药物ADME在药物开发中的应用

TheRoleofADMEinDrugDiscoveryandDevelopmentDecember20112Whathappensafteryoutakeapill...3•ADME−Absorption(Howmuch?Howfast?)−Distribution(Howextensive?Howfast?Whereisthedrugdistributed?)−Metabolism(Howfast?Whatmechanism/route?Whatmetabolites?Aretheyactive?Aretheytoxic?)−Excretion(Howfast?Whichroute?)•MeasureandunderstandADMEandyoucanpredictpharmacokinetics•ThesemeasurementsarekeyapplicationsforDrugdevelopmentanddiscoveryWhatisADME?4DrugDiscovery&DevelopmentTargetSelectionHTStoIDhitsPreclinicalTestingIND•Genomics•Proteomics•Metabolomics•Lipidomics•Animalmodels•Knockouts•ADME/Tox•CYPInhibition•MetStab•CaCo-2•ProteinBinding•GSHBinding•LogP•Solubility•Cell&AnimalTesting•PK/PD•Toxicology10106610103310101111•Label-FreeHTS•NumberoftestcompoundsSecondaryScreenHTHTADME/ADME/ToxToxSelectivityScreenMedChem/SARADME/ToxDevelopmentDiscovery5Whatisthe“right”answer?•FavorableADMEpropertiesisachievingabalance.Forexample,−Enoughsolubilitygiventhepermeabilityandexpecteddose−Enoughmetabolismsothatthedrugdoesn’taccumulatetotoxiclevelsbutnotsomuchthatitneedstobeadministeredmorethanonceaday−Enoughdifferentroutesofeliminationsuchthatcompromisingoneofthemdoesnotcausetoxiclevelsofthedrug−Evendrug-druginteractionscanbeacceptableifthedrugistheonlyonetobeeffectiveforalifethreateningindication•ResearchersconsidertheentireprofileofADMEproperties.Theyshouldnotoverfocusononeparameter.6Outline•Pemeability•InVitroDM−Metabolicstability−Drug-DrugInteraction(DDI)•CYPinhibition•CYPinduction•Reactionphenotyping7Outline•Pemeability•InVitroDM−Metabolicstability−Drug-DrugInteraction(DDI)•CYPinhibition•CYPinduction•Reactionphenotyping8Permeability•Theoralrouteisthemostconvenientandcheapestrouteofadministration•However,theintestinalwallcanpresentasignificantbarriertodrugentry•Permeabilityassaysmeasuretheabilityofdrugstopassthrougha“surrogate”oftheintestinalwall
Caco-2cells,ahumancolonderivedcellline
PAMPA,anartificialmembranesystem•Thesepermeabilitymeasurementshavebeenshowntocorrelatewithdrugabsorption9ExperimentalSystems•Caco-2−Cellsaregrownonamultiwellinsertandallowedtodifferentiatetoformabarrier−Compoundisaddedtothe“apical”side−Therateofappearanceofthecompoundonthe“basolateral”sideismeasuredbymassspec•PAMPA−Artificialmembraneismadeusinglipidsand/oroilandspottedinafilter−Compoundisaddedtothelowerchamber−TherateofappearanceofthecompoundintheupperchamberismeasuredbymassspecorUVKeyProduct:PremadePAMPAPlate10Caco-2vs.PAMPAEasy,fastpreparationRequirescellculture(upto21days)AssaypreparationLowHighCostTypeofPermeabilityTypeofmodelMeasurespassivepermeabilityinabsenceoftransportersoreffluxsystemsMeasuresthesumofpassiveandactivepermeabilityArtificialmembraneCellmonolayerPAMPACaco-2Note:PAMPAandCaco-2testscanbecomplementaryteststodeterminebothpassiveandactivepermeabilities.Caco-2testsalonemeasurethesumofpassiveandactivepermeabilitieswhichcannotbedecoupledwithouttheinformationobtainedfromPAMPAtests.11Outline•Pemeability•InVitroDM−Metabolicstability−Drug-DrugInteraction(DDI)•CYPinhibition•CYPinduction•ReactionphenotypingApplication:Metabolicstability13MetabolicStability•Scientistsareinterestedinmeasuringsomethingcalled“clearance”•Conceptually,thisistherateatwhichthedrugiseliminatedbythebody•Themetabolicenzymes“clear”thedrugfromthebodybymetabolizingittosomethingelse•Therateofclearanceisveryimportantindetermininghowmuchdruggetsthroughtheliverandhowlongadrugwillstayinthebody−“Howbigapillisneededandhowoftenyouneedtotakeit”14EnzymeLocationandP450Mechanism15PhaseI/PhaseIIReaction16PhaseI/IIDrugMetabolism17Metabolicstability•Researchersareinterestedinmeasuringsomethingcalled“clearance”•Conceptually,thisistherateatwhichthedrugiseliminatedbythebody•Themetabolicenzymes“clear”thedrugfromthebodybymetabolizingittosomethingelse•Therateofclearanceisveryimportantindetermininghowmuchdruggetsthroughtheliverandhowlongadrugwillstayinthebody−“Howbigapillisneededandhowoftenyouneedtotakeit”18ExperimentalApproach•Measurethelossofdrugcompoundovertimeinthepresenceofanactivemetabolicelement•Activemetabolicelementscouldbe:−Livermicrosomes(orS9)+NADPH−Hepatocytes•Alowdrugconcentrationisused−Betterapproximatesinvivoexposures−Hasbeenfoundtobemorepredictive•MeasuretheamountofparentdrugremainingbyLC-MS/MS•Calculatethehalflifeand“scaleup”toinvivobasedonwellknownscalingparametersKeyProducts:Metabolism-QualifiedHepatocytes,BDUltraPool™HLM150,Animallivermicrosomes19MicrosomalStabilityAssayMethodTestCompoundTestCompound(substrate2(substrate2µµM)M)NNOONNOOOHNADPH(1mM)NADPH(1mM)37°CMicrosomesMicrosomes(0.5mg/ml)(0.5mg/ml)3.63.84.4.24.44.6010203040Time(m)LogAreaSampleandanalyzebyLC-MS/MSEnzymes20ForexampleTestArticle020406080100120060120180240Incubationtime(min)Parentcompoundremaining(%)NeedData:1.Lossofdrugcompound(timedependent)2.Halflifeofcompound,T1/23.PredictClint(ul/min/million)4.MetabolitesmeasureApplication:CYPInhibition22CytochromeP450(CYP)Inhibition•WhenCYPsaretheprincipalrouteofclearance,anychangesinCYPactivitycandramaticallyaffectbloodlevelsofthedrug•SomedrugsareknowntobepotentinhibitorsofCYPs