口服药物的体外吸收模型.ppt

口服药物的体外吸收模型.ppt

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InVitroModelsforOralDrugAbsorptionOutlinenPhysicalandBiochemicalBarrierforOralDrugAbsorptionandInVitromodelsfordrugabsorptionnEvaluationoftheMethodCaco-2CellSuitabilitynUseofCaco2andFactorAffectingPermeationofNCEnContinuousDissolution/Caco-2SystemforFormulationScreeningofNCEBLightmicrographofmonkeysmallintestinalmucosaillustratingthethreelayersofthemucosa:

theepithelium（E）,thelaminapropria（L）,andthemuscularismucosa（M）.（takenfromJ.L.MaderaandJ.S.TrierinPhysiologyoftheGastrointestinalTract（L.R.Johnson,Ed.）,2ndEdition,RavenPress,NewYork,1987,p.1210）LEMIntestinalMucosa:

APhysicalandBiochemicalBarrierforOralDrugAbsorptionViewofIntestinalMucosa,Pre-1990ACurrentViewofIntestinalMucosa?

BasolateralApicalParacellularDiffusionTranscellularDiffusionMetabolismABEffluxTransportACDMultidrugResistanceTransportersandCYP3AsFormaConcertedBarriertoDrugAbsorptionMultidrugResistanceTransporters（MDR1,MRP2,BCRP）CYP3AsApicalsideBasolateralsideMetabolizedSubstratedrugDrugmetaboliteMDR1（P-glycoprotein,P-gp）Inhibitorse.g.GF120918ABSubstratese.g.DigoxinExamplesofDrugsThatareSubstratesforMDR1CancerdrugsVinblastineVincristineTaxolDaunorubicinEtoposideImmunosuppressiveDrugsCyclosporinAFK506SteroidsDexamethasoneProgesteroneCortisolHIVproteaseinhibitorsAmprenavirIndinavirCardiacdrugsDigoxinQuinidineAnti-goutagentColchicineAntibioticErythromycinAnti-tuberculousagentRifampinSchematicIllustrationofMDR1,MRP1,MRP2,MRP3,MRP5andBCRPinTransfectedPolarizedEpithelialCellsReference:

P.Borstetal.,BBA1461,347-357（1999）J.Jonkeretal.,JNCI92（20）,1651-1656（2000）MDR1MRP1MRP2MRP3MRP5ApicalsideBasolateralsideTightjunctionBCRPModelsUsedtoDeterminetheIntestinalMucosalPermeationCharacteristicsofaDrugCandidateOraldosingofanimalsorhumansIntestinalperfusionstudiesinhumansDosingofanimalsthroughachroniccannulaintheintestinallumenInSituintestinalperfusionstudiesinanimalsEffluxtransporter-geneknockoutmiceInVitropermeationstudiesusingexcisedhumanoranimalintestinalmucosaltissuesInVitropermeationstudiesusingcellculturemodelsPAMPA,IAMGastroPlusIntestinalPerfusionStudiesinHumansCorrelationbetweenFractionAbsorbedofd-GlucoseandPhenazoneinHumanRegionalPerfusionStudyDrugAbsorptionStudyinsituandinConsciousRatExtractionforLC/MS/MSAnalysisHPLCAnalysis0.2ml/minCentrifugationBloodFiltrationPerfusateInSituRatIntestinalPerfusionMesentericveinIleumAnimals:

MaleSprague-Dawleyratsweighing350-400gwereused.Perfusionsolution:

NaH2PO4（57.9mM）,Na2SO4（79.6mM）,pH7.5.PumpDonorBloodJugularveinPapp=DQDtAC0Basolateral（B）Apical（A）CellMonolayerBi-directionalTransportExperimentUsingCaco-2CellMonolayersandCalculationSubstratedrug（e.g.Digoxin）Apicallylocalizedeffluxtransporter（e.g.MDR1）Q/t:

Thelinearappearancerateofmassinthereceiversolution.A:

Cellmonolayersurfacearea.C0:

Theinitialconcentrationinthedonorsolution.PBtoAPAtoBRatioofPBtoA/PAtoB:

AnindicatorofeffluxactivityoftransporterincellmonolayerCorrelationbetweenOralAbsorptioninHumansandMembranePermeability（Papp）inCaco-2CellsP.Arturssonetal.,Adv.DrugDel.Rev.46:

27-43（2001）HistoryofCellCultureModelsUsedtoPredictDrugPermeationacrosstheIntestinalMucosaLate1980s-Mid1990s:

Caco-2cellsMid1990s:

Caco-2cellsvs.MDCKcellsLate1990s:

Caco-2cellsvs.MDCKcellsvs.MDCK-MDR1vs.MDCK-MRP2Caco-2CellMonolayerSystemAdvantagesCellspermitestimationof:

bothcellularuptakeanddrugtransepithelialtransportdrugtransportinthepresenceofmetabolicreactionspolarizedeffluxCellsare:

ofhumanorigin（colonadenocarcinama）viableforlongperiodsDisadvantagesRelativelong-periodoftimeforculture（3weeks）Underexpressionofkeydrug-metabolizingenzymes（e.g.,CYP3A4）Variableexpressionlevelsofeffluxtransporters（e.g.,MDR1,MRP2）PappvaluesinMDCKcellscorrelatedwellwiththoseinCaco-2cellsThecorrelationbetweenMDCKPappvaluesandhumanintestinalabsorptionissimilartotheresultsobservedwithCaco-2MDCKgrowfasterthanCaco-2（1weekvs3weeks）J.D.Irvineetal.,JPS88,28（1999）MDCKCellMonolayerSystemQuestionsConcerningMDCK-MDR1/MDCK-MRP2CellsastheModeloftheIntestinalMucosaMDR1/MRP2cDNAMDCKcellsWhencomparedwithCaco-2cells,dothesecelllines:

expressthesameprotein?

exhibitpolarizedefflux?

exhibitthesamekineticsandaffinityforsubstrates?

MDCK-MDR1/MRP2cellsExpressionofMDR1inCaco-2,MDCK-WT,andMDCK-MDR1CellsM1A1B2A2B3A3B4A4BKD250148M:

marker,A:

5gproteinB:

10gprotein.1:

Caco-2,2:

MDCK-WT,3:

MDCK-MDR1（lowpassage）,4:

MDCK-MDR1（highpassage）.Ref.:

Tang,F.,Horie,K,andBorchardt,R.T.（2002）AreMDCKcellstransfectedwithhumanMDR1geneagoodmodelofthehumanintestinalmucosa?

Pharm.Res.19765-772Tang,F.,Horie,K,andBorchardt,R.T.（2002）AreMDCKcellstransfectedwithhumanMRP2geneagoodmodelof