that had micromolar to nanomolar interferences for the toxin binding to AChBPs. Three hits (NSC42258, NSC121865, and NSC134754) can prolong the survival time of the mice if administered 30 min before injection with Cbtx, but only NSC121865 and NSC134754 can prolong the survival time if injected immediately after injection with Cbtx. These inhibitors serve as novel templates/scaffolds for the development of more potent and specific anticobratoxin.
Asunto(s)
Antivenenos/química , Proteínas Neurotóxicas de Elápidos/química , Proteínas Neurotóxicas de Elápidos/metabolismo , Simulación por Computador , Evaluación Preclínica de Medicamentos , Acetilcolina/metabolismo , Animales , Sitios de Unión , Proteínas Neurotóxicas de Elápidos/genética , Proteínas Neurotóxicas de Elápidos/toxicidad , Diseño de Fármacos , Elapidae , Humanos , Masculino , Ratones , Modelos Moleculares , Datos de Secuencia Molecular , Estructura Molecular , Unión Proteica , Conformación Proteica , Receptores Nicotínicos/metabolismoRESUMEN
NP506, the 3-{2,4-dimethyl-5-[2-oxo-5-(N'-phenylhydrazinocarbonyl)-1,2-dihydro-indol-3-ylidenemethyl]-1H-pyrrol-3-yl}-propionic acid, was designed as FGF receptor 1 inhibitor by computational study and found to be more active against endothelial proliferation of HUVEC after the rhFGF-2 stimulation than SU6668 with minimum effective dose of 10 microM. NP506 inhibited the tyrosine phosphorylation in FGF, VEGF, and PDGF receptors and the activation of extracellular signal-regulated kinase (ERK), c-Jun-N-terminal-kinase (JNK) and AKT after the rhFGF-2 stimulation. The introduction of the phenyl hydrazide motif to the position 5 of the pyrido[2,3-d]pyrimidine scaffold led to the inhibitory effect in two signaling pathways: inhibition of AKT activation in the phosphatidyl inositol 3'-kinase (PI13K)/AKT signaling pathway and the inhibition of ERK and JNK activation in MAPK pathway.