Non-cellular stroma

• Describe the components of the ECM

  Structural proteins, such as collagen, laminin and fibronectin, are organised as fibrillar structures embedded within a viscoelastic gel. The gel contains proteoglycans, glycoproteins, water and growth factors, as well as other metabolites secreted by cells. Crosslinking between chains creates this viscoelasticity.
• Describe the function proteoglycans in the ECM

  Proteoglycans are dominated by GAG chains, which are negatively charged and therefore attract water molecules. This allows hydration of the ECM, which provides a cushioning in normal tissues, allowing it to withstand forces. The negative charge also attracts growth factors.
• Describe how cancer cells alter the ECM

  Cancer cells upregulate Lysyl oxidase, which facilitates collagen crosslinking. This increases the number of crosslinks between collagen strands and makes the ECM stiffer. This can be felt as a palpable lump, which is one of the most noticeable signs of a tumour.
• Describe coordination of cell movement

  1. Actin polymerisation at the plus end protrudes the lamellipodium.
  2. Integrins form new focal adhesion points at the front of the cell
  3. Myosin motor proteins slide along actin filaments and cause contraction of the cell at the back and previous focal contacts at the back are released.
• Describe the role of integrins in cell adhesion

  Integrins function as heterodimers to transmit tensions across the plasma membrane. Integrins links fibronectin, which connects to collagen in the ECM, to adaptor proteins, which connect to the actin cytoskeleton.
• Describe how integrin conformational changes

  Integrin activity is regulated through changes in conformation. When bound to molecules on either side of the plasma membrane, integrins switch to an active conformation. This conformational change establishes a reversible mechanical linkage across the plasma membrane that can be switched on and off in order to let cells move through the tissue. Some integrins can bind to receptors on other cells, such as ICAM, which enables cell to cell adhesions.
• Describe ECM remodelling

  ECM must be degraded in order to allow cells to pass, which is mediated by two main groups of ECM degrading enzymes:

  • Matrix metalloproteinases: require bound Ca2+ or Zn2+ activity
  • Serine proteases: contain conserved serine residue in the active site

  These enzyme facilitate localised degradation of ECM in order to let cells pass through, whilst maintaining the overall ECM structure.
• Describe MMPs

  MMPs are released by myofibroblasts in response to TGF-beta. Degradation of the ECM allows for structural remodelling of the ECM, as well as releasing a variety of tethered growth factors. Examples include:

  • MMP-1: collagenase
  • MMP-2: gelatinase
• Describe how the activity of ECM degrading proteases is controlled

  • Some proteases are secreted in their inactive form:
  • localised activator then converts into their active form
  • Some proteases are confined by cell surface receptors
  • urokinase type plasminogen activator (uPA) is found at growing tips of some migrating cells and elevated in breast tumour tissue
  • Some proteases are inhibited by the actions of locally secreted inhibitors
  • e.g., tissue inhibitors of metalloproteinases
• What are invadopodia?

  These are cellular structures consisting of dynamic actin rich protrusions from the cell surface membrane and are usually found on cancer cells. Invadopodia degrade the ECM through local deposition of proteases and are key to cell invasion.
• Describe how a tumour manipulates the ECM to promote invasiveness

  Cancer cells degrade the basal lamina, allowing them to escape and invade the underlying connective tissue. Invading cancer cells can migrate along aligned collagen fibres that form tracks to facilitate tumour cell migration, while protease degrade ECM that forms a barrier to cell migration.
• Describe the epithelial to mesenchymal transition

  Cells start at the epithelium and then leave to revert to a mesenchymal cell fate and migrate to another location. EMT is aberrantly activated under pathological conditions such as cancer and is associated with cancer invasion and dissemination.
• Describe the different characteristics of mesenchymal cells

  • migratory
  • loss of cell to cell junctions
  • dynamic cell to ECM adhesions
• Describe the role of cadherins in EMT

  • EMT inducible transcription factors regulate expression of cadherin proteins in order to modulate cell adhesion.
  • In cancer progression, EMT can be utilised by cancer cells to contribute to malignancy
• Describe how adherens junctions are modulated

  • E-cadherin has a protein half life of 5-10h, making it highly susceptible to proteolytic degradation
  • Caspases degrade alpha and beta-catenin during cell death
  • Calpains are Ca2+ dependent cysteine proteases that modulate cadherin dependent cell to cell adhesions
  • Calpain is upregulated in breast tumour
• Describe the role of calpains in tumourigenesis

  • Calpains are switched on in response to activation of the oncogene Src
  • Calpain inhibitors suppress cell migration and proliferation
  • Calpain expression is upregulated in renal cell carcinomas that metastatsise to lymph nodes and in breast cancer tissue
• List therapies that target cancer invasion

  • Aspirin: restores E-Cadherin dependent cell to cell adhesions
  • MMP inhibitors: inhibit tumour induced degradation of ECM proteins
  • EGFR inhibitor: inhibit tumour growth and metastasis
  • VEGFR inhibitor: inhibits angiogenesis, tumour growth and metastasis