腫瘤細胞信號轉導課件

1、如何閱讀文獻在腫瘤學方面有哪些好的雜志如何快速閱讀文獻以獲取知識論文的主要結構如何閱讀文獻在腫瘤學方面有哪些好的雜志腫瘤細胞信號轉導課件腫瘤細胞信號轉導課件腫瘤的分子信號轉導腫瘤的分子信號轉導 + genomic instabilityfrom Hanahan and Weinberg 2000 + genomic from Hanahan and WeSignal Transduction and CancerLecture I: Growth Factors and ReceptorsOutline:What is Signal Transduction?What are Growth F

2、actors?How do they contribute to normal ST?How is this ST deregulated in Cancer?Signal Transduction and CancerLecture I: Growth Factors and ReceptorsWhat is Signal Transduction?Signal Transduction is the process by which a cell converts an extracellular signal into a response.Involved in:Cell-cell c

3、ommunicationCells response to environmentIntracellular homeostatsis- internal communication Lecture I: Growth Factors and Generic Signalling PathwaySignalReceptor (sensor)Transduction CascadeTargetsResponse Altered MetabolismMetabolicEnzymeGene RegulatorCytoskeletal ProteinAltered Gene ExpressionAlt

4、ered Cell Shape or MotilityAdapted from Molecular Biology of the Cell,(2002), 4th edition, Alberts et al. Generic Signalling PathwaySignComponents of SignallingWhat can be the Signal?External message to the cellPeptides / Proteins- Growth FactorsAmino acid derivatives - epinephrine, histamineOther s

5、mall biomolecules - ATPSteroids, prostaglandinsGases - Nitric Oxide (NO)PhotonsDamaged DNAOdorants, tastantsSignal = LIGANDLigand- A molecule that binds to a specific site on another molecule, usually a protein, ie receptorComponents of SignallingWhat cComponents of SignallingWhat are Receptors?Sens

6、ors, what the signal/ligand binds to initiate ST Cell surface Intracellular Hydrophillic LigandCell-Surface ReceptorPlasma membraneHydrophobic LigandCarrier ProteinIntracellularReceptorNucleusAdapted from Molecular Biology of the Cell,(2002), 4th edition, Alberts et al. Components of SignallingWhat

7、aCell Surface Receptor Types:Ligand-gated ion channelCell Surface Receptor Types:Cell Surface Receptor Types:2) G-Protein Coupled ReceptorCell Surface Receptor Types:Cell Surface Receptor Types:3) Enzyme-linked Receptor eg Growth Factor ReceptorsCell Surface Receptor Types:Growth FactorsLigands whic

8、h bind enzyme linked receptorsSignal diverse cellular responses including:ProliferationDifferentiationGrowthSurvival AngiogenesisCan signal to multiple cell types or be specificGrowth FactorsLigands which biFactorPrincipal SourcePrimary ActivityCommentsPDGFplatelets, endothelial cells, placentapromo

9、tes proliferation of connective tissue, glial and smooth muscle cellstwo different protein chains form 3 distinct dimer forms; AA, AB and BBEGFsubmaxillary gland, Brunners glandpromotes proliferation of mesenchymal, glial and epithelial cellsTGF-common in transformed cellsmay be important for normal

10、 wound healingrelated to EGFFGFwide range of cells; protein is associated with the ECMpromotes proliferation of many cells; inhibits some stem cells; induces mesoderm to form in early embryosat least 19 family members, 4 distinct receptorsNGFpromotes neurite outgrowth and neural cell survivalseveral

11、 related proteins first identified as proto-oncogenes; trkA (trackA), trkB, trkCErythropoietinkidneypromotes proliferation and differentiation of erythrocytesTGF-activated TH1 cells (T-helper) and natural killer (NK) cellsanti-inflammatory (suppresses cytokine production and class II MHC expression)

12、, promotes wound healing, inhibits macrophage and lymphocyte proliferationat least 100 different family membersIGF-Iprimarily liverpromotes proliferation of many cell typesrelated to IGF-II and proinsulin, also called Somatomedin CIGF-IIvariety of cellspromotes proliferation of many cell types prima

13、rily of fetal originrelated to IGF-I and proinsulinFactorPrincipal SourcePrimary Growth Factor ReceptorsGrowth Factor ReceptorsMost growth factors bind Receptor Tyrosine KinasesMost growth factors bind Recep腫瘤細胞信號轉導課件Growth Factor Receptor Activation IRTKRS/TKGrowth Factor Receptor ActivatGrowth Fac

