腫瘤學(xué)-信號(hào)傳導(dǎo)與疾病課件

1、信號(hào)傳導(dǎo)與疾?。⊿ignal transduction and disease）The Hallmarks of CancerCell-to-cell communication by extracellular signaling usually involves six stepssynthesis of signaling molecule by signaling cellrelease of the signaling molecule by the signaling celltransport of the signal to the target cellChanges in

2、targeted gene expression triggered by the receptor-signal complexremoval of the signal, which usually terminates the cellular responsedetection of the signal by a specific receptor protein細(xì)胞外因子通過(guò)與受體（膜受體或核受體）結(jié)合，引發(fā)細(xì)胞內(nèi)的一系列生物化學(xué)反應(yīng)，直至細(xì)胞生理反應(yīng)所需基因的轉(zhuǎn)錄表達(dá)開(kāi)始的過(guò)程。“wiring” of cell regulatory circuitsIntercellular S

3、ignaling:how cells communicate with one anotherIntracellularSignalingExamples of SignalingSYNAPTICPARACRINEENDOCRINEExtracellular Signals Bind to Different Types of Receptors 1951年，第一個(gè)生長(zhǎng)因子神經(jīng)生長(zhǎng)因子 NGF發(fā)現(xiàn) Rita Levi-Montalcini & Stanley Cohen 1955年，Receptor 的概念正式提出 1957年，第二信使cAMP發(fā)現(xiàn) Earl Sutherland50 年代中期

4、，磷酸化的發(fā)現(xiàn)和第一個(gè)蛋白激酶的分離 Edwin Krebs & Fischer 70年代，G蛋白和G蛋白連接受體的發(fā)現(xiàn)和克隆 Alfred G. Gilman & Martin Rodbell 發(fā)現(xiàn)癌基因v-src 的蛋白產(chǎn)物是蛋白酪氨酸激酶 Michael Bishop & Harold E. Varmus簡(jiǎn)要回顧The Nobel Prize in Physiology or Medicine 1986for their discoveries of growth factors Stanley Cohen Rita Levi-Montalcini “for his discoverie

5、s concerning the mechanisms of the action of hormones” The Nobel Prize in Physiology or Medicine 1971Earl W. Sutherland, Jr. The Nobel Prize in Physiology or Medicine 1994Alfred G. Gilman Martin Rodbell for their discovery of G-proteins and the role of these proteins in signal transduction in cells

6、for their discoveries concerning reversible protein phosphorylation as a biological regulatory mechanism The Nobel Prize in Physiology or Medicine 1992Edmond H. Fischer Edwin G. Krebs The Nobel Prize in Physiology or Medicine 1989for their discovery of the cellular origin of retroviral oncogenes J.

7、Michael Bishop Harold E. Varmus Robert F. Furchgott Louis J. Ignarro Ferid Murad “for their discoveries concerning nitric oxide as a signalling molecule in the cardiovascular system The Nobel Prize in Physiology or Medicine 1999 細(xì)胞外因子受體聯(lián)結(jié)蛋白G蛋白第二信使胞內(nèi)激酶核受體基本組成刺激細(xì)胞增殖的因子生長(zhǎng)因子（growth factor） 是一類以刺激細(xì)胞生長(zhǎng)為特征

8、的多肽 受體具有酪氨酸激酶活性 特異性 多樣性 家族性 交叉性細(xì)胞因子（cytokine） 白介素、 血細(xì)胞刺激因子、干擾素 受體本身不具有激酶活性激素和神經(jīng)遞質(zhì)細(xì)胞外因子其他重要的細(xì)胞外因子抗原腫瘤壞死因子（tumor necrosis factor, TNF）粘附分子(adhesion molecules) 纖粘連蛋白（Fibronectine, Fn ）層粘連蛋白（laminin, Lm） 膠原蛋白（collagen, coll）細(xì)胞外因子受體 酪氨酸激酶受體（ Receptor Tyrosine Kinases, RTK） G蛋白連接受體 （G protein-Coupled Rece

9、ptors) 細(xì)胞因子受體 （Cytokine Receptors） 粘附因子受體 （Adhesion Molecules Receptors）酪氨酸激酶受體Tyrosine-kinase ReceptorsClassExamplesStructural Features of ClassIEGF receptor, NEU/HER2, HER3cysteine-rich sequencesIIinsulin receptor, IGF-1 receptorcysteine-rich sequences; characterized by disulfide-linked heterotetr

