SorafenibisapotentmultikinaseinhibitorwithIC₅₀sof6nM,20nM,and22nMforRaf-1,B-Raf,andVEGFR-3,respectively.

IC50:6nM(Raf-1),20nM(VEGFR-3),22nM(BRAF),57nM(PDGFR-β),58nM(Flt3),68nM(c-KIT),90nM(VEGFR-2)^[1]

Sorafenib(BAY43-9006)alsoinhibitsBRAF^wt(IC₅₀=22nM),BRAF^V599E(IC₅₀=38nM),VEGFR-2(IC₅₀=90nM),VEGFR-3(IC₅₀=20nM),PDGFR-β(IC₅₀=57nM),c-KIT(IC₅₀=68nM),andFlt3(IC₅₀=58nM)inbiochemicalassays.InMDA-MB-231breastcancercells,SorafenibcompletelyblocksactivationoftheMAPKpathway.CellsarepreincubatedwithSorafenib(0.01to3μM),anddose-dependentinhibitionofbasalMEK1/2andERK1/2phosphorylation(IC₅₀,40and100nM,respectively)^[1].

Sorafenibdemonstratesbroadoralantitumorefficacyinpanelofhumantumorxenograftmodels.Sorafenibisgivenorallyat7.5to60mg/kg.Thereisnolethalityandnoincreaseinweightlossinanytreatedgrouprelativetothecorrespondingcontrolgroup.DailyoraladmiNISTrationofSorafenib(30to60mg/kg)producescompletetumorstasisduringtreatmentinfiveofthesixmodels^[1].Thesurvivalrateis73.3%inDiethylnitrosamine(DENA)groupand83.3%inSorafenibgroupcomparedto100%inthenormalcontrolgroup.DENAgroupshowsasignificantincreaseinliverindex(1.51-foldincrease,p<0.05) compared="" to="" normal="" control="" group,="" while="" treatment="" with="" sorafenib="" shows="" significant="" decrease=""><0.05) in="" liver="" index="" when="" compared="" to="" dena="" group.="" the="" liver="" index="" in="" sorafenib="" group="" significantly="" decreases="" to="" lower="" than="" its="" value="" in="" the="" normal="">^[2].

• [1].WilhelmSM,etal.BAY43-9006exhibitsbroadspectrumoralantitumoractivityandtargetstheRAF/MEK/ERKpathwayandreceptortyrosinekinasesinvolvedintumorprogressionandangiogenesis.CancerRes.2004Oct1;64(19):7099-109.

  [2].El-AshmawyNE,etal.Sorafenibeffectonliverneoplasticchangesinrats:morethanakinaseinhibitor.ClinExpMed.2016Apr16.

  [3].JinW,etal.Longnon-codingRNATUC338isfunctionallyinvolvedinsorafenib-sensitizedhepatocarcinomacellsbytargetingRASAL1.OncolRep.2017Jan;37(1):273-280.

  [4].LiM,etal.ActivationofanAKT/FOXM1/STMN1pathwaydrivesresistancetotyrosinekinaseinhibitorsinlungcancer.BrJCancer.2017Aug29.

TotestcompoundinhibitionagainstvariousRAFkinaseisoforms,SorafenibisaddedtoamixtureofRaf-1(80ng),wtBRAF,orV599EBRAF(80ng)withMEK-1(1μg)inassaybuffer[20mMTris(pH8.2),100mMNaCl,5mMMgCl₂,and0.15%β-mercaptoethanol]atafinalconcentrationof1%DMSO.TheRAFkinaseassay(finalvolumeof50μL)isinitiatedbyadding25μLof10μMγ-[³³P]ATP(400Ci/mol)andincubatedat32°Cfor25minutes.PhosphorylatedMEK-1isharvestedbyfiltrationontoaphosphocellulosemat,and1%phosphoricacidisusedtowashawayunboundrADIoactivity.Afterdryingbymicrowaveheating,aβ-platecounterisusedtoquantifyfilter-boundradioactivity^[1].MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly.

SorafenibisdissolvedinDMSOandstored,andthendilutedwithappropriatemediabeforeuse^[1].

TheMDA-MB-231humanmammaryadenocarcinomacelllinesareplatedat2×10⁵cellsperwellin12-welltissuecultureplatesinDMEMgrowthmedia(10%heat-inactivatedFCS)overnight.Cellsarewashedoncewithserum-freemediaandincubatedinDMEMsupplementedwith0.1%fattyacid-freeBSAcontainingvariousconcentrationsofBAY43-9006(0.01,0.03,0.1,0.3,1,3μM)in0.1%DMSOfor120minutestomeasurechangesinbasalpMEK1/2,pERK1/2,orpPKB.CellsarewashedwithcoldPBS(PBScontaining0.1mMvanadate)andlysedina1%(v/v)TritonX-100solutioncontainingproteaseinhibitors.Lysatesareclarifiedbycentrifugation,subjectedtoSDS-PAGE,transferredtonitrocellulosemembranes,blockedinTBS-BSA,andprobedwithanti-pMEK1/2(Ser²¹⁷/Ser²²¹;1:1000),anti-MEK1/2,anti-pERK1/2(Thr²⁰²/Tyr²⁰⁴;1:1000),anti-ERK1/2,anti-pPKB(Ser⁴⁷³;1:1000),oranti-PKBprimaryantibodies.Blotsaredevelopedwithhorseradishperoxidase(HRP)-conjugatedsecondaryantibodiesanddevelopedwithAmershamECLreagentonAmershamHyperfilm^[1].MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly.

