1 Molecular mechanism of cancer metastasis Dr. Yick-Pang Ching Department of Pathology Room L7-05, Faculty Medicine Building Tel: E.Mail:

2 Overview What is metastasis? Molecular mechanisms of metastasis Signalling pathways involved in metastasis

3 I) What is cancer metastasis?  Cancer defines as a population of cells that have lost their normal controls of growth and differentiation and are proliferating without check.  Metastasis is the process by which a tumor cell leaves the primary tumor, travels to a distant site via the circulatory system, and establishes a secondary tumor.

4 Forms of cancer metastasis

5 Preferential metastatic sites Primary tumourCommon distant site (s) Breast’ adenocarcinomaBone, brain, adrenal Prostate adenocarcinomaBone Lung small cell carcinomaBone, brain, liver Skin cutaneous melanomaBrain, liver, Bowel Thyroid adenocarcinomaBone Kidney clear cell carcinomaBone, liver, thyroid Testis carcinomaLiver Bladder carcinomaBrain NeuroblastomaLiver, adrenal

6 Reason for organ selectivity Mechanistic theory: determined by the pattern of blood flow. “ Seed and soil ” theory: the provision of a fertile environment in which compatible tumor cells could grow

7 Determining factors  Appropriate growth factors or extracellular matrix environment  Compatible adhesion sites on the endothelial lumenal surface  Selective chemotaxis at which the organ producing some soluble attraction factors to the tumor cells

8 II) Molecular mechanisms of metastasis

9 5 major steps in metastasis 1.Invasion and infiltration of surrounding normal host tissue with penetration of small lymphatic or vascular channels; 2.Release of neoplastic cells, either or single cells or small clumps, into the circulation; 3.Survival in the circulation; 4.Arrest in the capillary beds of distant organs; 5.Penetration of the lymphatic or blood vessel walls followed by growth of the disseminated tumor cells

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11 Stages of metastasis  Invasion : primary tumour cells enter circulation  Circulation to the secondary site of tumour growth  Colonisation : formation of secondary tumour

12 Tumor invasion 1.Translocation of cells across extracellular matrix barriers 2.Lysis of matrix protein by specific proteinases 3.Cell migration

13 Components of invasion a)Matrix degrading enzymes b)Cell adhesion c)Cell motility

14 a) Matrix degrading enzymes  Required for a controlled degradation of components of the extracellular matrix (ECM)  The proteases involved in this process are classified into serine-, cysteine-, aspartyl-, and metalloproteinase.

15 MMP family

16 Matrix metalloproteinases (MMP)  16 members, subdivided into 4 groups, based on their structural characteristics and substrate specificities  Soluble and secreted groups; collagenase, gelatinase and stromelysins  Membrane type (MT-MMP) group are anchored in the plasma membrane  A zinc ion in the active centre of the protease is required for their catalytic activities.

17 Regulation of MMP  MMP is controlled by an increased expression on a transcriptional level.  MMPs are calcium-dependent proteases, which are synthesized as a inactive proenzymes and are activated by the cleavage of a propeptide.  MMP activity is regulated by specific inhibitors, the tissue inhibitors of MMP (TIMPs). Binding TIMP to MMP is in a 1:1 stoichiometry.  MMP2 and MMP9, which cleave type IV collagen the major constituent of basement membrane, are believed to be of special importance

18 Serine proteases  Serine protease involved in ECM degradation are plasmin, plasminogen activators and cathepsin G.  Plasmin is believed to be the most important serine protease, firstly because its ability to degrade several matrix components like gelatin, fibronectin or laminin, and secondly by the possible activation of numerous proforms of MMPs by propeptide cleavage.  Plasmin is synthesized in its inactive proform, plasminogen, which can be converted to plasmin by plasminogen activator.

19 Plasminogen activator  Two main types : urokinase (uPA) and tissue (tPA).  uPA is bound to the surface of tumor cells by means of a specific receptor (uPAR)  There are specific inhibitors (PAI-1 and PAI-2) for the PA.

20 Interaction between tumour cells and the surrounding connective tissue

21 Cell adhesion and metastasis

22 b) Cell attachment 1.Integrin: cell-matrix adhesion 2.E-cadherin/catenin adhesion complex: cell-cell adhesion

23 1) Integrin  Heterodimeric transmembrane receptors consists of  and  subunits  Function to provide interactions between cells and macromolecules in the ECM  Integrin can affect the transcription of MMP genes

24 Integrin signaling

25 2) E-cadherin and catenin complex  Most important cell-cell adhesion molecules  Reduce expression of E-cadherin and catenin increase the invasiveness of tumor cells

26 Cadherin-mediated cell-cell adhesion

27 p120 catenin

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29 c) Cell migration 1.Small Rho GTPase family 2.Motility promoting factors

30 Small Rho GTPase Stimuli Cdc42 GTP Rac1 GTP Pak1 LIM kinase Cofilin Actin polymerisation MLC Kinase MLC Phosphorylation Contraction Stress fibers Detachment Filopodia Lamellipodia

31 Model of Rho GTPase regulation

32 Regulation of Rho GTPase

33 Cell movement

34 Rho GTPase is required for the transition of invasive phenotype

35 Signaling pathways related to integrin and small GTPase

36 E-cadherin and Rho GTPase signaling

37 Rho GTPase at different stages of tumour progression

38 2) Motility promoting factors  Hepatocyte growth factor/scattering factor  Insulin-like growth factor II  Autotaxin

39 HGF/scatting factor  Heterodimer of  and  chains  HGF normally acts as a paracrine growth factor, but in tumor cells it can act as an autocrine  HGF binds to the c-Met receptor and activated the downstream effectors

40 HGF induce cell scattering and invasion

41 HGF induce the formation of branched tubules

42 Signalling pathways responsible for MET-dependent invasive growth

43 HGF/Met regulate integrin, cadherin and MMPs during invasion