10、amersIIIPDGF receptors, c-Kitcontain 5 immunoglobulin-like domains; contain the kinase insertIVFGF receptorscontain 3 immunoglobulin-like domains as well as the kinase insert; acidic domainVvascular endothelial cell growth factor (VEGF) receptorcontain 7 immunoglobulin-like domains as well as the ki

11、nase insert domainVIhepatocyte growth factor (HGF) and scatter factor (SC) receptorsheterodimeric like the class II receptors except that one of the two protein subunits is completely extracellular. The HGF receptor is a proto-oncogene that was originally identified as the Met oncogeneVIIneurotrophi

12、n receptor family (trkA, trkB, trkC) and NGF receptorcontain no or few cysteine-rich domains; NGFR has leucine rich domainG蛋白連接受體 G-protein Coupled ReceptorsoutsideinsideG protein-coupled receptorsSchematic diagram of the general structure of G protein linked receptors. All receptors of this type co

13、ntain seven transmembrane a-helical regions. The loop between a helices 5 and 6, and in some cases the loop between helices 3 and 4, which face the cytosol, are important for interactions with the coupled G protein. olfactory R, adenosine R, cannabinoid R, .Family AFamily BGPCRFamily CFamily DFamily

14、 EFamily FGroup IGroup IIGroup IIIGroup VIGroup IVGroup VGroup IGroup IIGroup IIIGroup IIGroup IVGroup IIIGroup Iserotonin R, adrenergic R, dopamine R,histamine R, muscarinic R, .rhodopsins, endothelin R, thyrotropinreleasing hormone R, .bradykinin R, .angiotensin R, chemokine R, thrombin R, .melato

15、nin R, .calcitonin R, .parathyroid hormone/parathyroid-related-peptide Rglucagon R, growth hormone releasinghormone R, pituitary adenylyl cyclaseactivating peptide R, .latrotoxin R, .metabotropic glutamate R, .calcium R, .GABA-B R, .STE2 R, .STE3 R, .cAMP R, .bradykinin R, .G Protein-coupled Recepto

16、rsG Proteinai3asaolfai1ai2aoAaoBat1at2agazaqa11a14a15a16a12a136080100% amino acid identityasaiaqa12G Protein a SubunitsGs: linking -adrenergic receptors and adenylyl cyclase Gi: Inhibiting Certain Isotypes of Adenylyl CyclasesEach cytokine/hematopoietic receptor is a composite of structurally distin

17、ct protein domains. Members of this receptor family contain one, two, or even three polypeptides. The transmembrane domain consists of a single alpha helix of about 20 amino acids. Ligand binding occurs at a domain near the transmembrane domain 細(xì)胞因子受體 Cytokine ReceptorsLigand binding occurs in a reg

18、ion near the transmembrane domain.The cytosolic domain of a cytokine receptor does not possess intrinsic catalytic activity. Rather, the ligand-bound receptor recruits and activates the cytosolic tyrosine kinase JAK. JAK substrates include JAK itself, the receptor, IRS molecules, and the transcripti

19、onal activator STAT. Phosphorylated JAK also interacts with other signal transduction proteins through SH2 domains粘附因子受體Adhesion Molecules ReceptorscadherinsintegrinsIg超家族selectin連接蛋白Protein Binding Domain related to signal transduction SH2 domain SH3 domain PH domain Death Domain Src, is the produc

20、t of the first proto-oncogene to be characterized. Src, is a non-receptor tyrosine kinaseOther proteins have homologies to Src domainsSH Src-homology regions SH2 and SH3 domains: mediate protein-protein interactions in cellular signaling cascades: very common in proteins outside the Src family. SH2:

21、 binds peptides with consensus: (PTyr-Met/Val-X-Met) SH3: b-barrel. Interacts with proline-rich peptide targets The Src homology 2 (SH2) domain has been found in a number of signal transduction pathways. Its primary function is to bind phosphotyrosines and in doing so localizing different proteins n

22、ecessary to transmitt the proper function.SH2 DomainsPleckstrin homology domain (PH)Structural overview of DH-PH complexDHPHThe PH domain is able to interact with the DH domainPHSH2SH3pTyrproteinproteinX-P-P-X-PMembraneFunction of AdaptorG蛋白 經(jīng)典G蛋 小分子量G蛋白TYPES OF G-PROTEINSRAS proteins are small G-pr

23、oteinsCell proliferationMembrane boundGTP-dependent switchesActivate MAP kinasesSmall GTP-binding proteins include (roles indicated):initiation & elongation factors (protein synthesis).Ras (growth factor signal cascades).Rab (vesicle targeting and fusion).Ran (transport of proteins into & out of the