SorafenibisdissolvedinCremophorEL/ethanol(50:50;CremophorEL,95%ethylalcohol)at4-fold(4×)ofthehighestdose,foilwrapped,andstoredatroomtemperature(Mice)^[1].

Mice^[1]
FemaleNCr-nu/numiceareused.Micebearing75to150mgtumorsaretreatedorallywithSorafenib(7.5to60mg/kg),administereddailyfor9days.Ineachmodel,Sorafenibproducesdose-dependenttumorgrowthinhibitionwithnoevidenceoftoxicity,asmeasuredbyincreasedweightlossrelativetocontrolanimalsordrug-relatedlethality.Inparalleltotheantitumorefficacystudies,additionalgroupsoffourmicebearing100to200mgtumorsaretreatedorallywithvehicleorSorafenib(30to60mg/kg),administereddailyfor5days,whichistheshortesttreatmentdurationproducingcompletetumorstasisinthetreatedgroups.
Rat^[2]
Inthestudy,100-to120-gmalealbinoratsareutilized.Afteracclimatizationperiod,ratsareweighedandrandomlydividedintothreegroups:Group1(normalcontrolgroup;n=10)isgiventhevehicledailyfor8weeks.Group2(DENAgroup;n=15)receivei.p.singledoseof200mg/kgDENA.Group3(Sorafenibgroup;n=12)isgivenSorafeniborallyatadoseof10mg/kgdailyfor2weeks,6weeksafterDENAi.p.injection.Attheendoftheexperiment(8weeks),ratsareweighed,anesthetizedbyether,andkilled,andtheirliversaredissected.Freshliveriswashedtwicewithice-coldsaline,driedoncleanpapertowel,andweighed.Liverindexiscalculatedasliverweight(g)/finalbodyweight(g)×100.Theliverisdividedintofiveportions:oneportionispreservedin10%formalinforhistopathologicalexaminationandtheotherportionsareimmediatelyfrozeninliquidnitrogenandstoredat−80°C.MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly.

• [1].WilhelmSM,etal.BAY43-9006exhibitsbroadspectrumoralantitumoractivityandtargetstheRAF/MEK/ERKpathwayandreceptortyrosinekinasesinvolvedintumorprogressionandangiogenesis.CancerRes.2004Oct1;64(19):7099-109.

  [2].El-AshmawyNE,etal.Sorafenibeffectonliverneoplasticchangesinrats:morethanakinaseinhibitor.ClinExpMed.2016Apr16.

  [3].JinW,etal.Longnon-codingRNATUC338isfunctionallyinvolvedinsorafenib-sensitizedhepatocarcinomacellsbytargetingRASAL1.OncolRep.2017Jan;37(1):273-280.

  [4].LiM,etal.ActivationofanAKT/FOXM1/STMN1pathwaydrivesresistancetotyrosinekinaseinhibitorsinlungcancer.BrJCancer.2017Aug29.

464.83

C₂₁H₁₆ClF₃N₄O₃

284461-73-0

RoomtemperatureincontinentalUS;mayvaryelsewhere

DMSO:≥45mg/mL

SorafenibisdissolvedinDMSOandthendilutedwithsaline(thefinalDMSOconcentrationis<>^[3].
Sorafenibispreparedinvehicle(saline)^[4].

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

• [1].WilhelmSM,etal.BAY43-9006exhibitsbroadspectrumoralantitumoractivityandtargetstheRAF/MEK/ERKpathwayandreceptortyrosinekinasesinvolvedintumorprogressionandangiogenesis.CancerRes.2004Oct1;64(19):7099-109.

  [2].El-AshmawyNE,etal.Sorafenibeffectonliverneoplasticchangesinrats:morethanakinaseinhibitor.ClinExpMed.2016Apr16.

  [3].JinW,etal.Longnon-codingRNATUC338isfunctionallyinvolvedinsorafenib-sensitizedhepatocarcinomacellsbytargetingRASAL1.OncolRep.2017Jan;37(1):273-280.

  [4].LiM,etal.ActivationofanAKT/FOXM1/STMN1pathwaydrivesresistancetotyrosinekinaseinhibitorsinlungcancer.BrJCancer.2017Aug29.