腫瘤細(xì)胞信號(hào)轉(zhuǎn)導(dǎo)

如何閱讀文獻(xiàn)在腫瘤學(xué)方面有哪些好的雜志如何快速閱讀文獻(xiàn)以獲取知識(shí)論文的主要結(jié)構(gòu)腫瘤的分子信號(hào)轉(zhuǎn)導(dǎo)

+genomicinstabilityfromHanahanandWeinberg2000SignalTransductionandCancerLectureI:GrowthFactorsandReceptorsOutline:WhatisSignalTransduction? WhatareGrowthFactors?HowdotheycontributetonormalST?HowisthisSTderegulatedinCancer?LectureI:GrowthFactorsandReceptorsWhatisSignalTransduction?SignalTransductionistheprocessbywhichacellconvertsanextracellularsignalintoaresponse.Involvedin:Cell-cellcommunicationCell’sresponsetoenvironmentIntracellularhomeostatsis-internalcommunicationGenericSignallingPathwaySignalReceptor(sensor)TransductionCascadeTargetsResponseAlteredMetabolismMetabolicEnzymeGeneRegulatorCytoskeletalProteinAlteredGeneExpressionAlteredCellShapeorMotilityAdaptedfromMolecularBiologyoftheCell,(2002),4thedition,Albertsetal.ComponentsofSignallingWhatcanbetheSignal?ExternalmessagetothecellPeptides/Proteins-GrowthFactorsAminoacidderivatives-epinephrine,histamineOthersmallbiomolecules-ATPSteroids,prostaglandinsGases-NitricOxide(NO)PhotonsDamagedDNAOdorants,tastantsSignal=LIGANDLigand-Amoleculethatbindstoaspecificsiteonanothermolecule,usuallyaprotein,iereceptorComponentsofSignallingWhatareReceptors?Sensors,whatthesignal/ligandbindstoinitiateSTCellsurface

Intracellular

Hydrophillic

LigandCell-SurfaceReceptorPlasmamembraneHydrophobicLigandCarrierProteinIntracellularReceptorNucleusAdaptedfromMolecularBiologyoftheCell,(2002),4thedition,Albertsetal.CellSurfaceReceptorTypes:Ligand-gatedionchannelCellSurfaceReceptorTypes:2)G-ProteinCoupledReceptorCellSurfaceReceptorTypes:3)Enzyme-linkedReceptoregGrowthFactorReceptorsGrowthFactorsLigandswhichbindenzymelinkedreceptorsSignaldiversecellularresponsesincluding:ProliferationDifferentiationGrowthSurvivalAngiogenesisCansignaltomultiplecelltypesorbespecificFactorPrincipalSourcePrimaryActivityCommentsPDGFplatelets,endothelialcells,placentapromotesproliferationofconnectivetissue,glialandsmoothmusclecellstwodifferentproteinchainsform3distinctdimerforms;AA,ABandBBEGFsubmaxillarygland,Brunnersglandpromotesproliferationofmesenchymal,glialandepithelialcells

TGF-commonintransformedcellsmaybeimportantfornormalwoundhealingrelatedtoEGFFGFwiderangeofcells;proteinisassociatedwiththeECMpromotesproliferationofmanycells;inhibitssomestemcells;inducesmesodermtoforminearlyembryosatleast19familymembers,4distinctreceptorsNGF

promotesneuriteoutgrowthandneuralcellsurvivalseveralrelatedproteinsfirstidentifiedasproto-oncogenes;trkA(trackA),trkB,trkCErythropoietinkidneypromotesproliferationanddifferentiationoferythrocytes

TGF-activatedTH1cells(T-helper)andnaturalkiller(NK)cellsanti-inflammatory(suppressescytokineproductionandclassIIMHCexpression),promoteswoundhealing,inhibitsmacrophageandlymphocyteproliferationatleast100differentfamilymembersIGF-IprimarilyliverpromotesproliferationofmanycelltypesrelatedtoIGF-IIandproinsulin,alsocalledSomatomedinCIGF-IIvarietyofcellspromotesproliferationofmanycelltypesprimarilyoffetaloriginrelatedtoIGF-IandproinsulinGrowthFactorReceptorsMostgrowthfactorsbindReceptorTyrosineKinasesGrowthFactorReceptorActivationIRTKRS/TKGrowthFactorReceptorActivationIIGrowthsignalautonomy,Insensitivitytoanti-growthsignals,Resistancetoapoptosis:Uncouplecell’sgrowthprogramfromsignalsintheenvironment.Growthfactorsinnormalcellsserveasenvironmentalsignals.GrowthFactorSTandCancerGrowthfactorsregulategrowth,proliferation,andsurvival.Theseareallderegulatedincancer.HanahanandWeinberg,(2000)HallmarksofCancer,Cell(100)57GrowthfactorswithOncogenicPotentialPDGF,originallyshowntoregulateproliferation,wasalsoshowntohavehomologytov-sis,thesimiansarcomavirus.OtherviraloncogenesencodedproteinproductsthatweregrowthfactorsthatoftenoverexpressedincancersuchasTGF-a.Autocrine

signallingleadstoderegulatedgrowth.

