CustomerValidation
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| Description | SimeprevirisapotentHCVNS3/4Aproteaseinhibitor,andinhibitsHCVreplicationwithEC50of8nM. |
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| IC50&Target | EC50:8nM |
| InVitro | InHuh7-Luccells,antiviralactivityofsimeprevir(TMC435350)isdosedependent,andtheEC50andEC90valuesdeterminedforTMC435350are8nMand24nM,respectively.InhibitionofTMC435350onNS3/4Aproteaseistimedependent,andtheoverall Kisareestimatedtobe0.5nMforgenotype1aand0.4nMforgenotype1b,respectively[1].TMC435350isapotentinhibitorofHCVNS3/4Aprotease(Ki=0.36nM)andviralreplication(repliconEC50=7.8nM)[2]. |
| InVivo | Inrats,TMC435350(40mg/kg,p.o.)isextensivelydistributedtotheliverandintestinaltract(tissue/plasmaareaundertheconcentration-timecurveratiosof>35),andtheabsolutebioavailABIlityis44%[1]. |
| ClinicalTrial | ViewMoreCollapse |
| References |
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| PreparingStockSolutions |
Pleaserefertothesolubilityinformationtoselecttheappropriatesolvent. | ||||||||||||||||||||||||||
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| KinaseAssay [1] | InvitroinhibitionofNS3/4AactivityisdeterminedusingafluorescenceresonanceenergytransfercleavageassaywiththeRetS1peptidesubstrate,derivedfromthegenotype1aNS4A-4Bjunction,andbacteriallyexpressedfull-lengthNS3proteasedomain,supplementedwithanNS4Apeptide.Briefly,NS3/4AispreincubatedinthepresenceofTMC435350for10min,andthentheRetS1substrateisaddedandfluorescenceiscontinuouslymeasuredfor20min(excitation,355nm;emission,500nm).Cleavageofthesubstrateisexpressedasapercentageofthecleavageseenwiththevehiclecontrol.MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly. | ||||||||||||||||||||||||||
| CellAssay [1] | SimeprevirisdilutedininafinalDMSOconcentrationof0.5%intheabsenceofG418. Huh7-Luccellsareseededatadensityof2,500cells/wellina384-wellplateinDulbecco"smodifiedEagle"smediumplus10%fetalcalfserumandincubatedwitharangeofconcentrationsofseriallydilutedsimeprevir(TMC435350),inafinalDMSOconcentrationof0.5%intheabsenceofG418.After72hofincubation,SteadyLitereagentisaddedina1:1ratiotothemedium,andluciferasesignalismeasuredusingaViewLuxreader. MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly. | ||||||||||||||||||||||||||
| AnimalAdmiNISTration [1] | Twenty-fourmalespecific-pathogen-freeSprague-Dawleyrats,weighingbetween200and300gatthetimeofdosing,aredividedintoeightgroupsofthreeratseach.Sevengroupsaredosedorally(p.o.)bygastricintubationofavitaminEacetate-d-α-tocopherylpolyethyleneglycol1000succinate-polyethyleneglycol400solutionofSimeprevir(TMC435350)at2mL/kgbodyweighttoprovideadoseof40mg/kg.Onegroupisdosedintravenously(i.v.)byslowbolusinjectioninatailveinofa20%2-hydroxypropyl-β-cyclodextrinformulationofTMC435350(containingTMC435350,100mg/mL2-hydroxypropyl-β-cyclodextrin,0.1NNaOHtopH8.0±0.1,andmannitol-andpyrogen-freewater)at2mL/kgbodyweighttoprovideadoseof4mg/kg.Waterandfoodareavailableadlibitumduringthestudy.MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly. References |
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