Medchemexpress/GSK126(Synonyms: EZH2 inhibitor; GSK2816126A)/HY-13470/200mg

GSK126isapotent,highlyselectiveinhibitorofEZH2methyltransferaseactivitywithIC₅₀of9.9nM.

IC50:9.9nM(EZH2)^[1]

GSK126potentlyinhibitsbothwild-typeandmutantEZH2methyltransferaseactivitywithsimilarpotencies(K_i=0.5-3nM)independentofsubstrateused,andiscompetitivewithS-adenosyl-methionine(SAM)andnon-competitivewithpeptidesubstrates.GSK126ishighlyselectiveagainstothermethyltransferasesandmultipleotherproteinclasses(EZH1,IC₅₀=680nM)^[1].TreatmentofthreeSCLCcelllineswithGSK126,inducesgrowthinhibition.SCLCcelllines(Lu130,H209,andDMS53)aretreatedwith0.5,2,and8μMGSK126,andgrowthcurveisanalyzedbyWST-8assay.InhibitionofcellulargrowthbyGSK126treatmentisobservedat8μMinallthethreecelllines,whileLu130andH209aremoresensitivetoGSK126,evenatlowerdoses^[2].

GSK126isadmiNISTeredintraperitoneallyatadosevolumeof0.2mLper20gbodyweightinfemalebeigeSCIDmice.GSK126effectivelyinhibitstheproliferationofEZH2mutantDLBCLcelllinesandmarkedlyinhibitsthegrowthofEZH2mutantDLBCLxenograftsinmice^[1].

• [1].McCabeMT,etal.EZH2inhibitionasatherapeuticstrategyforlymphomawithEZH2-activatingmutations.Nature.2012Dec6;492(7427):108-12.

  [2].SatoT,etal.PRC2overexpressionandPRC2-targetgenerepressionrelatingtopoorerprognosisinsmallcelllungcancer.SciRep.2013May29;3:1911.

Thefive-memberPRC2complex(Flag-EZH2,EED,SUZ12,AEBP2,RbAp48)containingeitherwild-typeormutant(A677G,Y641N,Y641C,Y641H,Y641SorY641F)EZH2isprepared.GSK126isdissolvedinDMSOandtestedatconcentrationsof0.6nMto300nMwithafinalDMSOconcentrationof2.5%.Incontrasttowild-typeEZH2whichprefersH3K27me0asasubstrateinvitro,EZH2Y641mutantspreferH3K27me2andhavelittleactivitywithH3K27me0orH3K27me1.TheA677Gmutantisdistinctfromboththewild-typeandY641mutantformsofEZH2inthatitefficientlymethylatesH3K27me0,H3K27me1,andH3K27me2;therefore,histoneH3peptides(residues21-44;10μMfinal)witheitherK27me0(wildtype,A677GEZH2),K27me1(A677GEZH2),orK27me2(A677G,Y641N,Y641C,Y641H,Y641SandY641FEZH2)areusedasmethyltransferasesubstrates.GSK126isaddedtoplatesfollowedbyadditionof6nMEZH2complexandpeptide.AsthepotencyofGSK126isatornearthetightbindinglimitofanassayrunat[SAM]=K_m,_IC50valuesaremeasuredatahighconcentrationofthecompetitivesubstrateSAMrelativetoitsK_m(7.5μMSAMwheretheSAMK_mis0.3μM)^[1].MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly.

GSK126isdissolvedinDMSO(10mM)andstored,andthendilutedwithappropriatemediabeforeuse^[2].

JUB-andPTRF-introducedDMS53cellsareseededatdensityof1×10³cells/wellin96-wellplate,andcellulargrowthisanalyzedusingWST-8kitat12,36,60,and84h.CellulargrowthofLu130,H209,andDMS53withtreatmentbyDZNeporGSK126isalsoanalyzedusingWST-8kit.DZNepisdissolvedinPBSat5mM,andcellsareculturedatthefinalconcentrationof5μM.GSK126isdissolvedinDMSOat10mM,andcellsareculturedat0.5,2,and8μM^[2].MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly.

GSK126isdissolvedin20%captisoladjustedtopH4-4.5with1Naceticacid^[1].

Mice^[1]
GSK126orvehicleisadministeredintraperitoneallyatadosevolumeof0.2mLper20gbodyweight.PfeifferorKARPAS-422cells(1×10⁷)in100%MatrigelareimplantedsubcutaneouslyinfemalebeigeSCIDmice.Tumorsaremeasuredwithcalipers,andblockrandomizedaccordingtotumoursizeintotreatmentgroups.Forefficacystudies,10micearerandomizedineachtreatmentgroupbeforetheinitiationofdosingandGSK126treatmentisinitiatedoncethetumourvolumesareapproximately200mm³inthePfeifferandKARPAS-422studiesand500mm³intheKARPAS-422intermittentdosingstudy.Miceareweighedandtumorsmeasuredwithcaliperstwiceweekly.Two-tailedt-testsareconductedassumingtwosamplesofequalvariance.MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly.

• [1].McCabeMT,etal.EZH2inhibitionasatherapeuticstrategyforlymphomawithEZH2-activatingmutations.Nature.2012Dec6;492(7427):108-12.

  [2].SatoT,etal.PRC2overexpressionandPRC2-targetgenerepressionrelatingtopoorerprognosisinsmallcelllungcancer.SciRep.2013May29;3:1911.

526.67

C₃₁H₃₈N₆O₂

1346574-57-9

RoomtemperatureincontinentalUS;mayvaryelsewhere

10mMinDMSO

*"<1 mg/ml"="" means="" slightly="" soluble="" or="" insoluble.="" "≥"="" means="" soluble,="" but="" saturation="">