PDGFfamily Neurotrophins

Achain NGF Bchain(c-sis) BDNFFGFFamily NT3

acidicFGF Cytokines(Hematopoietic) basicFGF IL-2EGFFamily IL-3

EGF M-CSF TGF-a GM-CSF

GFReceptorswithOncogenicPotentialEGFR,kinaseactivitystimulatedbyEGF-1andTGF-ainvolvedincellgrowthanddifferentiation,waslinkedviasequencehomologytoaknownavianerythroblastosisvirusonocgene,v-erbB.Sincethen,manyoncogeneshavebeenshowntoencodeforGFRs.EGFRfamily InsulinReceptorfamily

erbB1(c-erbB) IGF-1(c-ros) erbB2(neu) Neurotrophins

FGFFamily NGFR(trk)

FGFR-1(fig) BDNFR(trk-B) FGFR-2(K-sam) NT3R(trk-C)PDGFRFamily

CSF-1R(c-fms) SLFR(c-kit) Inductionofcancerbyalternationsinseveraltypesofproteinsinvolvedincellgrowthcontrol

SignalTransductionandCancerLectureII:IntracellularSignallingOutline:Whataresomesignallingpathways? Whataretheircellbiologicaloutputs?Howdotheseresultinthecancerphenotype?Howcanweexploitsignallingpathwaysfortherapy?GenericSignal

TransductionRTKSignal

TransductionSignalTransductionDownstreameffectorsProteinSignalingModules(Domains)SH2andPTBbindtotyrosinephosphorylatedsitesSH3andWWbindtoproline-richsequencesPDZdomainsbindtohydrophobicresiduesattheC-terminioftargetproteinsPHdomainsbindtodifferentphosphoinositidesFYVEdomainsspecificallybindtoPdtlns(3)P(phosphatidylinositol3-phosphate)MechanismsforActivationofSignalingProteinsbyRTKsActivationbymembranetranslocationActivationbyaconformationalchangeActivationbytyrosinephosphorylationMechanismsforAttenuation&TerminationofRTKActivation1)Ligandantagonists2)Receptorantagonists3)Phosphorylationanddephosphorylation4)Receptorendocytosis5)Receptordegradationbytheubiquitin-proteosomepathwayActivationofMAPKPathwaysbyMultipleSignalsGrowth,differentiation,inflammation,apoptosis->tumorigenesisOverviewofMAPKSignalingPathwaysTheMAPKPathwayActivatedbyRTKPRTKST-PI3KpathwayProto-oncogenesthatEncodeforSignallingProteinsSerine/Threonine

Kinases

c-raffamily

akt

Non-receptorTyrosineKinases

src

ablReceptorassociatedbindingproteins

c-rasfamilyRasrecruitsRaftothemembraneSTintermediatescanbetargetsforanti-cancerdrugsKinases:RafSTintermediatescanbetargetsforanti-cancerdrugsKinases:Bcr-AblCellPatterningCellGrowthWntBMPHedgehogFGFWhataretheessentialelementsofanySignalingcascade?Signal–ligandDiffusibleorTetheredReceptorTransmembrane

(exceptforlipidsolubleligands)Transducers-effectorsTargetsGenesorCellularcomponentsWntSignalingPathwaySignalWntsReceptorFrizzledsTransducers-effectors

b-cateninTargetsGenes

cytoskeletonWingless(Wg):Drosophila

?morphogen

-diff.Concentrationsofligandelicitdifferent responsesinequivalentcells ?morphogenicmovementsandcellfatedeterminants