14、tor Receptor Activation IIGrowth Factor Receptor ActivatGrowth signal autonomy,Insensitivity to anti-growth signals,Resistance to apoptosis:Uncouple cells growth program from signals in the environment.Growth factors in normal cells serveas environmental signals. Growth Factor ST and CancerGrowth fa

15、ctors regulate growth, proliferation, and survival.These are all deregulated in cancer.Hanahan and Weinberg, (2000) Hallmarks of Cancer, Cell (100) 57Growth signal autonomy,Growth Growth factors with Oncogenic PotentialPDGF, originally shown to regulate proliferation, was also shown to have homology

16、 to v-sis, the simian sarcoma virus. Other viral oncogenes encoded protein products that were growth factors that often overexpressed in cancer such as TGF-a. Autocrine signalling leads to deregulated growth. PDGF familyNeurotrophinsA chainNGFB chain (c-sis)BDNFFGF FamilyNT3acidic FGFCytokines (Hema

17、topoietic)basic FGFIL-2EGF Family IL-3EGFM-CSFTGF-aGM-CSFGrowth factors with Oncogenic GF Receptors with Oncogenic PotentialEGFR, kinase activity stimulated by EGF-1 and TGF-a involved incell growth and differentiation, was linked via sequence homology to a known avian erythroblastosis virus onocgen

18、e, v-erbB. Since then, many oncogenes have been shown to encode for GFRs.EGFR familyInsulin Receptor familyerbB1 (c-erbB)IGF-1 (c-ros)erbB2 (neu) Neurotrophins FGF FamilyNGFR (trk) FGFR-1(fig)BDNFR (trk-B)FGFR-2(K-sam)NT3 R (trk-C)PDGFR FamilyCSF-1R (c-fms)SLF R (c-kit)GF Receptors with Oncogenic Po

19、腫瘤細胞信號轉導課件腫瘤細胞信號轉導課件腫瘤細胞信號轉導課件腫瘤細胞信號轉導課件Induction of cancer by alternations in several types of proteins involved in cell growth control Induction of cancer by alternaSignal Transduction and CancerLecture II: Intracellular SignallingOutline:What are some signalling pathways?What are their cell biolo

20、gical outputs?How do these result in the cancer phenotype?How can we exploit signalling pathways for therapy?Signal Transduction and CancerGeneric Signal TransductionGeneric Signal TransductionRTK Signal TransductionRTK Signal TransductionSignal TransductionDownstream effectorsProtein Signaling Modu

21、les (Domains)SH2 and PTB bind to tyrosine phosphorylated sitesSH3 and WW bind to proline-rich sequencesPDZ domains bind to hydrophobic residues at the C-termini of target proteinsPH domains bind to different phosphoinositidesFYVE domains specifically bind to Pdtlns(3)P (phosphatidylinositol 3-phosph

22、ate)Signal TransductionProtein SigMechanisms for Activation of Signaling Proteins by RTKsActivation by membrane translocationActivation by a conformational changeActivation by tyrosine phosphorylationMechanisms for Activation of SMechanisms for Attenuation & Termination of RTK Activation1) Ligand an

23、tagonists2) Receptor antagonists3) Phosphorylation and dephosphorylation4) Receptor endocytosis5) Receptor degradation by the ubiquitin-proteosome pathwayMechanisms for Attenuation & TActivation of MAPK Pathways by Multiple SignalsGrowth, differentiation, inflammation, apoptosis - tumorigenesisActiv

24、ation of MAPK Pathways byOverview of MAPK Signaling PathwaysOverview of MAPK Signaling PatThe MAPK Pathway Activated by RTKThe MAPK Pathway Activated by PRTK ST- PI3K pathwayPRTK ST- PI3K pathway腫瘤細胞信號轉導課件腫瘤細胞信號轉導課件Proto-oncogenes that Encode for Signalling ProteinsSerine/Threonine Kinasesc-raf fami

25、lyaktNon-receptor Tyrosine KinasessrcablReceptor associated binding proteinsc-ras familyProto-oncogenes that Encode fo腫瘤細胞信號轉導課件腫瘤細胞信號轉導課件腫瘤細胞信號轉導課件Ras recruits Raf to the membraneRas recruits Raf to the membraST intermediates can be targets for anti-cancer drugsKinases:RafST intermediates can be ta

