肿瘤细胞信号转导

如何阅读文献在肿瘤学方面有哪些好的杂志如何快速阅读文献以获取知识论文的主要结构肿瘤的分子信号转导

+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-transmembraneWntsßadrenergic,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