?“Beposterior”-cellfate

?“divide”-proliferation

?developmentalabnormalitieswhengenedeletedTheSignal:WntSharmadescribesawinglessmutationin1973√Sharma,1973Wingless-anewmutantinD.melanogaster.D.I.S.50:134√SharmaandChopra,1976,Effectofthewingless(wg1)mutationonwingandhalteredevelopmentinDrosophilamelanogaster.Dev.Biol.48:461-465LateritwasclonedpositionallyIntegrationofMMTVcausesmammarytumorsinmiceTumorsarepregnancydependentMMTVhasasteroidenhancerMicedevelopbreasttumorsbutonlyduringlactationGenewasdesignated-Int-1(integrationofMMTV)OtherinsertionsitesoccurredatotherGFse.g.FGFTumorsexhibitdominantGainofFunctionLesson:Ectopicactivationofagene>hyperplasia=OncogeneWingless+int-1=WntFlywgandMouseInt-1arehomologsGenesarecloned.Sequenceissimilar

102030405060708090100HWnt-1MGLWALLPSWVSTTLLLALTALPAALAANS----SGR-----WWGIVNIASSTNLLTDSKSLQLVLEPSLQLLSR-KQRRLIRQNPGILHSVSGGLQSAVFlyWgMDISYIFVICLMALCSGGSSLSQVEGKQKSGRGRGSMWWGIAKVGEPNNITP-----IMYMDPAIHSTLRRKQRRLVRDNPGVLGALVKGANLAI110120130140150160170180190200HWnt-1RECKWQFRNRRWNCPT---APGPHLFGKIVNRGCRETAFIFAITSAGVTHSVARSCSEGSIESCTCDYRR--RGP----------GGPDWHWGGCSDNIDFlyWgSECQHQFRNRRWNCSTRNFSRGKNLFGKIVDRGCRETSFIYAITSAAVTHSIARACSEGTIESCTCDYSHQSRSPQANHQAGSVAGVRDWEWGGCSDNIG210220230240250260270280290300HWnt-1FGRLFGREFVDSGEKGRDLRFLMNLHNNEAGRTTVFSEMRQECKCHGMSGSCTVRTCWMRLPTLRAVGDVLRDRFDGASRVLYGN---------------FlyWgFGFKFSREFVDTGERGRNLREKMNLHNNEAGRAHVQAEMRQECKCHGMSGSCTVKTCWMRLANFRVIGDNLKARFDGATRVQVTNSLRATNALAPVSPNA310320330340350360370380390400HWnt-1RGSN----------------------------------------------------------RASR----------AELLRLEPEDPAHKPPSPHDLVYFFlyWgAGSNSVGSNGLIIPQSGLVYGEEEERMLNDHMPDILLENSHPISKIHHPNMPSPNSLPQAGQRGGRNGRRQGRKHNRYHFQLNPHNPEHKPPGSKDLVYL410420430440450460470HWnt-1EKSPNFCTYSGRLGTAGTAGRACNSSSPALDGCELLCCGRGHRTRTQRVTERCNCTFHWCCHVSCRNCTHTRVLHECLNFlyWgEPSPSFCEKNLRQGILGTHGRQCNETSLGVDGCGLMCCGRGYRRDEVVVVERCACTFHWCCEVKCKLCRTKKVIYTCLNTheSignal:Wnt?Secretedproteinligandsof80-100aa?Lipidmodified

Latestbreakthrough(2003):purificationofactiveWnt requiresorganicextraction!!?Shortrangeacting?Sticktoextracellularmatrix?Gradients---->morphogenic??multipleWnts

(19inhuman/mouseand7inDrosophila)Wntssignalthroughserpentinereceptors2classesofsignalingreceptorsCatalyticTyrosineKinaseReceptors[RTKs]Ser/Thr

KinaseReceptors[BMPs]Serpentine/G-protein-coupled-receptors(GPCRs)/7-transmembraneWnts?adrenergic,dopamine,epinepherineetcTheReceptor:Frizzled?corereceptorforWnts?seven-passtransmembraneproteins?probablyG-proteincoupledreceptors?multipleFrizzleds

(10inhuman/mouseand4inDrosophila)?anewlyidentifiedco-receptorforWnts?singlepasstransmembraneprotein?relatedtofamilyoflipoproteinreceptorsLRP/arrow:WntSignalingregulatesgeneexpressionandcellpolaritycanonicalWntFzWntFzLrpLrpnon-canonicalCanonicalWntsignalingin2005/~rnusse/pathways/cell2//cgi/cm/CMP_5533b-cateninisthecytoplasmic-nuclearsignalingmediatorb-cateninb-catenin?armadilloinDrosophila

