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首页 2012新型抗流感病毒药物

2012新型抗流感病毒药物.pdf

2012新型抗流感病毒药物

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2012-05-02 0人阅读 举报 0 0 暂无简介

简介:本文档为《2012新型抗流感病毒药物pdf》,可适用于自然科学领域

Design,Synthesis,andinVitroBiologicalEvaluationofH,,TriazolecarboxamideDerivativesasNewAntiinfluenzaAAgentsTargetingVirusNucleoproteinHuiminCheng,†,⊥JuntingWan,†MengILin,‡YingxueLiu,†XiaoyunLu,†JinsongLiu,†YongXu,†JianxinChen,§ZhengchaoTu,†YihShyunECheng,‡andKeDing*,††KeyLaboratoryofRegenerativeBiologyandInstituteofChemicalBiology,GuangzhouInstitutesofBiomedicineandHealth,ChineseAcademyofSciences,KaiyuanAvenue,Guangzhou,People’sRepublicofChina‡GenomicsResearchCenter,AcademiaSinica,Taipei,,Taiwan,RepublicofChina§SouthChinaAgriculturalUniversity,WushanRoad,Guangzhou,People’sRepublicofChina⊥GraduateSchoolofChineseAcademyofSciences,#YuquanRoad,Beijing,China*SSupportingInformationABSTRACT:Theinfluenzavirusnucleoprotein(NP)isanemergingtargetforantiinfluenzadrugdevelopmentNucleozin()anditscloselyrelatedderivativeshadbeenidentifiedasNPinhibitorsdisplayingantiinfluenzaactivityUtilizingasaleadmolecule,wesuccessfullydesignedandsynthesizedaseriesofH,,triazolecarboxamidederivativesasnewantiinfluenzaAagentsOneofthemostpotentcompounds,b,inhibitedthereplicationofvariousHNandHNinfluenzaAvirusstrainswithICvaluesrangingfromtoμMCompoundbalsostronglyinhibitedthereplicationofHN(RG),amantidineresistantAWSN(HN),andoseltamivirresistantAWSN(HN,Y)virusstrainswithICvaluesinsubμMrangesFurthercomputationalstudiesandmechanisminvestigationsuggestedthatbmightdirectlytargetinfluenzavirusAnucleoproteintoinhibititsnuclearaccumulation■INTRODUCTIONInfluenzaisaseasonallyepidemicacuterespiratorydiseasepersistentlythreateningpublichealthcausedbyinfectionofribonucleicacid(RNA)influenzavirusesoforthomyxoviridaefamilyTheepidemicandpandemicsofinfluenzahavecausedseriousimpactonworldwidemorbidity,mortality,andeconomyThreedistincttypesofinfluenzavirusesareclassifiedbasedontheirserologicalsubtypes,amongwhichinfluenzaAvirusesarethemostvirulenthumanpathogensInfluenzaAvirusespossesseseightsegmentedRNAgenomesThecontinuousgeneticmutationandreassortmentofinfluenzaAvirusesmaycausetheevasionofprimaryhumanimmunitiesandleadtotheresistanceagainstcurrenttherapiesMoreimportantly,theviralgeneticshiftmayeventuallycausethebreakthroughofinterspeciesandinterhumantransmissionbarrierswhichwillbedisastroustopublichealth−Currently,twotypesofsmallmoleculardrugsareavailableforthetreatmentofinfluenzaAincludingMionchannelblockers(ie,amantidineandrimanitidine)andneuraminidaseinhibitors(ie,oseltamivirandzanamivir)−However,theapplicationofMionchannelblockers,particularlyforamantidine,hasbeenstrictlylimitedbecauseoftherapidemergenceofdrugresistanceandtheoccurrenceofcentralnervoussideeffectsOseltamivirresistantviruseshavealsobeenreportedsince,,andalmostalltheseasonalinfluenzaHNvirusescirculatingintheUnitedStateswereresistanttoOseltamivirinthe−fluseason−Mostrecently,casesofreassortmentbetweenswineinfluenzaandinfluenzaA(HN)inhumanwerereported,whichhighlightstheemergentneedfordevelopingnewantiinfluenzaagentswithnewmechanismofactionTheinfluenzavirusnucleoprotein(NP)isencodedbythefifthgenomesegmentandabundantlyexpressedduringthecourseofinfectionComparingwiththeviralsurfacespikeproteinslikehemagglutininandneuraminidase,theinfluenzavirusnucleoproteinishighlyconserved−Duringthevirallifecycle,nucleoproteinbindswithinfluenzaviralRNAsegmentsandpolymerasesubunitproteins(PB,PB,andPA)toformvirusribonucleoprotein(vRNP)complex,whichistransportedtohostcellnucleustotriggerviralRNAtranscription,replication,andvirionassemblyOwingtoitsindispensablerolesinnumerousstagesofviralmultiplication,influenzavirusnucleoproteinhasbeenconsideredasanoveltargetfornewantiinfluenzadrugdevelopmentSeveralnucleoproteininhibitorshavebeenreportedtodisplaypromisingantiinfluenzavirusactivities−Nucleozin()isthearguablyfirstsmallmolecularinfluenzavirusnucleoproteininhibitorreportedbyYuenandSuindependently,StudieshavedemonstratedthatanditsrelatedderivativescouldpotentlytriggertheaggregationofnucleoproteinandinhibititsnuclearaccumulationtodisplayReceived:October,Published:February,Articlepubsacsorgjmc©AmericanChemicalSocietydxdoiorgjm|JMedChem,,−potentantiinfluenzavirusactivitiesFurthermutationalstudiessuggestedthattheseinhibitorsmightdirectlybindtonucleoproteinand“induceformationofhigherordernucleoproteinoligomers”thatareunabletomigrateintothenucleus−However,itremainselusivethathowtheinhibitorsinducedtheformationofnucleoproteinoligomersNoneofthecurrentinhibitorshasbeenapprovedforclinicalinvestigationItishighlydesirabletoidentifynewinhibitorsfortofurthervalidatenucleoproteinasnovelmoleculartargetforantiinfluenzaAdevelopmentHerein,wereportthestructuraldesign,synthesis,andinvitrobiologicalevaluationofH,,triazolecarboxamidederivativesasnewantiinfluenzaAagentsusingastheleadcompound■CHEMISTRYCompoundsandwerepreparedbyusingethylbenzoyloxobutanoateasthesamestartingmaterial(Scheme)Thecondensationcyclizationofethylbenzoyloxobutanoate()withhydroxylaminehydrochlorideorhydrazinehydrateyieldedethylmethylphenylisoxazolecarboxylate()orethylmethylphenylHpyrazolecarboxylate(),respectivelyCompoundorwashydrolyzedandreactedwith(chloronitrophenyl)piperazinetoproducethedesigned((chloronitrophenyl)piperazinyl)(methylphenylisoxazolyl)methanone()or((chloronitrophenyl)piperazinyl)(methylphenylHpyrazolyl)methanone()((Chloronitrophenyl)piperazinyl)(′biphenyl)methanone()wasreadilypreparedbyadirectcondensationofphenylbenzoicacid()with(chloronitrophenyl)piperazineCompoundsweresynthesizedbyusingatraditional“”cycloadditionasthekeystep−Briefly,theanilineswerediazotizedandthenreactedwithNaNtoproducetheazidobenzenesCompoundsweredirectlyreactedwithSchemeSynthesisofCompounds,,andaaReagentsandconditions:(a)NHOH·HCl,ethanolaq,°Covernight,(b)NaOH,MeOH:THF:HO=::,overnight,(c)(COCl),dichloromethane,h,then(chloronitrophenyl)piperazine,EtN,dichloromethane,h,−(d)NH·HO,ethanol,refluxovernight,(e)BBr,DCM,−°C,h,SchemeSynthesisofCompoundsandtheXRayStructureofIntermediateaaaReagentsandconditions:(a)HOethylacetate,thenHCl,NaNO(b)NaN,°C,h,(c)toluene,reflux,overnight,()(d)LiOH,MeOH:THF:HO=::,overnight,(e)(COCl),dichloromethane,DMFcat,h,then(chloronitrophenyl)piperazine,EtN