24、 nucleus).Rho (regulation of actin cytoskeleton)All GTP-binding proteins differ in conformation depending on whether GDP or GTP is present at their nucleotide binding site. Generally, GTP binding induces the active state.第二信使胞內(nèi)激酶 MAPK pathway (Mitogen-activated protein kinase pathway)Erk (extracellu

25、lar-signal regulated kinase):p44/p22JNK (c-Jun N-terminal kinase); SAPK (stress-activated protein kinase)p38 cAMP-PKA pathway cGMP-PKG pathway DAG-PKC pathway PI-3K pathway Ca2+-CAMK pathway MAP kinase pathwayThe MAP kinase CascadeMAP-kinaseMAP-kinase-kinaseMAP-kinase-kinase-kinasechanges in protein

26、 activitychanges in gene expression核受體INACTIVE RECEPTORACTIVE RECEPTOR核受體Tyrosine Kinase Receptors signal pathwayG protein-coupled Receptors pathwayTGF signal pathwayTNF signal pathwayWnt signal pathway主要信號(hào)傳導(dǎo)通路Mechanism of Tyrosine Kinase Receptors When hormone binds to the extracellular domain the

27、receptors aggregate Mechanism of Tyrosine Kinase Receptors When the receptors aggregate, the tyrosine kinase domains phosphorylate the C terminal tyrosine residues Mechanism of Tyrosine Kinase Receptors This phosphorylation produces binding sites for proteins with SH2 domains. GRB2 is one of these p

28、roteins. GRB2, with SOS bound to it, then binds to the receptor complex. This causes the activation of SOS. Mechanism of Tyrosine Kinase Receptors SOS is a guanyl nucleotide-release protein (GNRP). When this is activated, it causes certain G proteins to release GDP and exchange it for GTP. Ras is on

29、e of these proteins. When ras has GTP bound to it, it becomes active. Mechanism of Tyrosine Kinase Receptors Activated ras then causes the activation of a cellular kinase called raf-1 Mechanism of Tyrosine Kinase Receptors Raf-1 kinase then phosphorylates another cellular kinase called MEK. This cau

30、se the activation of MEK Mechanism of Tyrosine Kinase Receptors Activated MEK then phosphorylates another protein kinase called MAPK causing its activation. This series of phosphylating activations is called a kinase cascade. It results in amplification of the signal Mechanism of Tyrosine Kinase Rec

31、eptors Among the final targets of the kinase cascade are transcriptions factors (fos and jun showed here). Phosphorylation of these proteins causes them to become active and bind to the DNA, causing changes in gene transcription Signal Transduction Through Receptor Tyrosine Kinases The G Protein Act

32、ivation/Inactivation Cycle cAMP PathwayIP3 PathwaySignaling moleculeCell surface receptorG proteinEnzymeTarget proteinIntracellular mediatorcAMPCa+Canonical, SMAD-dependent TGF- signaling pathwayNon-canonical TGF- signaling and crosstalk with other signaling pathwaysDEATH signals TNF-alphaCD95 ligan

33、d / Fas ligandDeath receptors, e.g. CD95Death domainsBinding to the receptor induces receptors to cluster and trimerize After binding to ligand, receptors form trimersDeath domainsFADD (Fas associated death domain protein) is recruited via its death domain DED (death effector domain) of FADD recruit

34、s pro-caspase 8 via its DED FADD-CASP8 complex brings multiple pro-caspase 8 molecules in close proximity(induced proximity aggregation results in their cross-activation). This is the DISC -complex. DISC complexActivated caspase-8 (a heterotetramer) is released from DISC into the cytoplasm caspase-8

35、 cleaves pro-apoptotic BID protein.BID interacts with the pro-apoptotic BCL2 relatives BAX and BAKThis amplifies apoptosis induction through the cell-extrinsic pathway Nature Reviews Cancer 2; 420-430 (2002); TARGETING DEATH AND DECOY RECEPTORS OF THE TUMOUR-NECROSIS FACTOR SUPERFAMILYTNF- and NF-Bc

36、ell surface TNF- receptorNF-B inactive in cytoplasm with IB kinase removes IB NF-B trans-locates to nucleusNF-kappa B is an anti-apoptotic factorVia NF-kappaB TNF blocks its own cell death potential chemotherapy activates NF-kB within tumor cells NF-kB inhibitors augment chemotherapy many common syn