26、rgetST intermediates can be targets for anti-cancer drugsKinases:Bcr-AblST intermediates can be target腫瘤細胞信號轉導課件Cell PatterningCell GrowthWntBMPHedgehogFGFCell PatterningWntWhat are the essential elements of any Signaling cascade?Signal ligandDiffusible or TetheredReceptor Transmembrane (except for

27、lipid soluble ligands)Transducers - effectorsTargets Genes or Cellular componentsWhat are the essential elementWnt Signaling PathwaySignalWntsReceptor FrizzledsTransducers - effectorsb-cateninTargets Genes cytoskeletonWnt Signaling PathwaySignalWingless (Wg): Drosophila morphogen - diff. Concentrati

28、ons of ligand elicit differentresponses in equivalent cells morphogenic movements and cell fate determinants “Be posterior” - cell fate “divide” - proliferation developmental abnormalities when gene deletedThe Signal: WntWingless (Wg): DrosophilaThe Sharma describes a wingless mutation in 1973 Sharm

29、a, 1973 Wingless - a new mutant in D. melanogaster. D. I. S. 50: 134 Sharma and Chopra, 1976, Effect of the wingless (wg1) mutation on wing and haltere development in Drosophila melanogaster. Dev. Biol. 48: 461-465Later it was cloned positionallySharma describes a wingless muIntegration of MMTV caus

30、es mammary tumors in miceTumors are pregnancy dependentMMTV has a steroid enhancerMice develop breast tumors but only during lactationGene was designated - Int-1 (integration of MMTV)Other insertion sites occurred at other GFs e.g. FGFTumors exhibit dominant Gain of FunctionLesson:Ectopic activation

31、 of a gene hyperplasia= OncogeneIntegration of MMTV causes mamWingless + int-1 = WntFly wg and Mouse Int-1 are homologsGenes are cloned. Sequence is similar 10 20 30 40 50 60 70 80 90 100H Wnt-1 MGLWALLPSWVSTTLLLALTALPAALAANS-SGR-WWGIVNIASSTNLLTDSKSLQLVLEPSLQLLSR-KQRRLIRQNPGILHSVSGGLQSAVFly Wg MDISY

32、IFVICLMALCSGGSSLSQVEGKQKSGRGRGSMWWGIAKVGEPNNITP-IMYMDPAIHSTLRRKQRRLVRDNPGVLGALVKGANLAI 110 120 130 140 150 160 170 180 190 200H Wnt-1 RECKWQFRNRRWNCPT-APGPHLFGKIVNRGCRETAFIFAITSAGVTHSVARSCSEGSIESCTCDYRR-RGP-GGPDWHWGGCSDNIDFly Wg SECQHQFRNRRWNCSTRNFSRGKNLFGKIVDRGCRETSFIYAITSAAVTHSIARACSEGTIESCTCDYSHQ

33、SRSPQANHQAGSVAGVRDWEWGGCSDNIG 210 220 230 240 250 260 270 280 290 300H Wnt-1 FGRLFGREFVDSGEKGRDLRFLMNLHNNEAGRTTVFSEMRQECKCHGMSGSCTVRTCWMRLPTLRAVGDVLRDRFDGASRVLYGN-Fly Wg FGFKFSREFVDTGERGRNLREKMNLHNNEAGRAHVQAEMRQECKCHGMSGSCTVKTCWMRLANFRVIGDNLKARFDGATRVQVTNSLRATNALAPVSPNA 310 320 330 340 350 360 370 3

34、80 390 400H Wnt-1 RGSN-RASR-AELLRLEPEDPAHKPPSPHDLVYFFly Wg AGSNSVGSNGLIIPQSGLVYGEEEERMLNDHMPDILLENSHPISKIHHPNMPSPNSLPQAGQRGGRNGRRQGRKHNRYHFQLNPHNPEHKPPGSKDLVYL 410 420 430 440 450 460 470 H Wnt-1 EKSPNFCTYSGRLGTAGTAGRACNSSSPALDGCELLCCGRGHRTRTQRVTERCNCTFHWCCHVSCRNCTHTRVLHECLNFly Wg EPSPSFCEKNLRQGILGT

