The

225

    ;EwingFamilyofTumorsingFamilyotIu

    ;C,ancjFeng’

    ;‘DepatmentofCancer,MianyangCen—

    ;traIHospital,SichuanProvince621000,

    ;China.

    ;DepartmentofCancer.FuzhouHospi—

    ;laIofNankingMilitaryDistrict.Fujian

    ;Province350001.China.

    ;0CancerCenterofXinqiaoHospita1.the

    ;ThirdMilitaryUniversity,Chongqing,

    ;400037,China.

    ;Correspondenceto:DonglinWang

    ;E—mail:a65223682@cta.cq.cn

    ;ThisworkwassupportedbytheNational

    ;NaturaIScienceFoundationofChina.

    ;(No:30100189)

    ;ReceivedFeburary20,2004;accepted

    ;June18,2004.

    ;ChineseJoumalofClinicalOncolgy

    ;Email:COCR@eyou.comTel(Fax):86—22-2352—2919

    ;AB$TRACrThereisevidencefhaf95%offheEwingfamilyoffumors(EF-r) ;haveaEWS—FLI一1fusiongene.EWS—FLI一1isatranscriptionfactorwitha ;pivotaIfunctionanditisknownlobindloaspeciaIDNAsequence.Research ;hasdemonstratedlhallheEWS—FLI一1fusiongeneoccurrenceisrelatedlo ;lheEFT.andithasbeenusedlodiagnose.1realandserveasabasisforEFT ;prognosis.WehavebrieflysummarizedlheprogressoflheEWS-FLI一1

    ;fusiongeneinbasicandclinicaIinvestigationwithinthepastseveraIyears. ;KEYWORDS:Ewingfamilyofhsmor,fudongene.刚S—Fu一1.

    ;SincethereportofthePhiladephiachromosome(PHchromosome)in1960, ;mostofthenewfusiongenes,whichhavebeenidentified, ;arefoundinleukemiaandmalignantlymphoma.Withthedevelopment ;ofmolecularbiology,manyspecifictranslocationsofchromosomes ;havebeenfoundinsolidtumors,especiallyinsoft—tissuesarcomas

    ;derivedfrommesodermfTable1).[“

    ;TheEwingfamilyoftumors(EFT),whichderivefrom

    ;neuroectoderm,includeclassicEwing’ssarcoma,Askintumorsofthe

    ;chestwall,andperipheralprimitiveneuroectodermaltumorsofboneor ;softtissues.ThefirstcaseofEwingfamilyoftumorswasdescribed

    ;nearly80yearsago.butnowitoccursannuallyintheUSAata ;frequencyof2.7casespermillionchildrenundertheageof15years ;annuallyintheUSA.AlthoughEFTcanOCCt~atanyage,thegreat ;majorityofcasesoccuratlessthan20yearsofage.Approximately95% ;ofEFThaveaspecifictranslocation,t(1l;22)(q24;q12),whichresults ;infusionoftheEWSandFLI—lgeneandproductionofanewfusion

    ;proteinEWS—FLI一1.Experimentsandclinicalcaseshaveshownthat ;EWS—FLI一1isinvolvedintumorigenesisandprogressionofEFT,andit ;hasbecomeusedfordiagnosis,treatmentandabasisofprognosis.H-7] ;oftheEWS—FU一1fusiongene

    ;Thespecifict(11;22)(q24;q12)chromosomaltranslocationoccurring ;in95%ofEFTiscausedbyrearrangementofthe5’halfoftheEWS

    ;geneonchromosome22q12withthe3’halfoftheFLI一1geneon

    ;

    ;226ChineseIournalofClinicalOncology2004/Volume1/Number3 ;Table1.Partiallistofthechromosometranslocationsandfusiongenesinsoft-tissuesarcomas

    ;PNET:primitiveneuroectodermaltumor;NK:notknown;RMS:rhabdomyosarcoma ;chromosomel1q24.IllThe5’halfoftheEWSgene

    ;containsatransformationdomainwhichcontainsan

    ;abundanceofglutamineandthe3’halfoftheEWSgene

    ;includesaTATA-bindingprotein(TBP)codetypicalof ;aRNA—bindingfactor.EWSisafunctional

    ;RNA—bindingproteinthat,alongwithTLSandanovel

    ;TATA—bindingprotein—associatedfactor,TAFII68

    ;(alsoknownasRBP56),formsanewfamilyofrelated

    ;proteins.BothTLSandEWScanfunctionas

    ;TATA—bindingprotein—associatedfactors(TAFIIs).[41

    ;ThissuggeststhatEWSmaymediatethestepsof

    ;nascenttranscriptswiththebasaltranscription

    ;apparatus.However,theEWSportionofEWS—FLI—l

    ;mayberedirectedtootherproteintargetsbecauseof

    ;chromosomalconformationchangesinducedbythe

    ;genefusion.[81FLI—lisamemberoftheETS

    ;(erythroblastosisvirus—associatedtransforming

    ;sequences)familyoftranscriptionfactorswhichcan ;activatespecifictargetgenesbybindingtotheircognate ;DNAsequencesthroughtheirDNA—bindingregions.