37、thetic (e.g., aspirin), and traditional (e.g., green tea, curcumin) remedies target, at least in part, the NF-kB signaling pathway The Wnt signaling pathwayIntegrin 傳導(dǎo)通路Networks of Signal Transduction600 G protein-coupled receptors+Multiple gene families and combinationsof G protein subunits20G isof

38、orms6 G isoforms12 G isoforms+Multiple gene families for selected effector proteinsAdenylyl cyclasesPhospholipasesIon channelsThe phenomenon of crosstalk increases the complexity and variety of signal transduction pathways in a cell. The net result is that a single cellular response may be sensitive

39、 to several different extracellular stimuli. Crosstalk is, in part, a reflection of multiple gene families for all major protein components of signal transduction pathways. The magnitude of amplification within this cellular cascade structure often exceeds 10+4. That is, the binding of one molecule

40、of ligand to a cell-surface receptor leads a change of 10,000-fold in the intracellular concentration of a metabolic product. 腫瘤臨床中的信號(hào)傳導(dǎo) 信號(hào)傳導(dǎo)異常與腫瘤發(fā)生 腫瘤治療增殖失控：生長(zhǎng)因子： c-Sis生長(zhǎng)因子受體: HER-2 蛋白激酶: c-SrcG蛋白: c-Ras細(xì)胞周期調(diào)控因子: Rb凋亡受阻TNFFas/FasLBcl/Baxp53侵襲與轉(zhuǎn)移IntegrinE-CadherinVEGF信號(hào)傳導(dǎo)異常與腫瘤發(fā)生腫瘤發(fā)生與發(fā)展是多因素作用、多基因參與、經(jīng)

41、過(guò)多個(gè)階段形成。腫瘤中常見(jiàn)的信號(hào)傳導(dǎo)異常Familial adenomatosis polyposis coli （FAP,APC)Deleted in colorectal carcinoma(DCC) Based on genetic analysis, at least two pathways are characterized in detail, which lead to colon cancer development. One pathway (indicated with red arrows) initiates with mutations in the APC gen

42、e followed by mutations in K-ras, deleted in colorectal cancer (DCC) and p53 genes. The second pathway (indicated with blue arrows) is initiated by mutations in the MMR genes (hMSH3, hMSH) and other genes (TGFbetaIIR, IGFIIR, PTEN, BLM, Tcf-4, Bax and E2F4). Beside these there are many other less ch

43、aracterized pathways with a high degree of overlapping among them. At least, seven gene mutations are needed to develop a normal epithelial cell into carcinoma. Frontiers in Bioscience 10, 1118-1134, May 1, 2005Model for genetic alterations in the development of colorectal cancerAdaptorGRFs12,13,61,

44、63mutationGTPaseGTP結(jié)合活性半衰期延長(zhǎng)3-9倍結(jié)腸癌、胰腺癌等 突變的ras降低了Ras蛋白水解GTP為GDP的能力。降低了自身內(nèi)源的GTPase的活性，導(dǎo)致Ras與GTP持續(xù)結(jié)合具有促進(jìn)細(xì)胞增長(zhǎng)的作用。G TArg Valcolorectal carcinomas 30-60%, K-rasnon-melanoma skin cancer 30-50%, H-ras hematopoietic neoplasia of myeloid origin 18-30%, K-and N-rasSeminoma 25-40%, K-ras In other tumors, a mu

45、tant ras gene is found at a lower frequency:for example, in breast carcinoma (0-12%, K-ras), Neuroblastoma (0-10%, K- and N-ras)Wnt-1 was found as an oncogene activated by the Mouse Mammary Tumor Virus in murine breast cancer. APC was first isolated as a tumor suppressor gene in human colon cancer.

46、After establishing that APC and beta-catenin bind to each other activating mutations in the human beta-catenin gene were found in human colon cancer and melanomas .These mutations alter specific beta-catenin residues important for GSK3 phosphorylation and stability . The role for Frat/GBP in cancer

47、is illustrated by its activation by proviral insertion in mouse lymphomas . A recent example of the link between cancer and Wnt signaling comes from the identification of the Drosophila gene legless as a homolog of Bcl-9 a gene implicated in B cell malignancies.Interestingly, mutations in the human

48、AXIN1 gene were reported in human hepatocellular carcinomas. TCF1 can also act as a tumor suppressor gene , as Tcf1 mutant mice develop adenomas in the gut and mammary glands Several components of the Wnt signaling pathway have been implicated in human tumors or experimental cancer models.EMT : epit

