Mousemodelsofsarcomascriticaltoolsinour understandingofthepathobiology SeanMPost Abstract SarcomasareneoplasticmalignanciesthattypicallyariseintissuesofmesenchymaloriginTheidentificationof novelmole ID: 610270
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REVIEWOpenAccess Mousemodelsofsarcomas:criticaltoolsinour understandingofthepathobiology SeanMPost Abstract Sarcomasareneoplasticmalignanciesthattypicallyariseintissuesofmesenchymalorigin.Theidentificationof novelmolecularmechanismsleadingtosarcomaformationandtheestablishmentofnewtherapieshasbeen hamperedbyseveralcriticalfactors.First,thistypeofcancerisrarelyobservedintheclinicwithfewerthan15,000 newlycasesdiagnosedeachyearintheUnitedStates.Anothercomplicatingfactoristhatsarcomasareextremely heterogeneousastheyariseinamultitudeoftissuesfrommanydifferentcelllineages(e.g.bone(osteosarcoma), fat(liposarcoma),andmuscle(myosarcoma)).Thescarcityofclinicalsamplescoupledwithitsinherent heterogeneitycreatesachallengingexperimentalenvironmentforcliniciansandscientists.Facedwiththese challenges,therehasbeenextremelylimitedadvancementintreatmentoptionsavailabletopatientsascompared toothercancers.Inordertogleaninsightintothepathobiologyofsarcomas,scientistsarenowusing invivo mousemodelswhosegenomeshavebeenspecificallytailoredtocarrygenedeletions,geneamplifications,and pointmutationscommonlyobservedinhumansarcomas.Theuseofthesemodelorganismshasbeensuccessful inincreasingourknowledgeandunderstandingofhowalterationsinrelevantoncogenic,tumorsuppressive,and signalingpathwaysdirectlyimpactsarcomagenesis.Itisthegoalofmanyinthebiologicalcommunitythattheuse ofthesemousemodelswillserveaspowerful invivo toolstofurtherourunderstandingofsarcomagenesisand potentiallyidentifynewtherapeuticstrategies. Keywords: Sarcoma,Mousemodels,p53,Retinoblastoma(Rb),Translocation Background Sarcomasarearareformofcancerwithlessthan15,000 newcasesdiagnosedeachyearintheUnitedStates. Thoughrare,sarcomasarehighlydebilitatingmalignan- ciesastheyareoftenassociatedwithsignificantmorbidity andmortality.Sarcomasarebiologicallyveryheteroge- fromaplethoraofdifferenttissuesandcelltypes.They areclassicallydefinedbytheirtissueoforiginandareadd- itionallystratifiedbytheirhistopathologyorpatient sage atdiagnosis[1,2].Whiletheseclassificationshaveproven useful,modernbiologicalandclinicaltechniqueshavethe abilitytofurtherstratifysarcomasbasedontheirgenetic profile[1,3,4].Cytogeneticandkaryotypeanalyseshave revealedtwodivergentgeneticprofilesinsarcomas.The firstandmostsimplegeneticprofileistheobservationof translocationeventsinsarcomaswithanotherwise normaldiploidkaryotype.Ontheotherhand,mostsarco- masdisplayamorecomplexgeneticphenotype,suggest- inggenomicinstabilityplaysanimportantroleinmany sarcomas. Historicalperspective Muchofourcurrentknowledgeregardingsarcomabiol- ogyhasbeenascertainedthroughexperimentationusing highdoseirradiation,viralinfections, invitro cellline studies,andxenograftsmodels.Oneoftheearliestanimal studiesinvestigatedtheimpactoftheRoussarcomavirus onthedevelopmentofsofttissuesarcomas[5].Our knowledgeregardingradiation-inducedsarcomagenesis allyexposedtoradiumandanimalmodelssubjectedto highdoseradiationdevelopedsarcomas[6,7].Whilethe plightofthesepatientsandthesubsequentanimalexperi- mentsledtotheidentificationofacauseandeffectfor somesarcomas,theseobservationswereunabletoidentify themoleculareventsresponsibleforsarcomagenesis. Correspondence: spost@mdanderson.org DepartmentofLeukemia,M.D.AndersonCancerCenter,1515Holcombe Blvd,Houston,TX77030,USA CLINICAL SARCOMA RESEARC H ©2012Post;licenseeBioMedCentralLtd.ThisisanOpenAccessarticledistributedunderthetermsoftheCreativeCommons AttributionLicense(http://creativecommons.org/licenses/by/2.0),whichpermitsunrestricteduse,distribution,and