geneticsdeterminedthatitfunctioneddownstreamofWg?b-catenininmammaliansystemidentifiedascomponent ofcelladhesionjunctions?subcellularlocalizationofproteincontroversialforyears?purificationofb-cateninandcloningofgenein1991byP.McCraeandB.Gumbinershowedthatarmadilloandb-cateninareorthologuesThetransducer/effector:Armadillorepeatstructureofb-cateninLEF/TCFWntCK1GSK-3bAPCb-cateninaxinFrizzledLRPE-cadherinWntsignalingpathwayC.Liuetal.2002.Cell108:837.Complicatedphosphorylationcontrolsb-cateninstabilityHowdoesb-cateninreachtargetgenes??LEF/TCFtranscriptionfactors?HMG(HighMobilityGroup)proteins?mammalianLEF-1andTCF-1identifiedinTlymphocytes in1991?twomoremembersclonedbylowstringencyscreeningofLibrariesanddegeneratePCRin1993?b-cateninwasusedinayeasttwo-hybridassayandLEF-1wasclonedasaninteractingproteinin1997 -endpointofthepathwaydetermined -mergedtwoindependentgroupsofscientists -subcellularlocalizationofb-catenin

finallysettledGeneralStructureofLEF/TCFTranscriptionFactorsb-catenin

bindingCo-repressorbindingDNAbinding/bendingalt.COOHTCF-1TCF-3TCF-4NLSHMGNLEF-1BBBEEE94%96%99%55%52%64%TargetGenesofWntSignaling?cellcycleregulatorsandtranscriptionfactors -c-MYC -cyclinD1?tissuespecificgenes?tissueremodelingproteins -matrixmetalloproteinases -ephrinreceptorsandligands -adhesionproteins?angiogenesis -VEGFIntheabsenceofWntsignaling:NLSHMGNLEF-1BNLSHMGNdnLEF-1BGrouchoLEF/TCFWntGSK-3baxinFrizzledLRPAPCLEF/TCFLEF/TCFLEF/TCFLEF/TCFLEF/TCFLEF/TCFLEF/TCFLEF/TCFActivationofLEF-1targetgenescantransformcellsAoki,M.etal.1999.Proc.Natl.Acad.Sci.USA.96:139-144anchorage-independentgrowthcontactinhibitioncolonyformationAdenomatous

PolyposisColi?IdentifiedbyJoannaGrodenandRayWhiteasatumorsuppressorgenesufferingLOHinfamilieswithveryhighratesofcoloncancer.?TruncationmutationsorlossoftheentiregeneoccursinmostSporadiccoloncancers?http:///~rnusse/wntwindow.htmHereditarycolorectalcancer(~15%)FamilialAdenomatous

Polyposis(FAP)-<1%allcolorectalCAGermlinemutationsinAPCgene.Acceleratedtumourinitiation.RatelimitingstepissomaticmutationinotherAPCalleleMedianageofcancerdiagnosis42yrs.Despitesharedgenotypes,notallclinicaldiseaseissimilar(diseasemodifyinggenesorenvironmentalinfluences?)Oftendevelopextracolonicmanifestations.MousemodelAPCminHereditaryNonPolyposisColorectalCancer(HNPCC)-2-4%allcolorectalCAMutationsinDNAmismatchrepair(MMR)genes-germline-genome-wide microsatelliteinstability(MI).Earlycluescamefrombacterialstudies.Adenomasformatthesamerateasthegeneralpopulation,butthereisacceleratedtumourprogressionMedianageofcancerdiagnosisalso42yrs.FAP HNPCC Sporadic Sporadic Adenomas CancersIncidence1:7000 1:500 1in2 1in20APCmutation>90% >80% >80%(prevalance) (germline) (somatic) (somatic)MMRdeficiency >90% <3% 13%(prevalance)MMRgene >70% ? ~65%mutationsAPCshuttlemode-speculativel

WntsignalingandcolorectalcancerMajorfunctionofAPCistheregulationofcelluar

b-cateninlevels.ActivationofwntpathwayincoloncancerdrivescellproliferationTcf-responsivegenes:

c-myc,cyclinD1,PPARd-fibronectinandmatrilysin(anextracellularmetalloproteinase)CNSMutationclusterregion-allresultinproteintruncationRacGEFGraybars-b-cateninbindingsites.APCmayplayaroleincell-celladhesion(Cadherins)Redbars-Axin/Conductinbindingsites(lostinmutations)Redarrows-nuclearexportsignals.MutantAPCaccumulatesinthenucleusAsefbindingactivatesRacatmembranes,inducingmembraneruffling thereforepossiblyaffectingcellmotilityMT-microtubulebindingsite.APCisinvolvedinlinkingmicrotubulestokinetochores thereforemutationscancontributetogenomicinstabilityb-catenindestructioncomplexAxinandAPCphysicallyinteract.APCmutationsincolonCAlackAxin bindingsites.-b-cateninbindstoAPC.APC/AxincomplexregulatesGSK3b

kinaseactivity.BindstoAxin. ThereforeAxinmayserveascaffoldingfunction.AxinandAPCarealsoGSK3bsubstrates,andphosphorylationincreases theirabilitytobindb-catenin.HowwntsignalsinhibitGSK3bactivityisunclear.Dishevellediscritical.WntsignalresultsindephosphorylationofAxinPP2Adephosphorylates