49、helial-mesenchymal transition腫瘤的轉(zhuǎn)移及EMT （Epithelial-Mesenchymal Transition）Different types of EMTJ. Clin. Invest. 119:14201428 (2009). doi:10.1172/JCI39104Contribution of EMT to cancer progressionScience Vol 331,Mar25, 2011; Cell 147,Oct14, 2011Biphasic activities of the TGF- signaling pathway during

50、 tumorigenesis: from the tumour suppressor to the tumour promoter腫瘤治療腫瘤治療 單克隆抗體Herceptin: HER-2 STI571(Gleevec): Bcr-Abl May 2001magic bullet Hematopoietic stem cellMatureBloodCellsCell DifferentiationLeukemiaCytogenetic Abnormality of CML:The Philadelphia ChromosomeChronic Myelogenous Leukemia (CML

51、). The Leukemia & Lymphoma Society. P-32 20M. Rev 10/99.p210Bcr-Abl Fusion Protein Tyrosine KinaseExtracellular spaceY177BAP-1GRB2CytoplasmSH3SH2SH1CBLSHCCRKLThe Philadelphia Chromosome: t(9;22) Translocation22bcrablPhbcr-ablFUSION PROTEINWITH TYROSINEKINASE ACTIVITY99+Structure of BCR-ABL Fusion Pr

52、oteinsBcr-Abl Signal Transduction PathwaysGleevec-Tyrosine Kinase InhibitorGoldman JM. Lancet. 2000;355:1031-1032.Bcr-AblATPSubstrateSTI571Y = TyrosineP = PhosphateBcr-AblSubstratePPPPStructure of Gleevec (Imatinib Mesylate)Class: Phenylaminopyrimidines, 589.7 mwCH3SO3HOConclusionsCML is a result of

53、 a single genetic mutation termed the Philadelphia ChromosomeGleevec is a Tyrosine Kinase Inhibitor that targets the protein product of the mutationOngoing trials show Gleevec to be effective in Chronic Phase CMLAcceleratedBlast CrisisResponse to therapy is generally seen in 1 monthGleevec is genera

54、lly well-toleratedchemist Nicholas Lydon, a former researcher for Novartis, oncologist Brian Druker of Oregon Health and Science University (OHSU), Charles Sawyers of Memorial Sloan-Kettering Cancer Center Lasker-DeBakey Clinical Medical Research Award converting a fatal cancer into a manageable chr

55、onic condition.ForwardAdd one compound/wellSelect compound that produces phenotype of interestidentify protein targetReverseFind ligand for protein of interest by screeningAdd ligand to cellsAssay for phenotypeChemical BiologyMicroRNAs in caner: tiny players in big diseaseZhao QianDepartment of Path

56、ophysiologyShanghai JiaoTong University School of MedicineCell, Vol 75, 843-854, 3 December 1993The C. elegans Heterochronic Gene lin-4 Encodes Small RNAs with Antisense Complementarity to lin-14Rosalind C. Lee, Rhonda L. Feinbaum, and Victor Ambros endogenous 1925nt non-protein-coding RNAs up to 10

57、00 human miRNAsconservatism, tissue specificity and time-ordered in expression are the obvious characteristics of miRNA negatively regulate the expression of genes at the posttranscriptional level by blocking translation or/and degrading target mRNAs each microRNA regulates 100-200 target genesmicro

58、RNAs have been shown to control a wide range of biological functions such as stem cell development, cellular proliferation, differentiation and apoptosis.Marilena V. Iorio and Carlo M. Croce J Clin Oncol .2009.24.0317 Biogenesis, processing, and maturation of microRNAsMechanisms of microRNA regulati

59、on About half of the annotated human miRNAs map within fragile regions of chromosomes, which are areas of the genome that are associated with various human cancers different miRNAs are deregulated in primary human tumors tumor suppressor gene: let-7d, miR-15a, miR-16-1, et al. oncomirs : miR-155, mi

60、R-17-92 cluster, et al. B cell lymphomaCLLtumor suppressor geneoncomir目前發(fā)現(xiàn)與乳腺癌發(fā)生和發(fā)展密切相關(guān)的癌基因和抑癌基因有：c-myc基因 HER2基因 (C-erbB-2)RAS基因MTDH基因EZH2基因BRCA1 BRCA2P53P21P16.與乳腺癌發(fā)生和發(fā)展密切相關(guān)microRNAs:miR-21 miR-29miR-155miR-27amiR-25miR-10bmiR-125a/bmiR-145miR-17-5PmiR-26a.MiR-200 miRNAs as prognostic tools miRNAs