    ;RecentresearchhasshownthatFLI—lhasaclose

    ;relationshiptocellimmortalizationresultingincancer. ;StudiesalsohaveshownthatFLI—lhasarelationto

    ;vascularizationandgrowthofneuroectoderm.The5’

    ;halfoftheFLI—lgenecontainsatransformationdomain

    ;andthe3’halfoftheFLI—lgeneisaDNA—binding

    ;domain(DBD).TheETSfamilymembersaredefined

;bythepresenceofahighlyconserved85一aminoacid

    ;domainnamedtheETSdomain.Theconserveddomain ;isthemainpartoftheDBDintheETSfamily. ;ComputeranalysisshowsthattheN—terminalofthe

    ;ETSdomainhasaahelicalstructureandthe ;C—terminalhasaconservedstructure.TheDNA ;sequenceboundbyETScontainsapurine—rich

    ;sequencecenteredaroundaconservedGGAA,1【

    ;polynucleotideandisnotthesameindifierentmembers ;oftheETSfamily.【9】IntheEWS—FLI—lfusionprotein.

    ;theRNA—bindingmotifcontainingtheC—terminalhalf

    ;OfEWSisreplacedbytheDBDoftheFLI—lprotein.

    ;TheEWS—FLI—lisformedbythetransformation

    ;domainofEWSandDBDofFLI—1.Itisbelievedthat

    ;theEWS—FLI—lisanewtransformationfactor.which ;canrecognizeaspecialdomainupstreamfi-omthegene ;andcanactivatethetransformation.IntheEWS—FLI—l

    ;fusionprotein.boththeN—terminaldomainofEWSand

    ;theDBDofFLI—larenecessaryforthetransforming ;activity.Researchhasindicatedthattheactivationof ;transformationbytheEWS—FLI—lismuchstrongerthan

    ;thatoftheFLI—linnormalcells.anditevencan

    ;transfcIrillculturedNIH3T3cells.I10]Toensurethatthe ;DBDdomaininthe3’endofFLI—linEwing’ssarcoma

    ;canrecognizeandbindwithaspecialgenesequence, ;Maoeta1.【ll1testedthebindingactivityof22different ;genesequencesandfoundthatACCGGAAGTwasthe ;best.

    ;Ithasbeendemonstratedinclinicalstudies.that ;variousEFTshavedifierentclinicalbehaviors, ;progressionandtherapeuticefficacies.These ;characteristicshaveacloserelationtothedifferent ;fusiontypesthatarecausedbyvariationsi11the ;

    ;locationsoftheEWSandFLI—lgenomicbreakpoints.

    ;Meloteta1.?reportedthatatleastl2typesof

    ;EWS—FLI—lchimerictranscriptshadbeenobservedin ;clinicalinvestigations.andthefusionofEWSexon7to ;FU一1exon6(type1)orFLI一1exon5(type2)accounted

    ;forabout60%and25%ofEWS—FLI—lfusions

    ;respectively.Thedifferentfusiontypeshaveno ;significantdifferenceinbiologicalactivityandthe ;fusionoftheN—terminaldomainofEWS.encodedby

    ;exonsl一7,withDBDofFLI一1,encodedbyexon9,are

    ;theminimalcomponentsneedforactivety.The ;differenceinclinicalbehavior,progressionand ;therapeuticefficacyforEFTisduetothecomponentsof ;EWSandFLI一1.Theclinicalbehaviorandtherapeutic ;efficacyoffusiontype1isfoundintumorsless ;malignant.In1999,Lineta1.【圳usedanelectrophoretic

    ;mobilityshiftassay(EMSA)todetecttheradioactivity ;oftheisotope—labeledDNA.containingthe

    ;ACCGGAAGTsequence.in7difierentEWS—FLI—l

    ;fusiontypesofEFTandfoundthattheywerealmostthe ;same.TheyalsoconstructedareporterplasmidpS2by ;ligatingtheACCGGAAGTsequenceintotheSacIand ;KpnIsitesofpGL3一Promoter(Promega).Tocompare

    ;theratioofluciferaseactivity.thepS2wastransferred ;usingliposomesinto7differentEwing’ssarcomacells

    ;andcontrolcellsfHelaandNIH3T3).Theresults ;showedthattheratioofluciferaseactivityinthe ;Ewing’ssarcomacellswasmuchhigherthanthatin

    ;conffolcellsfP=0.003)andthattheACCGGAAGT ;wasthebestbindingsequenceofEWS—FLI—1.