35、HGRQCNETSLGVDGCGLMCCGRGYRRDEVVVVERCACTFHWCCEVKCKLCRTKKVIYTCLNWingless + int-1 = WntFly wg aThe Signal: Wnt Secreted protein ligands of 80-100aa Lipid modifiedLatest breakthrough (2003): purification of active Wntrequires organic extraction! Short range acting Stick to extracellular matrix Gradients

36、- morphogenic? multiple Wnts (19 in human/mouse and 7 in Drosophila)The Signal: Wnt Secreted prWnts signal through serpentine receptors2 classes of signaling receptorsCatalyticTyrosine Kinase Receptors RTKsSer/Thr Kinase Receptors BMPsSerpentine/G-protein-coupled-receptors(GPCRs) /7-transmembraneWnt

37、sadrenergic, dopamine, epinepherine etcWnts signal through serpentineThe Receptor: Frizzled core receptor for Wnts seven-pass transmembrane proteins probably G-protein coupled receptors multiple Frizzleds (10 in human/mouse and 4 in Drosophila) a newly identified co-receptor for Wnts single pass tra

38、nsmembrane protein related to family of lipoprotein receptorsLRP/arrow:The Receptor: Frizzled core Wnt Signaling regulates gene expression and cell polaritycanonicalWntFzWntFzLrpLrpnon-canonicalWnt Signaling regulates gene eCanonical Wnt signaling in 2005/rnusse/pathways/cell2/cgi/cm/CMP_5533Canonic

39、al Wnt signaling in 200b-catenin is the cytoplasmic-nuclear signaling mediatorb-cateninb-catenin is the cytoplasmic-nb-catenin armadillo in Drosophilagenetics determined that it functioned downstream of Wg b-catenin in mammalian system identified as componentof cell adhesion junctions subcellular lo

40、calization of protein controversial for years purification of b -catenin and cloning of gene in 1991 by P. McCrae and B. Gumbiner showed that armadillo and b-catenin are orthologuesThe transducer/effector:b-catenin armadillo in DrosoArmadillo repeat structure of b-cateninArmadillo repeat structure o

41、f LEF/TCFWntCK1GSK-3bAPCb-cateninaxinFrizzledLRPE-cadherinWnt signaling pathwayLEF/TCFWntCK1GSK-3bAPCb-cateniC. Liu et al. 2002. Cell 108:837.Complicated phosphorylation controls b-catenin stabilityC. Liu et al. 2002. Cell 108:8How does b-catenin reach target genes? LEF/TCF transcription factors HMG

42、 (High Mobility Group) proteins mammalian LEF-1 and TCF-1 identified in T lymphocytesin 1991 two more members cloned by low stringency screening ofLibraries and degenerate PCR in 1993 b-catenin was used in a yeast two-hybrid assay andLEF-1 was cloned as an interacting protein in 1997- endpoint of th

43、e pathway determined- merged two independent groups of scientists- subcellular localization of b-catenin finally settledHow does b-catenin reach targeGeneral Structure of LEF/TCF Transcription Factorsb-catenin bindingCo-repressor bindingDNA binding/bendingalt.COOHTCF-1TCF-3TCF-4NLSHMGNLEF-1BBBEEE94%

44、96%99%55%52%64%General Structure of LEF/TCF TTarget Genes of Wnt Signaling cell cycle regulators and transcription factors-c-MYC-cyclin D1 tissue specific genes tissue remodeling proteins- matrix metalloproteinases- ephrin receptors and ligands- adhesion proteins angiogenesis- VEGFTarget Genes of Wn

45、t SignalingIn the absence of Wnt signaling:NLSHMGNLEF-1BNLSHMGNdnLEF-1BGrouchoIn the absence of Wnt signalinLEF/TCFWntGSK-3baxinFrizzledLRPAPCLEF/TCFLEF/TCFLEF/TCFLEF/TCFLEF/TCFLEF/TCFLEF/TCFLEF/TCFLEF/TCFWntGSK-3baxinFrizzledLRActivation of LEF-1 target genes can transform cellsAoki, M. et al. 1999

46、. Proc. Natl. Acad. Sci. USA. 96:139-144anchorage-independent growth contact inhibition colony formationActivation of LEF-1 target genAdenomatous Polyposis Coli Identified by Joanna Groden and Ray White as a tumor suppressorgene suffering LOH in families with very high rates of colon cancer. Truncat

