KU-55933-DataSheet-生命科學試劑-MedChemExpress

Hotline:400-820-3792Inhibitors?ScreeningLibraries?Proteinswww.MedChemEKU-55933Cat.No.:HY-12016CASNo.:587871-26-9分?式:C??H??NO?S?分?量:395.49作?靶點:ATM/ATR;Autophagy作?通路:CellCycle/DNADamage;PI3K/Akt/mTOR;Autophagy儲存?式:Powder-20°C3years4°C2yearsInsolvent-80°C6months-20°C1month溶解性數(shù)據(jù)體外實驗DMSO:80mg/mL(202.28mM;Needultrasonic)MassSolvent1mg5mg10mgConcentration制備儲備液1mM2.5285mL12.6425mL25.2851mL5mM0.5057mL2.5285mL5.0570mL10mM0.2529mL1.2643mL2.5285mL請根據(jù)產(chǎn)品在不同溶劑中的溶解度選擇合適的溶劑配制儲備液；?旦配成溶液，請分裝保存，避免反復凍融造成的產(chǎn)品失效。儲備液的保存?式和期限：-80°C,6months;-20°C,1month。-80°C儲存時，請在6個?內(nèi)使?，-20°C儲存時，請在1個?內(nèi)使?。體內(nèi)實驗請根據(jù)您的實驗動物和給藥?式選擇適當?shù)娜芙?案。以下溶解?案都請先按照InVitro?式配制澄的儲備液，再依次添加助溶劑：(為保證實驗結(jié)果的可靠性，澄的儲備液可以根據(jù)儲存條件，適當保存；體內(nèi)實驗的?作液，建議您現(xiàn)?現(xiàn)配，當天使?；以下溶劑前顯?的百分?指該溶劑在您配制終溶液中的體積占?；如在配制過程中出現(xiàn)沉淀、析出現(xiàn)象，可以通過加熱和/或超聲的?式助溶)1.請依序添加每種溶劑：10%DMSO>>40%PEG300>>5%Tween-80>>45%salineSolubility:≥2.5mg/mL(6.32mM);Clearsolution1/3MasterofBioactiveMolecules—您?邊的抑制劑?師www.MedChemE2.請依序添加每種溶劑：10%DMSO>>90%(20%SBE-β-CDinsaline)Solubility:≥2.5mg/mL(6.32mM);Clearsolution3.請依序添加每種溶劑：10%DMSO>>90%cornoilSolubility:≥2.5mg/mL(6.32mM);ClearsolutionBIOLOGICALACTIVITY?物活性KU-55933有效的ATM抑制劑，IC50和Ki值分別為12.9和2.2nM；對ATM的選擇性?對DNA-PK，PI3K/PI4K，ATR和mTOR?。IC50&TargetATMDNA-PKmTORPI3K12.9nM(IC50)2500nM(IC50)9300nM(IC50)16600nM(IC50)體外研究KU-55933(10μM)blockstheionizingradiation-inducedp53serine15phosphorylation.KU-55933hasadose-dependenteffectininhibitingthisATM-dependentphosphorylationeventwithanestimatedIC50of300nM.KU-55933ablatestheionizingradiation-inducedphosphorylationoftheseATMsubstrates.KU-55933specificallyinhibitsATMbutnottheotherDNAdamage-activatedPIKKs,ATR,andDNA-PK[1].KU-55933inducespATM,p53,E2F1andpATR,noticeablyupregulatesthenuclearfractionofE2F1atthe0.5htimepoint[2].MetforminincreasesATMandAMPKphosphorylation,aswellasSHPproteinlevelinprimaryhepatocytes,andthisstimulatoryeffectofmetforminisrepressedbyaspecificATMkinaseinhibitorKU-55933[3].PROTOCOLKinaseAssay[1]ATMforuseintheinvitroassayisobtainedbyimmunoprecipitationwithrabbitpolyclonalantiserumraisedtotheCOOH-terminal400aminoacidsofATMinbuffercontaining25mMHEPES(pH7.4),2mMMgCl2,250mMKCl,500μMEDTA,100μMNa3VO4,10%v/vglycerol,and0.1%v/vIgepal.ATM-antibodycomplexesareisolatedfromnuclearextractbyincubatingwithproteinA-Sepharosebeadsfor1hourandthenthroughcentrifugationtorecoverthebeads.Inthewellofa96-wellplate,ATM-containingSepharosebeadsareincubatedwith1μgofsubstrateglutathioneS-transferase-p53N66(NH2-terminal66aminoacidsofp53fusedtoglutathioneS-transferase)inATMassaybuffer[25mMHEPES(pH7.4),75mMNaCl,3mMMgCl2,2mMMnCl2,50μMNa3VO4,500μMDTT,and5%v/vglycerol]at37°Cinthepresenceorabsenceofinhibitor.After10minuteswithgentleshaking,ATPisaddedtoafinalconcentrationof50μMandthereactioncontinuedat37°Cforanadditional1hour.Theplateiscentrifugedat250×gfor10minutes(4°C)toremovetheATM-containingbeads,andthesupernatantisremovedandtransferredtoawhiteopaque96-wellplateandincubatedatroomtemperaturefor1.5hourstoallowglutathioneS-transferase-p53N66binding.ThisplateisthenwashedwithPBS,blotteddry,andanalyzedbyastandardELISAtechniquewithaphospho-serine15p53antibody.ThedetectionofphosphorylatedglutathioneS-transferase-p53N66substrateisperformedincombinationwithagoatantimousehorseradishperoxidase-conjugatedsecondaryantibody.EnhancedchemiluminescencesolutionisusedtoproduceasignalandchemiluminescentdetectioniscarriedoutviaaTopCountplatereader.MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly.2/3MasterofBioactiveMolecules—您?邊的抑制劑?師www.MedChemECellAssay[1]1BRorAT4cellsareseededin10-cmPetridishesandtreatedonday2(80to90%confluence).Cellsarepreincubatedfor1hourwithKU-55933orvehiclecontrolandthenexposedto5Gyofionizingradiation.Timecoursesofcellcycledistributionareperformed,andtheoptimaltimefordiscriminationofpopulationsisselectedas16hours.Allsubsequentexperimentsareperformedatthe16-hourtimepoint.CellsarestainedwithpropidiumiodideaccordingtostandardprotocolsandanalyzedbyFACSwithaFACScalibur.Exponentiallygrowing(50-70%confluent)SW620cellsin60mmdishesareexposedtoKU-55933orDMSOfor1hbeforeadditionofetoposide(finalconcentrationof0.1and1μM)for16hbeforeharvesting,propidiumiodidestainingandanalysisasabove.MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly.戶使?本產(chǎn)品發(fā)表的科研?獻?CancerCell.2021Apr12;39(4):566-579.e7.?MolCell.2022Apr14:S1097-2765(22)00290-8.?NucleicAcidsRes.2020Sep18;48(16):9109-9123.?NatCommun.2019Aug21;10(1):3761.?JExpClinCancerRes.2019Aug22;38(1):370.Seemorecustomervalidationsonwww.MedChemEREFERENCES[1].HicksonI,etal.Identificationandcharacterizationofanovelandspecificinhibitoroftheataxia-telangiectasiamutatedkinaseATM.CancerRes.2004Dec15;64(24):9152-9[2].KhalilHS,etal.PharmacologicalinhibitionofATMbyKU55933stimulatesATMtranscription.ExpBiolMed(Maywood).2012Jun;237(6):622-34.Epub2012Jun22.[3].KimYD,etal.OrphannuclearreceptorSHPnegativelyregulatesgrowthhormone-mediatedinductionofhepaticgluconeo