,dichloromethane,h,JournalofMedicinalChemistryArticledxdoiorgjm|JMedChem,,−differentalkynoatestoobtainthekeyintermediatesethylphenylH,,triazolecarboxylates()andthebyproductsethylphenylH,,triazolecarboxylates()Compoundsandcouldbeseparatedbysilicagelcolumnchromatography,andthestructuresofwerefurtherdeterminedbyXraycrystallographicanalysisofarepresentativecompounda(Scheme)Withthekeyintermediatesinhands,compoundswerereadilyobtainedbyhydrolysisandcouplingwithsubstitutedpiperazinesor,diazepanewithgoodyields■RESULTSANDDISCUSSIONTakingcompoundastheleadcompound,weinitiallydesignedisoxazolcarboxamide(a),H,,triazoleFigureDesignofnewantiinfluenzaAagentsusingascaffoldhoppingstrategyTableInhibitoryActivitiesoftheCompoundsagainstInfluenzaAReplicationIC(μM)compdnRRRRRRRCCc(μM)HNaHNbabababaClNOMeHHHH>bClNOMeOMeHHH>cClNOMeHOMeHH>dClNOMeHHOMeH>eClNOMeClHHH>fClNOMeHClHH>gClNOMeHHClH>>hClNOMeOHHHH>iClNOMeMeHHH>jClNOMeOEtHHH>kClNOMeOiPrHHH>lClNOMeCOMeHHH>mClNOMeOMeHHOMe>nClNOHOMeHHH>>oClNOEtOMeHHH>>>pClHMeOMeHHH>>qHNOMeOMeHHH>>rHHMeOMeHHH>>>sOMeNOMeOMeHHH>>>tClNOMeOMeHHH>>AMDd>>aInfluenzaAHK(HN)stainsbInfluenzaAWSN(HN)stainscAntiproliferationagainstMDCKcellsdAMD:amantidineThedataaremeansofresultsfromatleastthreeindependentexperimentsJournalofMedicinalChemistryArticledxdoiorgjm|JMedChem,,−carboxamide(a),Hpyrazolcarboxamide(a),andbiphenylcarboxamide(a)asnewantiinfluenzaAagentsbyusingascaffoldhoppingstrategy(Figure)TheantiinfluenzaAvirusactivitiesofthecompoundswerepreliminarilyevaluatedusingMadin−Darbycaninekidney(MDCK)cellbasedCPE(cytopathiceffectprotection)assaysUnderthescreeningconditions,inhibitedthereplicationofHN(AHK)andHN(AWSN)strains,withICvaluesofandμM,respectively,whichwerecomparabletothereporteddata,TheresultsalsosuggestedthatallthefourdesignedcompoundspotentlyinhibitedthereplicationofHN(AHK)andHN(AWSN)strains,amongwhichH,,triazolecarboxamide(a)displayedthegreatestpotencyandinhibitedthereplicationofHN(AHK)andHN(AWSN)strains,withICvaluesofandμM,respectively(Table)Althoughcompoundaisaboutfoldlesspotentthantheoriginalleadcompound,itrepresentedanewantiinfluenzaAagentwithadifferentchemicalscaffoldTherefore,furtherstructuraloptimizationwasconductedtoimproveitsantiinfluenzaactivityTheresultsweresummarizedinTableSimilartothepreviousobservationonthederivativesofcompound,,asubstitutedgroupinphenylringAmighthavegreatimpactontheantiinfluenzaactivityoftheH,,triazolecarboxamidecompoundsForinstance,whenamethoxylgroupwasintroducedatRpositionofcompounda,theantiinfluenzaactivitywasobviouslyimprovedandtheresultingcompoundbdisplayedICvaluesofandμMagainstthereplicationofHN(AHK)andHN(AWSN)strains,respectively,whichwasabouttimesmorepotentthantheoriginalcompoundaHowever,whenamethoxylgroupwasintroducedinRorRposition(bandc),thepotencywasdecreasedTheRCl−substitutedcompoundewasalsomorepotentthanthecorrespondingRorRsubstitutedcompoundforg,whichsuggestedthatRmightbeafeasiblepositionforfurtheroptimizationTherefore,avarietyofothermoietiessuchashydroxyl(h),methyl(i),ethoxyl(j),oracetyl(k)groupswereintroducedforfurtherinvestigationHowever,itwasdisappointingthatnoneofthesesubstitutionsfurtherimprovedthepotencyThe′,′dimethoxylcompoundm(R,Rdisubstitutedcompound)alsodisplayedgoodpotencyagainstthereplicationofHN(AHK)andHN(AWSN)strainsTheimpactofRwasalsoinvestigated,andtheresultsindicatedthatremovalofthemethylgroup(n)orreplacingitwithaslightlybiggerethylgroup(o)causeddramaticdecreaseofthepotencyInvestigationonthesubstitutedgroupinphenylringBrevealedthatboththenitro(R)andchloro(R)werenecessaryfortheantiinfluenzaactivityThepotenciesweresignificantlylostbyremovalofthenitroorandchlorogroups(p−s)Itwasalsodemonstratedthatreplacementofthepiperazinelinkerwith,diazenpane(t)ledtoabouttimedecreaseofthepotency(comparingwithb)Utilizingtheknowledgeofthestructure−activityrelationshiponcompounds,methoxylorchlorosubstitutedderivativesofcompoundsa,a,andawerealsodesignandsynthesizedTheresultingcompoundsb,b,andbdisplayedimprovedantiinfluenzaactivitiesagainstHN(AHK)andHN(AWSN)strains,withICvaluescomparabletothatofbAntiproliferativeactivitiesofthecompoundsagainstMDCKcellswerealsoevaluatedtomonitorthepotentialcytotoxiceffectsUndertheassayconditions,displayedmoderatetoxicityagainsttheMDCKcellswithICvalueofμMHowever,noneoftheH,,triazolecarboxamidecompoundsshowedobviouscellulargrowthinhibitionagainstMDCKcellsunderμM(Table),indicatingthatthecompoundsbeardgreatselectivityindexes(SI=CCIC)Althoughbwasabouttimeslesspotentthancompound,itdisplayedequalpotencywithamantidineagainstthereplicationofinfluenzaAHK(HN)stainswithgreatselectivityindexFurthermore,balsopotentlyinhibitedthereplicationofAWSN(HN)stainswhichwereresistanttotheclinicalMblockeramantidine(Table),suggestingthatthiscompoundmightbeusedanewleadcompoundfornovelantiinfluenzadrugdiscoveryTherefore,furtherbiologicalevaluationswereperformedtovalidateitsantiinfluenzaactivityPlaguecountingbasedviralyieldreductionassayisoneofthemostdirectandreliablemethodsforevaluatingtheinhibitoryeffectsonviralreplicationTherefore,theantiinfluenzafunctionofbwasfurthervalidatedbyusingaplaquereductionassayAsshowninFigure,compoundbdosedependentlyinhibitedtheplaqueformationofMDCKcellsbyAWSN(HN)influenzavirusinfectionTheplaqueformationwascompletelyinhibitedbybunderaconcentrationofμMHighlyconsistentwiththeresultsfromourCPEassay,compounddisplayedbetterplaguesuppressiveeffectthanbandinducedacompleteabrogationofviralplaguesatμMTheviralyieldreductionactivitiesofandbwerefurthervalidatedbyusingmicroplatecountingofinfectiousparticles(McIP)assay,andthededucedICvalueswereandμM,respectivelyTheantiinfluenzaactivitiesofbwerealsoevaluatedagainstapanelofotherinfluenzaAvirusstrains,andtheresultsweresummarizedinTableItwasshownthatbalsopotentlyinhibitedseveralotherHNandHNinfluenzaAvirusstrainswithvariedICvaluesinlowμMrange,exceptfortheHN(Br)strains(IC>μM)ItwasalsonoteworthythatbpotentlyinhibitedthereplicationofHN(RG)strains,tamifluresistantWSN(Y)AWSN(HN)strains,andAGuangdongChickenHNinfluenzavirusstrainswithICvaluesof,,andμM,respectivelyCompoundhadbeenreportedasanucleoproteininhibitorthattriggeredtheaggregationandinhibitedthenuclearaccumulationofinfluenzavirusnucleoproteins,GiventhestructuralsimilarityoftheH,,triazolecarboxamideFigureCompoundbpotentlyinhibitedtheplaqueformationofinfluenzavirusinfectedMDCKcellsMonolayerMDCKcellswereinfectedwithAWSN(HN)atMOI(multiplicityofinfection)andoverlaidwithagarcontainingvariousconcentrationsofnucleozin()(upperpanel)andb(bottompanel)Athpostinfection,plaqueswerevisualizedbystainedwithcrystalvioletTheresultsshownarerepresentativeofatleastthreeindependentexperimentsJournalofMedicinalChemistryArticledxdoiorgjm|JMedChem,,−derivativestothatof,wehypothesizedthatthenewcompoundsmightalsotargetinfluenzaAnucleoproteinsTherefore,theeffectofbonnucleoproteinswasinspectedunderfluorescencemicroscopy(Figure)Notsurprisingly,bpotentlyinhibitedthenuclearaccumulationofinfluenzavirusAnucleoproteinandcausedthenucleoproteinstobetrappedinthecytoplasmandscatteredrandomlyinHN(AWSN)virusinfectedMDCKcells,indicatingnucleoproteinsmightbethemoleculartargetoftheH,,triazolecarboxamideanaloguesMostrecently,theXraycrystalstructureofnucleoproteincomplexwithacompoundanaloguewasdisclosedItwasdemonstratedthateachinhibitorbridgedtwomoleculesofNP(ie,NPAandNPB)proteintoformahexamericcomplexTheinteractionsoftheinhibitorwiththeYregioninNPAandtheYregioninNPBwerecriticalforbindingoftheinhibitortointerfaceofNPAandNPB(FigureA,B)TheimportanceoftheYandYregionsinnucleoproteinhadalsobeenobservedbypreviousmutationalstudies,Tounderstandthepotentialinteractionofbwithinfluenzanucleoproteins,acomputationalstudywasperformedTheresultssuggestedthatbboundtonucleoproteinswithasimilarmodetothatofthenucleozinanalogues(Figure)ThecarbonylgroupofbformedanessentialhydrogenbondwiththeOHgroupoftheSresidueinNPBThenitrochlorophenylmoietymightformstrongπ−πstackinginteractionwiththeYresidueofNPA,whilethemethoxylphenylgroupandthepiperazinemoietycouldachieveimportanthydrophobiceffectwiththeYsidechainofNPBTofurthervalidatetheproposedinteraction,theantiinfluenzaactivityofbweredeterminedagainstANewJerseystrainsbearingYHmutationandrecombinantWSNvirusesharboringeitherwildtypeNP(Y)ormutatedNP(H)usingasareferencecompoundSimilartocompound,bisapotentantiinfluenzaagenttotheWSNviruswithwildtypeNP(Y)However,bothYHmutatedisogenicWSNvirusandYHmutatedANewJerseystrainswereresistanttob(IC>μM),highlightingthecriticalinteractionofbwiththesetworesiduesTheseresultswerehighlyconsistenttoourcomputationalmodelingresultsandfurthersupportedthatvirusnucleoproteinmightbethemoleculartargetoftheH,,triazolecarboxamideantifluenzaagents■CONCLUSIONInsummary,usingscaffoldhoppingandbioisostericreplacementstrategies,wesuccessfullydesignedandsynthesizedaseriesofH,,triazolecarboxamidederivativesasnewantiinfluenzaAagentsbasedonthechemicalstructureofcompoundOneofthemostpotentcompoundsbpotentlyinhibitedthereplicationofvariousHNandHNinfluenzaAvirusstrainwithICvaluesrangingfromtoμMFurthermore,balsopotentlyinhibitedthereplicationofHN(RG),amantidinresistantAWSN(HN),andtamifluresistantWSN(Y)AWSN(HN)withICvaluesof,,andμM,respectivelyFurthercomputationalstudyandmechanisminvestigationsuggestedthatbmightdirectlytargetinfluenzavirusAnucleoproteintoinhibititsnuclearaccumulationOurstudyprovidedanewseriesofleadc

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2012新型抗流感病毒药物

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