    ;ClinicaI

    ;gene

    ;Itisobviousthatthemoleculardiagnosisofcancer ;dependsonspecialmolecularmarkers.Becausespecial ;molecularmarkersusedfordiagnosisarefew,the ;clinicalapplicationofmoleculardiagnosisislimited. ;Fusiongenesoftumorscausedbyspecialchromosomal ;translocationsarehighlyspecificandhaveapotentialof ;becomingatoolformoleculardiagnosis,histological ;typingandjudgingtherapeuticefficacy,reoccurrence ;andprognosis.[131Clinicalresearchesarenowfocusing ;onhowtousetheEWS..FLI..1fusiongenetodetermine ;EWS-FLI-1fusiongene/GangFengeta1.227

    ;theoptimalEFTtherapy.

    ;Clinicianshavedifficultyindiagnosis,treatmentand ;prognosisofEFT,becausethehistologicaland ;immunohistochemicaldifferencesarefewbetweenEFT ;andthesmallblueroundcelltumors(SBRCP).We ;knowthat95%ofEFThaveaspecificchromosome ;translocation,(11;22)(q24;q12),andthatthefusionof ;EWSandFLI一1leadstohighexpressionofFLI一1.To

    ;evaluatetheexpressionofFLI一1in132SBRCP

    ;determinedbypathology.Folpeeta1.[14]usedanti—FLI一1

    ;polyclonalantibodyandimmunohistochemical

;technology.TheyfoundthattheexpressionofFLI—1

    ;wasseenin29of41(71%)ES/PNET,7of8(88%) ;lymphoblasticlymphomas,1ofasingle(100%) ;desmoplasticround—celltumorfithastheEWS—FLI一1

    ;gene),0of8poorlydifferentiatedsynovialsarcomas ;(PDSS),0of32rhabdomyosarcomas(RMS),0of30 ;neuroblastomas(NB),0of8esthesioneuroblastomas ;(ENB),0of3Wilms’tumorsand0ofasingle

    ;mesenchymalchondrosarcoma.Theirresearch ;demonstratedthatimmunohistochemistrycould ;distinguishEFTfromSBRCP.

    ;AtonetimeantibodiesforCD99weresuggestedasa ;specificmarkerforEFT.butnowweknowthatitiSnot ;specific.TheexpressionofCD99isonthecell ;membranebutEWS—FLI一1isinthenucleus.Specificity ;foranuclearmarkerisgreaterthanoneonthecell ;membrane.Forexample.NBandENBtumorsare ;positivewhentestedbyFLI—lantibodybuttheyare

    ;almostalwaysnegativewhentestedbyCD99antibody. ;TheexpressionofFLI一1isalsohelpfulindistinguishing ;EFTfromothertumorsthatmaybeCD99一positive,

    ;suchasPDSS.

    ;Inrecentyears,autologousbonemarrow

    ;transplantationhasbeenusedtotreatEFT.Toavoid ;re—implantationofthetumorcellsandEFT

    ;re—occurrence,onemustmakesurethatnotumorcells ;remaininthegraft.Traditionaltechnologyhasbeen ;usedtodetecttheexistenceoftheEFTcells,butthe ;sensltl’Vl’tyisnothigh.around1%.UsingRT—PCR

    ;technology.Vichaeta1.【7】foundtheEWS—FLI一1fusion

    ;genetobenegativein31bonemarrowsamplesand5 ;bloodsamples,and7differentfusiontypeswerefound ;in7autologousbonemarrowtransplantations.This ;

    ;228ChineseJournalofClinicalOncology2004/Volume1/Number3

    ;researchsuggeststhatRT—PCRismoresensitivethan

    ;histologyindetectingtheexistenceofEFTcells,anda ;moreimportantassayinconductingautologousbone ;marrowtransplantations.

    ;TheEWS—FLI—lfusiongeneisfoundin95%ofEFT

    ;andthisfusiongeneisfundamentalfortheoccurrence ;anddevelopmentofEFT.EWS—FLI—lisatranscription

    ;factorthathasapotentactivitybybindingwitha ;specialDNAsequencewhichleadstoahighexpression

;ofitselfandthedown—streamgenes.TheEWS—FLI—l

    ;hasalreadybeenusedinthediagnosis.treatmentand ;prognosisofEFT,andisbecomingatargetforgene ;therapy.Genefusionisverycommonincancer.The ;importantgoalnowishowtoutilizetheknowledge ;abouttheEWS—FLI.1fusiongeneinrelationtoEFTas ;wellastoothertumors.

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