47、ion mutations or loss of the entire gene occurs in mostSporadic colon cancers /rnusse/wntwindow.htmAdenomatous Polyposis ColihttpHereditary colorectal cancer ( 15%)Familial Adenomatous Polyposis (FAP) - 90% 80% 80% (prevalance) (germline)(somatic)(somatic)MMR deficiency 90% 70% ? 65% mutations FAP H

48、NPCC SporadAPC shuttle mode - speculativel Wnt signaling and colorectal cancerMajor function of APC is the regulation of celluar b-catenin levels.Activation of wnt pathway in colon cancer drives cell proliferationTcf-responsive genes: c-myc, cyclin D1, PPARd- fibronectin and matrilysin (an extracell

49、ular metalloproteinase)APC shuttle modeWnt signaling CNSMutation cluster region - all result in protein truncationRac GEFGray bars - b-catenin binding sites. APC may play a role in cell-cell adhesion (Cadherins)Red bars - Axin/Conductin binding sites (lost in mutations)Red arrows - nuclear export si

50、gnals. Mutant APC accumulates in the nucleusAsef binding activates Rac at membranes, inducing membrane rufflingtherefore possibly affecting cell motilityMT - microtubule binding site. APC is involved in linking microtubules to kinetochorestherefore mutations can contribute to genomic instabilityCNSM

51、utation cluster region - ab-catenin destruction complexAxin and APC physically interact. APC mutations in colon CA lack Axinbinding sites.- b-catenin binds to APC.APC/Axin complex regulates GSK3b kinase activity. Binds to Axin.Therefore Axin may serve a scaffolding function.Axin and APC are also GSK

52、3b substrates, and phosphorylation increasestheir ability to bind b-catenin.How wnt signals inhibit GSK3b activity is unclear. Dishevelled is critical.Wnt signal results in dephosphorylation of AxinPP2A dephosphorylates Axin. Its catalytic subunit binds Axin while itsregulatory subunit binds APC. Th

53、e regulatory subunit of PP2Ais mutated in a subset of colon CA. How is PP2A activity regulated?- Where is the intracellular localization of the destruction complex?b-catenin destruction complexA腫瘤細胞信號轉導課件APC mutationWild type APCAPC mutations result in increased genomic instabilityAPC mutationWild t

54、ype APCAPC mMouse Model - APCminMultiple intestinal neoplasia (min). APC gene mutation. Truncated protein atcodon 850.Htz have increased propensity for tumors. Tumors acquire somatic mutation in wild type APC allele.Tumors located in upper GI tract (not colorectal).Genetic background of mouse influe

55、nces tumor load (?modifiers).MOM-1 - possibly secreted phospholipase A2.APC1638T lacks C-terminal domain that binds tubulin, EB1-like proteins.homozygous ES cells have high degree of chromosomal instabilitybut homozygous mice do NOT exhibit increased tumor susceptibilityCooperating Oncogenes.Cycloox

56、ygenase 2: deletion of COX-2 gene suppresses intestinal polyposisin APCD716 mice. COX-2 levels are increased in premalignant polyps.But COX-2 is expressed in interstitial cells not intestinal epithelium.Smad4: deletion of Smad4 in APCD716 mice resulted in more aggressivetumors (compound htz mice). H

57、ighlights the role of TGF signal in tumor progression.DNA methyltransferase: compound htz have reduced polyp numbers(epigenetic events?).Mouse Model - APCminMultiple iTumour ProgressionTGF signaling mutationsreceptor II mutations detected in regions of high grade dysplasia but absent in adenomas. In

58、 tumours with microsatellite instability (MI)mutations correlate with progression of adenomas to cancermutations in TGF signaling components (e.g., smad4) - non MI tumoraccelerate/worsen murine (APCmin) intestinal cancer modelCell-cell adhesive complex mutations - cadherins, b-catenins, others?3. Me

59、talloproteinase activation - matrilysin is a tcf-responsive geneTumour ProgressionTGF signa compaction of the early embryo- morphogeneticmovement of cells- establishment ofcell fates, and polarityloss of cell-cell and cell-matrix recognitiontissue invasion motilitynormal developmentcancer progressio

60、n“epithelial mesenchymal” transition compaction of the - morphogenHedgehog Signaling PathwaySignalHedgehogReceptor PatchedTransducers - effectorsCubitus InterruptusTargets Genes Hedgehog Signaling PathwaySign Mutations in Hedgehog signaling in humans embryos yields cyclopia(a form of holoprosencepha