reproductioninanymedium,providedtheoriginalworkisproperlycited. Post ClinicalSarcomaResearch 2012, 2 :20 http://www.clinicalsarcomaresearch.com/content/2/1/20 Tomoreaccuratelyinvestigatethegeneticandmo-lecularchangesmanifestedinsarcomas,scientistsbeganusingpatientderivedsarcomacelllines.Thesecelllineshavealsoaddedtoourunderstandingofthesarcomadiseaseprogressioninvivo,throughtheiruseinxeno-graftexperiments[8-10].Eventhoughthesecelllineexperimentshavegreatlyadvancedourunderstandingofsarcomas,theyhaveseverelimitations.First,patient-derivedcelllinesaretypicallyisolatedduringsurgicalre-sectionoflatestagetumors[11].Thus,thesecellshaveundergonenumerousgeneticalterations,complicatingourabilitytoidentifythecriticalprimaryandsecondarygeneticcausesofthesecancers.Second,celllinesiso-latedfromindividualspossessdiversegeneticback-groundsashumansharbormillionsofsinglenucleotidepolymorphiccombinations[12].Finally,someofthecur-rentlyavailablesarcomacelllineshavebeenpassagedformorethanageneration.Theimpactofcellcultureshockiswelldocumentedandnodoubtaltersthemuta-tionrateandgeneticstabilityofthesecelllines[13].Howeachoftheseimpactsanindividualtumororitsresponsetotherapyislargelyunknown.Asecondcomplicationarisesfromtheuseofimmuno-compromisedxenograftmousemodelstransplantedwithhumansarcomacells.Theseexperimentshavetheabilitytotestthetumorformingpotentialofaparticularcellline;however,theyfailtofaithfullyrecapitulatethetrueinvivoenvironmentofasarcomaastheylackafunc-tionalimmunesystem[14].Itiswidelyappreciatedthattheimmunesurveillancesystemplaysacriticalroleintumorprevention[15].Furthermore,stromalinteractionsbetweenthehostandtheinjectedcelllinesdiffersignifi-cantlyandundoubtedlyalternormalmicroenvironmentinteractions.Giventhesecaveats,ithasbecomeimperativethatresearchersgeneratemoreaccurateanimalmodelsthatwillallowscientiststodirectlyinvestigatethemechan-ismsofsarcomagenesis.Inthisreview,wewillhighlightseveralmodelsengineeredtoharborknowntransloca-tionsthoughttodrivehumansarcomagenesisaswellastumorpronemodelswithanincreasedpropensityforsarcomaformation.Whilethisreviewisnotmeanttobecomprehensiveofallsarcomamodels,wewilldiscusshowspecificgeneticalterations,pathways,andanimalmodelsmayserveaspreclinicalmodelsforfuturestud-ies,andthusprovideaframeworkforotherstudiesexaminingtheimpactoftranslocationsorderegulatedpathways.SarcomasdefinedbytranslocationAsalludedtoabove,somesarcomasharbordiploidkaryo-typesbutposseschromosomaltranslocation,suggestingadirectcorrelationbetweenthetranslocationeventandtheetiologyofthedisease[16].Thespecificityofindividualtranslocationsarelikewiseusefuldiagnosticindicatorsofspecificsarcomas.Ewing'ssarcomascommonlycarryat(11;22)(q24:q12)reciprocaltranslocationresultinginagenefusionproductbetweentheRNAbindingpro-teinEwsandthetranscriptionfactorFli1[17,18].Giventhattherearefewerthan300newEwingssarcomacasesintheUnitedStateseachyear,ourunderstandingofthediseaseprocessisquitelimited.Therefore,inordertodirectlyinterrogatetheimpactofthefusiongeneontumorformation,severallaboratorieshavegeneratedmousemodelsexpressingantransgene.Alveolarrhabdomyosarcomas,likeEwingssarcomas,arealsooftendefinedbythepresenceoftranslocationevents,mostcommonlyt(2;13)(q35;q14)andt(1;13)(p36;q14)[19,20].However,themajorityofthesearethet(2;13)(q35;q14)translocationwhichresultsinthefusionofthetranscriptionfactorPax3withthetrans-activationdomainof[21].LikeEwingssarcoma,alveolarrhabdomyosarcomasareexceedinglyrare,withfewerthan100newcasesayearreportedintheUnitedStates.Sinceclinicalsamplesaredifficulttoobtain,ourknowledgeofthisdiseaseisquitesparse.Tocombatthisdilemma,severalmousemodelsmimickingthealveolarrhabdomyosarcomatranslocationeventshaverecentlybeengenerated.ThegenerationandcharacterizationofthealveolarrhabdomyosarcomaandEwingssarcomamousemodelsandtheirimpactontumorformationwillbedetailedinlatersections.SarcomaswithcomplexkaryotypesIncontrasttosarcomasidentifiedashavingdiploidkar-yotypes,themajorityofsarcomasbelongtothemorekaryotypicallycomplexgroup.Cytogeneticandkaryo-typicanalysesofundifferentiatedpleomorphicsarcomas,pleomorphicrhabdomyosarcomas,embryonalrhabdo-myosarcomas,andosteosarcomashaverevealedtheirgenomestobeunstableanddisorganizedasevidencedbymultipledeletions,amplifications,andchromosomalfusions[22].Molecularanalyseshaveshownthatmanyofthecanonicaltumorsuppressorpathways,suchasthep53andretinoblastomapathwaysareablatedinthesetumors[22].Furthermore,somesarcomasalsoharboractivatingoncogenicmutations;suchasexpres-sionofoncogenicK-ras.Together,disruptionofthesegenesandpathwaysarethoughttobeadrivingforceinsarcomagenesis.Unlikethedirectcorrelationbetweenasinglechromo-somaltranslocationeventindiploidsarcomas,itismorechallengingtoidentifywhichofthenumerousmuta-tions,deletions,oramplificationsdrivethedevelopmentofsarcomaswithcomplexcytogenetics.Furthermore,theextremeheterogeneityinthesesarcomasisalsoachallengeforcliniciansattemptingtodeveloppersona-lizedtreatmentstrategies.Giventhesecomplexities,weClinicalSarcomaResearchPage2of9http://www.clinicalsarcomaresearch.com/content/2/1/20 willhighlightsomeofthecriticalpathwaysthoughttobealteredduringsarcomagenesisbelow.TumorsuppressorandoncogenicpathwaysinvolvedinsarcomagenesisThep53pathwayThep53tumorsuppressorpathwayisoneofthemostwellcharacterizedpathwaysincancers[23].Thegeneencodesatranscriptionfactorrequiredfortheacti-vationofnumerousDNAdamage-dependentcheckpointresponseandapoptoticgenes[24,25],andthusitsactiv-itiesareoftenablatedinmanycancers.Inadditiontolossofp53functionsviainheritedgermlinemutations,thep53pathwayiscommonlydisruptedbypointmutationsinthegeneduringsporadicsarcomagenesis[26].However,eventhoughgenealterationsarewidelyregardedashavingasignificantimpactonsarcomagen-esis,manysarcomasretainwildtype,yetphenotypic-allydisplayalossofp53function.Thesefindingssuggestthatchangesinothercomponentsofthep53pathway;suchasamplificationofMdm2,anegativeregulatorofthep53pathway,mayresultinp53inactivation[27,28].Furthermore,bothmiceandhumanswithelevatedlevelsofMdm2duetoahighfrequencysinglenucleotidepoly-morphismintheMdm2promoter(Mdm2SNP309)aremoresusceptibletosarcomaformation[29-31].Add-itionally,deletionorsilencingofArfArfhuman),aninhibitoroftheMdm2-p53axis,oftenresultsindevelopmentofsarcomas.Together,thesedataindi-catethatwhileinactivationofthep53pathwayisobservedinthevastmajorityofhumansarcomas,themechanismsleadingtodisruptionofthepathwaycanvarygreatly.TheretinoblastomapathwayTheretinoblastoma(Rb)pathwayrepresentsasecondmajortumorsuppressorpathwayderegulatedinmanysar-comas.Individualsinheritingagermlinetypicallydevelopcancersoftheeyeearlyinlife[32-34].However,inadditiontoretinalcancers,thesechildrenhaveasignificantlyhigherpropensitytodevelopsarcomasthanthegeneralpopulation[35].Whileinheritanceofagerm-linealterationsincreasessarcomarisk,therearealsonumerousexamplesofsporadicsarcomasharboringspon-mutationsanddeletions,particularlyosteosar-comasandrhabdomyosarcomas[36].Furthermore,Ikn4aanegativeregulatoroftheCDK-cyclincomplexesthatphos-phorylateandactivateRb,isoftendeletedinsarcomas[37,38].Together,thesefindingsillustratetheimportanceoftheRbpathwayinsarcomagenesis.OncogenicsignalingInadditiontolossoftumorsuppressorpathways,sarcoma-genesisisalsodrivenbyaberrantoncogenicsignaling.TheRassignalingpathwayinparticularisthoughttobealteredduringsarcomadevelopment[39].DeregulationoftheRaspathwayaberrantlystimulatescellularproliferation,whichinandofitselfimpingesonthep53andRbpathways,col-lectivelydemonstratingthesignificantcross-talkbetweenthesethreeseparatebutoverlappingpathways.Giventhenumeroussignalingpathwayspotentiallydis-ruptedinsarcomas,therehasbeenacriticalneedtointer-rogatehoweachofthesegenesanddivergentpathwaysimpactsarcomagenesisinaprospectivemanner.Sincethesestudiesarenearlyimpossibleinhumanpatients,scientistsandcliniciansarenowusingmicegeneticallytai-loredforsuchstudies(Table1).Below,wewillhighlightseveralwellcharacterizedgeneticallyengineeredmousemodelsharboringcommongeneticalterationsobservedinsarcomabiology.MousemodelsofsarcomasFormanyyears,mousemodelshaveservedaspowerfultoolsinourinterrogationofthemechanismsregulatinghumancancers.However,itwasnotuntiltheprevalenceofgeneticallymanipulablemousemodelsinthe1980and90sthatwebecamefullycapableofexaminingthedirectcausesofmanycancersinaninvivosetting.Al-thoughwedonotfullyunderstandthediseaseprocessesofsarcomagenesis,wenowhaveamplebiologicalreagentsinwhichtoexploretheseprocesses,severalofwhicharedetailedbelow.MousemodelsharboringtranslocationssSarcomaSarcomaswithsimplediploidkaryotypesoftenhavechromosomaltranslocationsthatdirectlyimpactsarcoma-genesis.ToidentifytheimpactoftheEws-Fli1transloca-tion,t(11;22)(q24:q12),inEwingssarcoma,miceharboringtransgenehavebeengenerated.Expressionoftransgeneislethalwhenexpressedinsometis-sues[57].Therefore,tolimitthislethalphenotype,theEws-transgenemustbeconditionallyexpressedinspecificcelltypesusingtheCre-recombinase-loxPsystem[58].Cre-loxPtechnologieshavetheabilitytodeleteentiregenes,specificexons,orevenremoveinhibitorsoftrans-genicexpressioninspecificcelllineagesortissues[59].Usingthissystem,transgenicmiceharboringalatentEws-transgeneweregeneratedandcrossedwithmiceexpressingCre-recombinaseunderthecontrolofthePrxpromoter[44],resultingintheactivationofthetransgenespecificallyinosteogenicmultipotentcells.Al-thoughthesemicedevelopedmultipleboneabnormalities,theyultimatelyfailedtoproducesarco-mas.Thisfindingsuggeststhatwhilethet(11;22)(q24:q12)translocationisacommoneventinEwingssarcoma,itis,byitself,unabletostimulateacancerphenotypewhichindicatesthatotheraccompanyingmutations(orhitsClinicalSarcomaResearchPage3of9http://www.clinicalsarcomaresearch.com/content/2/1/20 thegenome)arerequiredforfranktumorformation.Toaddressthis,miceexpressingthetransgenewerethencrossedtomiceharboringPrx-Cre-directeddeletion.Themicerapidlydevelopedpoorlydifferentiatedsarcomas(medianageof21weeks);whilemediateddeletionofp53aloneresultedinthedevelopmentofosteosarcomas(medianageof50weeks),demonstratingthecooperativeinteractionsbetweenEws-Fli1andp53insarcomas.AlveolarrhabdomyosarcomasAlveolarrhabdomyosarcomasareoftencharacterizedbyt(2;13)(q35;q14)translocations.Knock-inmiceharboringthet(2;13)(q35;q14)translocationhavebeengeneratedbyknockingthegeneintothePax-3locus,result-inginaPax-3-FkhrfusiongeneunderthecontroloftheendogenousPax-3promoter[49].SimilartotheCre;Ews-Fli1studies,thesemicedidnotdevelopsarco-mas,butdiddisplaynumerouscongenitaldefects,sug-gestingthePax3-Fkhrfusiongeneisimportantinnormalmurinedevelopmentbutrequiresadditionalgen-etichitsforsarcomadevelopment.Inordertogenerateamorerobustalveolarrhabdomyosarcomamodel,micespecificallyexpressingaPax3-FkhrtransgeneinthemuscleundertheinfluenceofMyf6-Cre-mediatedacti-vationweregenerated[50,51].Surprisingly,thesemice Table1MousemodelsofhumansarcomasTumortypeGenealteration(s)AgentusedSignificanceProposedkaryotypeReferenceSoftTissueSarcomaNDRoussarcomaViralinfectioninfluencessarcomagenesisUnknown[OsteosarcomaNDRadiationRadiationinfluences7osteosarcomaformationinrabbitsUnknown[Sporadic/Variedp53NoneLossofresultsinsarcomaformationComplex[Sporadic/VariedmutationNoneMutationsinp53resultsinsarcomaformationComplex[Sporadic/31Variedworw/oNoneMutationsinthep53pathwayresultinsarcomaformationComplex[PoorlyDifferentiatedworw/oPrx-CreEws-Fli1micefailtoinducesarcomagenesisintheabsenceofTranslocationbutcomplexwithp53/Rblossloss44]PoorlyDifferentiatedSarcoma/Osteosarcomaworw/oRbandp53deletionsPrx-CreEws-Fli1micefailtoinducesarcomagenesisbutreducetimeofonsetintheabsenceofTranslocationbutcomplexwithwith45]OsteosarcomaTax;p19ArfNoneExpressionofTaxintheabsenceofp19ArfresultsinosteosarcomaformationComplex[OsteosarcomaRbandp53-CreDeletionofcooperateintheboneleadingtoosteosarcomagenesisComplex[Osteosarcoma/Leiomyosarcomaandp107deletionshaploinsufficiencycoupledwithdeletionresultsinlowpenetrantSarcomaComplex[DevelopmentaldefectsPax-3-FkhrfusionNonePax3-FkhrfusionproductfailstoproducesarcomaswhenexpressedfromPax3promoterTranslocation[RhabdomyosarcomasPax3-Fkhrw/andw/oMyf6-CreExpressionofthePax3-FkhrtransgenerequireslossofrhabdomyosarcomaformationTranslocationbutcomplexcomplex50,51]Rhabdomyosarcomas/VarioussarcomasPtch1heterozygosityw/andw/oMyf6-Cre,Myf-5Cre,Pax7-CreERLossofinconjunctionwithhaploinsufficiencyresultsinrhabdomyosarcomagenesisComplex[UndifferentiatedPleomorphicMutantK-rasexpressionandAdenoviralCreMutantK-rasexpressionandcooperateinthedevelopmentofundifferentiatedpleomorphicsarcomasComplex[PleomorphicRhabdomyosarcomasMutantK-rasexpressionandofCreintotheMutantK-rasexpressionandcooperateinthedevelopmentofundifferentiatedpleomorphicsarcomasComplex[RhabdomyosarcomasMutantK-rasexpressionandlossormutationAh-CreExpressionofmutantp53facilitatesamorerapiddevelopmentofrhabdomyosarcomasthanlossofinthemutantK-rasbackgroundComplex[ClinicalSarcomaResearchPage4of9http://www.clinicalsarcomaresearch.com/content/2/1/20 alsofailedtodisplayasarcomaphenotype.However,concomitantdeletionofArf,orintheMyf6-Cre;Pax3-Fkhrmiceresultedinarhabdomyosar-comaphenotype[50,51].Thesestudiesillustratethecomplexitiesinalveolarrhabdomyosacromagenesisandimplicatethep53andRbpathwaysinthedevelopmentofPax3-Fkhr-dependentsarcomas.AdditionalsarcomamousemodelsregulatedbytransloactioneventsSynovialsarcomas/myxoidliposarcomasTheidentificationofcommontranslocationeventshasgreatlyassistedinourunderstandingofsarcomagenesisandhasledtothegenerationofmousemodelswiththepowertoexaminetheirimpact.Inadditiontothetrans-locationsnotedabove,chromosomalrearrangementst(X;18)andt(12;16)(q12;p11)arecommonlyobservedinsynovialandliposarcomas,respectively(Table2).Mousemodelsmimickingthet(X;18)translocation,viaexpres-sionofthechimericproteinSYT-SSX2,resultinsyn-ovialsarcomaswithhighpenetrance[60,61].Likewise,expressionofTLS-CHOP,afusionproteinthatmimicsthet(12;16)(q12;p11)translocation,resultedinmyxoidroundcellliposarcomas[62].Giventherarenatureofthesetumors,thesemousemodelsmakeexcellentplat-formsforinvestigatingthepathobiologyofthesedis-easesaswellaspre-clinicaltherapeuticmodels[76,77].SarcomamousemodelswithcomplexgeneticsSarcomasofthebone(osteosarcomas)Incontrasttothesarcomasdrivenprimarilybyspecifictranslocations,themajorityofsarcomaspossesshighlyaneuploidgenomesduetodisruptionsintumorsuppres-sorpathwaysandaberrantoncogenicactivation.Osteo-sarcomasareoneofthemostwellstudiedtypesofsarcomaswithcomplexgeneticsgiventhedevelopmentofnumerousknock-out,knock-in,andtransgenicanimalmodelsavailableforthisdisease.Thegenerationandcharacterizationoftumorsfrom-nullandheterozygousknock-outmicedemonstratedtheimport-anceofp53inosteosarcomas[40,41].Theroleofp53inosteosarcomasisfurtherhighlightedbytumorana-lysisofknock-inmicecontainingamutantcopyofp53R172H(correspondingtotheR175Hhot-spotmuta-tioninhumans)[42,43].Animportantdifferentiationbetweentheknock-outandknock-inmiceisthatsarcomasdevelopedametastaticgainoffunctionphenotype,faithfullyrecapitulatingthepheno-typeobservedinthehumandisease[42,43].Thegener-ationofthemutantR172Hmousemodelprovidesresearchers,forthefirsttime,withtheabilitytoinvesti-gatemetastaticosteosarcomadiseaseprogressioninainvivosetting.Inadditiontodirectablationofp53function,transgenicmiceoverexpressingthep53regula