Axin.ItscatalyticsubunitbindsAxinwhileits regulatorysubunitbindsAPC.TheregulatorysubunitofPP2A ismutatedinasubsetofcolonCA.HowisPP2Aactivityregulated?-Whereistheintracellularlocalizationofthedestructioncomplex?APCmutationWildtypeAPCAPCmutationsresultinincreasedgenomicinstabilityMouseModel-APCminMultipleintestinalneoplasia(min).APCgenemutation.Truncatedproteinat

codon850.Htzhaveincreasedpropensityfortumors.Tumorsacquiresomaticmutation inwildtypeAPCallele.TumorslocatedinupperGItract(notcolorectal).Geneticbackgroundofmouseinfluencestumorload(?modifiers). MOM-1-possiblysecretedphospholipaseA2.APC1638TlacksC-terminaldomainthatbindstubulin,EB1-likeproteins. homozygousEScellshavehighdegreeofchromosomalinstability buthomozygousmicedoNOTexhibitincreasedtumorsusceptibilityCooperatingOncogenes.Cyclooxygenase2:deletionofCOX-2genesuppressesintestinalpolyposis inAPCD716mice.COX-2levelsareincreasedinpremalignantpolyps. ButCOX-2isexpressedininterstitialcellsnotintestinalepithelium.Smad4:deletionofSmad4inAPCD716miceresultedinmoreaggressive tumors(compoundhtzmice). HighlightstheroleofTGFsignalintumorprogression.DNAmethyltransferase:compoundhtzhavereducedpolypnumbers (epigeneticevents?).TumourProgressionTGFsignalingmutationsreceptorIImutationsdetectedinregionsofhighgradedysplasia

butabsentinadenomas.Intumourswithmicrosatelliteinstability(MI)mutationscorrelatewithprogressionofadenomastocancermutationsinTGFsignalingcomponents(e.g.,smad4)-nonMItumor accelerate/worsenmurine(APCmin)intestinalcancermodelCell-celladhesivecomplexmutations-cadherins,b-catenins,others?3.Metalloproteinaseactivation-matrilysinisatcf-responsivegenecompactionoftheearlyembryo-morphogeneticmovementofcells-establishmentofcellfates,andpolaritylossofcell-cellandcell-matrixrecognitiontissueinvasionmotilitynormaldevelopmentcancerprogression“epithelialmesenchymal”transitionHedgehogSignalingPathwaySignalHedgehogReceptorPatchedTransducers-effectors

Cubitus

InterruptusTargetsGenes

?MutationsinHedgehogsignalinginhumansembryosyieldscyclopia (aformofholoprosencephaly)imagesareonlyforthestout-hearted.?Inadults,mutationsinHedgehogsignalinggivesphenotypesinstemcellandprogenitorpopulations.Increasedsignalinggivestumors,lesssignalinggivesshort-livedstemcells?MostrecentadvanceisthatmanytumorsshowelevatedHhsignaling.Cyclopamine(firstobtainedfromthecornlily)haspromisefortherapeuticinterventionofcancer.TheSignal:Hedgehog?Secretedproteinligand-heavilyprocessed?Lipidmodifiedwithcholesterol?Shortrangeacting?Sticktoextracellularmatrix?Gradients---->morphogenic??threeligandsinmammals:Indian,Desert,SonicThehedgehoggeneencodesanovelmembrane-linkedligandimportantforlocalpatterningofmanytissues.Theprimarytranslationproductcontainsasignalpeptidethatiscleavedtoproducea45kDapolypeptideprecursor.Cleavageofthissecretedprecursorproducesa20kDaN-terminalfragmentassociatedwiththeplasmamembraneandwithinductiveactivityplusa25kDafragment.TheN-terminalfragmentbecomestetheredtothemembraneviaahydrophobiccholesterolmoiety(itdoesn’tcontainanyhydrophobicresidues). Aseries