.cp_wz tabla {borde superior: 1px sólido #ccc; borde izquierdo: 1px sólido #ccc; } .cp_wz table td {borde derecho: 1px sólido #ccc; borde inferior: 1px sólido #ccc; padding: 5px 0px 0px 5px;} .cp_wz table th {border-right: 1px solid #ccc; border-bottom: 1px solid #ccc; padding: 5px 0px 0px 5px;} \ n Peso molecular: 308.31 AR-A014418 es un inhibidor de GSK3β selectivo y competitivo con ATP con IC50 y Ki de 104 nM y 38 nM, sin inhibición significativa en otras 26 quinasas probadas. \ n AR -A014418 inhibe la fosforilación de tau en un sitio específico de GSK3 (Ser-396) en fibroblastos 3T3 que expresan la proteína tau humana de cuatro repeticiones con IC50 de 2,7 μM, y protege las células N2A cultivadas de la muerte inducida por el bloqueo de la vía PI3K / PKB. En cortes de hipocampo, AR-A014418 inhibe la neurodegeneración mediada por el péptido beta-amiloide. Mientras que en las células NGP y SH-5Y-SY, AR-A014418 reduce los marcadores neuroendocrinos y suprime el crecimiento celular del neuroblastoma. En el modelo de ratón ALS con la SOD1 humana mutante G93A, AR-A014418 (0-4 mg / kg, ip) retrasa el inicio de síntomas, mejora la actividad motora, ralentiza la progresión de la enfermedad y pospone el punto final de la enfermedad. Además, AR-A014418 produce un efecto de inhibición sobre la nocicepción inducida por ácido acético y formalina en ratones al modular la señalización del receptor metabotrópico y NMDA, así como la transmisión de TNF-α e IL-1β en la médula espinal. \ n Inhibidor selectivo de GSK3 permeable a las células. \ n
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GSK3 Scintillation Proximity Assay
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The competition experiments are carried out in duplicate with 10 concentrations of the inhibitor in clear-bottomed microtiter plates. The biotinylated peptide substrate, biotin-AAEELDSRAGS(PO3H2)PQL, is added at a final concentration of 2 μM in an assay buffer containing 6 milliunits of recombinant human GSK3 (equal mix of both α and β), 12 mM MOPS, pH 7.0, 0.3 mM EDTA, 0.01% β-mercaptoethanol, 0.004% Brij 35, 0.5% glycerol, and 0.5 μg of bovine serum albumin/25 μl and preincubated for 10-15 min. The reaction is initiated by the addition of 0.04 μCi of [γ-33P]ATP and unlabeled ATP in 50 mM Mg(Ac)2 to a final concentration of 1 μM ATP and assay volume of 25 μl. Blank controls without peptide substrate are used. After incubation for 20 min at room temperature, each reaction is terminated by the addition of 25 μl of stop solution containing 5 mM EDTA, 50 μM ATP, 0.1% Triton X-100, and 0.25 mg of streptavidin-coated SPA beads corresponding to ∼35 pmol of binding capacity. After 6 h the radioactivity is determined in a liquid scintillation counter. Inhibition curves are analyzed by non-linear regression using GraphPad Prism.
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Ensayo celular: [1]
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Cell lines
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N2A cells
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Concentrations
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~50 μM
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Incubation Time
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24 h
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Method
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Cell viability is assessed by calcein/propidium iodide uptake. Calcein AM is taken up and cleaved by esterases present within living cells, yielding yellowish-green fluorescence, whereas PI is only taken up by dead cells, which become orange-red fluorescent. In brief, N2A cells are cultured for 2 days in vitro and then treated with 50 μM LY-294002 in the presence of AR-A014418 or vehicle (DMSO) for 24 h. Subsequently, N2A cells are incubated for 30 min with 2 μM PI and 1 μM calcein-AM. The cultures are then rinsed three times with Hanks' buffered saline solution containing 2 mM CaCl2, and the cells are visualized by fluorescence microscopy using a Zeiss Axiovert 135 microscope. Three fields (selected at random) are analyzed per well (∼300 cells/field) in at least three different experiments. Cell death is expressed as percentage of PI-positive cells from the total number of cells. In every experiment, specific cell death is obtained after subtracting the number of dead cells present in vehicle-treated cultures.
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Estudio con animales: [2]
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Animal Models
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ALS mouse model with the G93A mutant human SOD1
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Formulation
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Normal saline
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Dosages
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~4 mg/kg
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Administration
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i.p.
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Conversión de diferentes modelos de animales basados en BSA (valor basado en datos del Borrador de Directrices de la FDA)
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Species
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Baboon
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Dog
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Monkey
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Rabbit
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Guinea pig
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Rat
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Hamster
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Mouse
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Weight (kg)
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12
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10
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3
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1.8
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0.4
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0.15
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0.08
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0.02
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Body Surface Area (m2)
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0.6
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0.5
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0.24
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0.15
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0.05
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0.025
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0.02
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0.007
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Km factor
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20
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20
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12
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12
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8
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6
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5
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3
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Animal A (mg/kg) = Animal B (mg/kg) multiplied by
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Animal B Km
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Animal A Km
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Por ejemplo, para modificar la dosis de resveratrol utilizada para un ratón (22,4 mg / kg) a una dosis basada en el BSA para una rata, multiplique 22,4 mg / kg por el factor Km para un ratón y luego divida por el factor Km para una rata. Este cálculo da como resultado una dosis equivalente para ratas de resveratrol de 11,2 mg / kg.
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Rat dose (mg/kg) = mouse dose (22.4 mg/kg) ×
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mouse Km(3)
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= 11.2 mg/kg
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rat Km(6)
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Información química
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Molecular Weight (MW)
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308.31
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Formula
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C12H12N4O4S
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CAS No.
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487021-52-3
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Storage
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3 years -20℃Powder
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6 months-80℃in solvent (DMSO, water, etc.)
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Synonyms
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GSK-3β Inhibitor VIII
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Solubility (25°C) *
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In vitro
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DMSO
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62 mg/mL
(201.09 mM)
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Water
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<1 mg/mL
(
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Ethanol
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<1 mg/mL
(
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* <1 mg/ml means slightly soluble or insoluble.
* Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.
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Chemical Name
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N-[(4-Methoxyphenyl)methyl]-N'-(5-nitro-2-thiazolyl)urea
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Calculadora de molaridad Calculadora de dilución Calculadora de peso molecular
Grupos de Producto : PI3K / Akt / mTOR > Inhibidor GSK-3