-tor,Mdm2,aswellasmiceharboringasinglenucleotidepolymorphismintheMdm2promoter,haveanincreasedrisktodevelopsarcomas[31,68].Furthermore,trans-genicmiceexpressingtheviraloncogeneTax,coupledwithdeletionofArf,developedhighlypenetrantos-teosarcomas[46].Together,theseresultsfurtherdem-onstratetheimportanceofablatingthep53pathwayinosteosarcomagenesis.Inhumans,lossoftheRbpathwayhasalsobeenimplicatedintheetiologyofosteosarcomas.However,inthemouse,homozygousdeletionofresultsinanembryolethalphenotypeduetoplacentaldefects[69].Therefore,inordertoinvestigatetheroleofRbinbonemalignancies,researchersagainemployedtheCre-loxPsystemtodeletespecificallyinthebone.UnlikethecriticalroleofRbinhumanosteosarcomas,micelackinginosteocytesdonotdevelopcancers[47].However,whencoupledwithloss,lossexacerbatesthep53-dependentosteosarcomaphenotype,withmostmicesuc-cumbingtotheirdiseasewithin150days[45,47].Asacaveattothefindingthat-lossalonedidnotinduceosteosarcomas,thereissignificantredundancyintheRb Table2AdditionalmousemodelsofhumansarcomasTumortypeGenealteration(s)AgentusedSignificanceProposedkaryotypeReferenceSynovialSarcomaSYT-SSX2fusionMyf5-CreExpressionoftheSYT-SSX2transgeneresultedin100%penetrantsynovialsarcomasTranslocation[MyxoidLiposarcomaTLS-CHOPfusionw/p53deletionPrx-CreDeletionofcooperatesintheformationofliposarcomasTranslocation[NeurofibromaMPNSTworw/op19Arf3.9Periostin-CreDeletionofandp19ArfstimulateMPNSTdevelopmentComplex[UterineleiomyosarcomaLmp2deletionNoneLossofLmp2resultsinuterineleiomyosarcomaformationComplex[UterineleiomyosarcomaTDGF1/CRIPTOoverexpressionMMTV-promoterTDGF1/CRIPTOexpressionresultsinuterineleiomyosarcomadevelopmentComplex[ClinicalSarcomaResearchPage5of9http://www.clinicalsarcomaresearch.com/content/2/1/20 pathwayinmice.Rbconsistsofthreefamilymembers(p105,p107,andp130)andeachsharessimilarstructureandfunction[70].Assuch,concomitantlossofbothp107inmousedidinfactresultinalowpenetrantosteosarcomaphenotype[48,71].Takentogether,thesestudiesdemonstratetheabsoluterequirementforablationofthep53pathwayinosteosarcomagenesisandsuggestthatpRbplaysaco-operativeroleinosteosarcomagenesis.SofttissuesarcomasUndifferentiatedpleomorphicsarcomasUndifferentiatedpleomorphicsarcomasaresofttissuesarcomastypicallyobservedinadultsthatarisefromcellsofunknownorigin,and,likeosteosarcomas,displaycomplexgeneticsresultingfromderegulationofmultiplepathways.Investigationsintothecellularoriginofbothundifferentiatedpleomorphicsarcomasandembryonalrhabdomyosarcomashaveidentifiedtheimportanceofthep53andRbpathwaysintheetiologyofbothmalig-nancies[52].Inadditiontotheimportanceofthesetwotumorsuppressorpathways,theKras-signalingpathwayhasalsobeenimplicatedinthedevelopmentofundiffer-entiatedpleomorphicsarcomas[53,54].MiceharboringalatentcopyofoncogenicKrasLSLG12D(silencedbyafloxedloxP-stop-loxP(LSL)cassette)andtwofloxed)thatweresimultaneouslyactivatedtoexpressmutantanddeletefollowinginjectionofadenoviral-Creintothemuscle,rapidlydevelopedsarco-maswithsignificantmetastaticpotential.Detailedmolec-ularanalysisoftheAd-cre;KrasG12Dtumorsrevealedanexpressionprofilesimilartothoseobservedinhumanundifferentiatedpleomorphicsarcomas[54].Together,thesedatasupporttheideathatbothablationoftumorsuppressorpathwaysandactivationofoncogenescooper-atetodrivesarcomagenesis.RhabdomyosarcomasUsingtheCre-LoxPstrategytosimultaneouslyactivateala-tentoncogenicK-rasG12Valleleanddeletethe2-10allelesinmyocytes,itwasdemonstratedthatmicerapidlydevelopsarcomasthatarehistopathologicallysimilartopleomorphicrhabdomyosarcomasobservedinhumans[55].AlthoughtheundifferentiatedpleomorphicandrhabdomyosarcomastudiesusedsimilarmousemodelstoidentifytheroleofmutantK-rasand-lossinsarcoma-genesis,theseexperimentsresultedinsomewhatdifferentmalignancies.Thus,giventhecellularsimilaritiesbetweenundifferentiatedpleomorphicsarcomasandrhabdomyo-sarcomas[52],itisimperativetofurtherinvestigatesarco-magenesisintheKras-LSLG12DFl210/Fl2mousemodelsusingmultiplemyospecificCre-expressingtrans-genicmiceinordertopreciselyascertainhowthesepath-wayssynergiesinspecifictissues.WhileeachoftheKras-LSL10/Fl2studiesmentionedaboverevealtheimportanceofp53andK-rasinmyocytespecificsarcomagenesis,theyfailedtoac-curatelyrepresentthemostcommontypeofalterationtothegeneinhumancancers(e.g.p53mutations).Arecentstudyexaminedtheimpactofp53insarcoma-genesismoreaccuratelybynotonlydeletingalsoexpressingthemutant(correspondingtothehumanp53R175hotspotmutation)inthemuscle[56].UsingtheKrasLSLG12V;p53Fl210/Fl2KrasLSLG12VR172H/Fl2allelesincombinationwithAh-Creexpres-sion,itwasrevealedthatexpressionofmutantp53,eveninthecontextofheterozygosity(e.g.,p53R172H/+),hadamoredeleteriouseffectthansimplylosingonewildtypeallele.TheseAh-Cre;KrasG12VR172H/miceformedrhabdomyosarcomaswithhighpenetranceascomparedtolessthan10%rhabdomyosarcomasformationintheCre;KrasG12Vmice.Inaddition,unlikethetumorsAh-Cre;KrasG12Vmice,thetumorsfromtheAh-Cre;KrasG12V;p53R172H/micealsorecapitulatedthemetastaticphenotypetypicallyobservedinhumanrhabdomyosarcomas.AdditionalsarcomamousemodelsregulatedbydrivermutationsNeurofibromatosis/leiomyosarcomasGiventheextremeheterogeneityofsarcomaswithregardstotissueoforigin,itisobviousthatalterationstonumerousgenes,pathways,andsignalingcomplexesplayanimportantroleinthepathobiologyofsarco-mas.Whilethisreviewdoesnotcoverallgeneticalterationsresponsibleforsarcomadevelopment,therearenumerousadditionalgenesthatimpactthisdisease(Table2).Forexample,alterationsinexpressionoftumorsuppressorgenes,suchasNeurofibromatosistype1(NF1),likewiseimpacttheetiologyofsomesar-comas.Mousemodelsthatcarrygenomicdeletionsand/ortissue-specificCre-mediateddeletionofresultinneurofibromas[72].TheseNF1-dependentphenotypesarefurtherexacerbatedwheniscon-comitantlydeletedwithothertumorsuppressors(e.g.;)resultinginmoreaggressivephenotypesasevidencedbymalignantperipheralnervesheathtumorformation[63,64].Tofurtherillus-tratethatlossofasinglegeneimpactssarcomaforma-tion,miceharboringanLMP-2deletionresultedinspontaneousuterineleiomyosarcomas[65].Thispro-videsevidenceofitsroleasatumorsuppressorandapotentialbiomarkerinhumandisease[66,73].Inadditiontolossoffunctionalterations,overexpressionofteratocarcinoma-derivedgrowthfactor1,alsoknownasCRIPTO,resultsinleiomyosarcomasbyde-regulationoftheWNTpathway[67].ClinicalSarcomaResearchPage6of9http://www.clinicalsarcomaresearch.com/content/2/1/20 Thevastdifferencesinthecellularoriginsofsarcomas,thelackofavailabilityoftumorspecimens,andthehetero-geneityinherentwithinindividualtumorshasimpededourabilitytofullyunderstandthebiologyofsarcomas.However,giventheavailabilityofnumerousgeneticknock-outs,knock-ins,andconditionalallelescoupledwiththebevyoftissue-specificCre-recombinaseexpres-singmouselines,wenowhavetheabilitytosystematicallyandprospectivelyinterrogatehowindividualgenesandmutationsimpactsarcomagenesis.Goingforward,tumoranalysisfrommultiplemurinederivedtumortypescanbecomparedandcontrastedinordertoidentifycriticalchangesinspecificsarcomas.Thesemousemodelshaveclearlydemonstratedthatwhiletherearedrivermuta-tions/translocations,sarcomagenesisis,infact,amulti-hitdisease.Theuseofthesemousemodelsmimickingthehumandiseaseconditionleadstoacriticalquestion:whattherapeuticapproachescanbetakentolessentheimpactofthesedebilitatingdiseases?First,wemustrecognizethatthesemousemodelsdemonstratethesynergismbetweenmultiplepathwaysandthuscombinatorialtreatmentstrat-egiesareneededtocombatthesecancers.Fortreatmentofpatientswithtranslocations,onecanenvisionatargetedtherapeuticapproach,likethatwhichhasbeenobservedinthetreatmentofchronicmyeloidleukemia.Theadditionoftyrosinekinaseinhibitors(TKIs),suchasima-tinib,whichinhibitstheactivityoftheBCR-ABLfusiongene,hasreducedCMLfromadeathsentencetoaman-ageableandstabledisease.Canthescientific/clinicalcom-munitydesigntargetdrugstothetranslocationeventsobservedinsarcomas?Theuseofthesemousemodelsmayserveasanexcellentpreclinicalplatformforsuchstudies.Treatingandalleviatingthediseaseprocessinsarcomaswithcomplexgeneticsmayprovemoredifficultthaniden-tifyingtargetedtherapies.However,giventhatmanygroupshaveidentifiedtheimportanceofspecificpathwaysinsarcomagenesis,suchasthep53pathway,wehaveastartingpoint.PreclinicaldrugslikePRIMA1-MetandNCS319726havebeenshowntorestoremutantp53activ-ities[74,75].Thesedrugscouldberapidlyscreenedforef-ficacyinmutantp53sarcomamodels.Moreover,thep53pathwayisalsoinactivatedbydysregulationofitsproteinpartners,Mdm2andp19Arf.TheemploymentofMdm2-p53antagonists,suchasNutlin-3andRITAmayproveef-ficaciousinreactivatingthep53pathwayandthusprovideatherapeuticbenefit.Also,lossofp19ARFduetopromotermethylationisacommoneventinsarcomagenesis.There-fore,theseanimalmodelsmayproveusefulinexaminingtheimpactofhypomethylatingagents,suchasazacytidineordasatinib,insarcomas.Incaseswherespecificoncogenesareknowntodrivetumorformation,suchasactivatedK-ras,theuseofcompoundsthatinhibitK-rastargets(suchasMEK)couldbebeneficial.TheefficacyofaMEK-inhibitorlikeARRY-162couldbereadilyexaminedinmousemodelspossessingamutatedK-rassignalingpathway.Allofthesepotentialchemotherapeuticagents,ifproveneffectiveininvivoclinicalmodels,couldprovidearationaleforpersonalizedandtargetedtherapyinsarcomapatients.Whilemousemodelscannotcompletelypredicttheout-comeofeachdisease,theycanprovidevaluableandcriticalinformation,particularlyinexceedinglyraretypesofsarco-masorwhenlowpenetrantsinglenucleotidepolymorph-ismsconfounddataanalysis.Rb:Retinoblastoma;Cre:Cre-recombinase.CompetinginterestsTheauthordeclarethathehavenocompetinginterests.SMPwrotethemanuscript.AcknowledgementsIwouldliketothankMs.XiaoruiZhangforreviewingthemanuscript.Received:27March2012Accepted:16July2012Published:4October20121.LasotaJ,Fanburg-SmithJC:Geneticsforthediagnosisandtreatmentofmesenchymaltumors.SeminMusculoskeletRadiol2.DavicioniE,etalMolecularclassificationofrhabdomyosarcomagenotypicandphenotypicdeterminantsofdiagnosis:areportfromtheChildren'sOncologyGroup.AmJPathol3.HelmanLJ,MeltzerP:Mechanismsofsarcomadevelopment.NatRev4.TaylorBS,etalAdvancesinsarcomagenomicsandnewtherapeuticNatRevCancer5.RousP:ASarcomaoftheFowlTransmissiblebyanAgentSeparablefromtheTumorCells.JExpMed6.MartlandHS:Theoccuranceofmalignancyinradioactivepersons.AmericanJournalofCancer7.SabinFR,DoanCA,ForknerCE:TheProductionofOsteogenicSarcomataandtheEffectsonLymphNodesandBoneMarrowofIntravenousInjectionsofRadiumChlorideandMesothoriuminRabbits.JExpMed8.BudachV,etalRadioresponsivenessofahumansofttissuesarcomaxenografttodifferentsingleandfractionatedregimens.Strahlenther9.KodousekR,etalHistopathologicalandultrastructuralobservationsofsomehumantumorxenograftspassagedinathymicnudemice.I.Humanosteosarcomaxenograftwithfeaturesofviral(oncornaAandCtyperetrovirus)involvement.ActaUnivPalackiOlomucFacMed10.PimmMV,BaldwinRW:Serologicalaspectsofrattumourxenograftgrowthinathymicnudemice.BrJCancer11.LinPP,etalSurgicalmanagementofsofttissuesarcomasofthehandandfoot.12.IstrailS,etalWhole-genomeshotgunassemblyandcomparisonofhumangenomeassemblies.ProcNatlAcadSciUSA13.SherrCJ,DePinhoRA:CellularSenescence:MinireviewMitoticClockorCultureShock?14.PelleitierM,MontplaisirS:Thenudemouse:amodelofdeficientT-cellMethodsAchievExpPathol15.WherryEJ:Tcellexhaustion.NatImmunol16.FletcherJA:Cytogeneticsandexperimentalmodelsofsarcomas.CurrOpinOncolClinicalSarcomaResearchPage7of9http://www.clinicalsarcomaresearch.com/content/2/1/20 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