Metalloproteinases

• Metalloproteinases


  Divergent roles in cancer biology

  
• Module ME3034, Course cancer, Revised Not started, Date

  
• ECM


  • A 3D, non-cellular structure, provides physical support for tissue integrity and elasticity
  • Dynamic structure that is constantly remodelled to control tissue homeostasis
• ECM components


  • Basement membrane - separates the epithelium from the surrounding stroma
  • Interstitial CT matrix - surrounds cells and provides structural scaffolding for tissues
• Fibrous ECM components


  • Collagen
  • Elastin
  • Fibronection
  • Laminin

  
• Non-fibrous ECM components


  • Proteoglycans
  • Polysaccharide
• Examples of MMPs in health and disease

  • Development
  • Immune system
  • Matrix maturation
  • Stem cell differentiation
  • Wound healing
  • Bone remodeling
  • Inflammation
  • Arthritis
  • Angiogenesis
  • Cancer

  
• MMPs in cancer progression

  • Promote tumor growth by regulating proliferation and apoptosis
  • Activating GFRs through release of cell membrane associated ligands
  • Proteolytic inactivation of cell death Rs FAS
  • Create tumour promoting micro environment
  • Promote invasion and metastasis

  
• Failures of early therapies
• Anti-target


  Molecule/protein with essential or host protective roles in the normal state of a cell or tissue function. Inhibition of its activity results in clinically unwanted side effects and initiation or worsening of disease
• Target


  Molecule or protein that unambiguously contributes to a disease. Inhibition of a validated drug target by a drug reverses the disease course or at the very least holds it static or slows disease progression; at best, drug targeting restores the normal state of a cell or tissue function

  
• MMPs


  • Pro-peptide domain - needs to be cleaved in order for protein to become activated
  • Catalytic domain Zn is present to allow catalytic activity

  
• Activation and chemical strategies for inhibition

  1. Access to Zn is blocked by pro-domain as it is bound to Zn via S

  2. Proteolytic cleavage of pro-domain to allow entry of water

  3. Activated MMP
• BMMPI


  MMPi which is coupled to a molecule which has the ability to guide the drug into the bone
• MT1MMP


  Migration, invasion and metastasis
• Protease activated prodrug


  1. Toxic warhead is bound to a peptide to form prodrug

  2. Enters the cancer cell via R-mediated endocytosis

  3. The cleavage by tumor enriched protease will occur

  4. Removal of remaining peptide stub by non-specific exopeptidases

  5. Prodrug > drug - kills cancer cells

  
• Ideal characteristic for protease activated prodrug development

  • Enzyme is well characterised biologically and its cleavage preferences are known
  • Protease is present and active at higher levels in the TME and has low expression or lack of activity in normal tissues
  • Absent or inactive in the circulation to minimise systemic toxicity
  • High affinity and selectivity for the designed prodrug, leading to rapid release of the active form of the drug in vivo
• ADAMs


  A disintegrin and metalloprotease are a family of type 1 transmembrane glycoproteins, related to snake venom MMPs and disintegrins

  
• Implications of ADAMs


  • Sperm egg binding and fusion
  • Cell fate determination in the NS
  • Cell migration
  • Axon gorwth and guidance
  • Muscle development and myoblast fusion
  • Adipogenesis
  • Diverse aspects of immunity
  • Cell/cell or cell/matrix interactions
  • Epithelial maturation
  • Neuro-protection
  • Cell signalling
• ADAMs


  • Pro domain and metalloprotease domain
  • Disintegrin domain - can bind integrins
  • IC domains: Y tyrosines
  • Great variability in IC domain

  
• ADAMs and remodeling of ECM
• ADAMs and adhesion
• ADAMs and shedding: EGFR activation

  Direct proliferation response transduction mitogenic signals through Ras-MAPK pathway, suggesting a direct role in cell survival

  
• Regulated intra membrane proteolysis


  ADAM cleaves the EC region > conformation change of protein > y-secretase cleaves in the juxtamembrane region of the IC section > signalling

  
• ADAM15


  • Overexpressed in breast, prostate and pancreatic cancer, haematological malignancies
  • Enhances heterotypic and homotypic cell-cell interaction
  • Regulates the permeability of endothelial cells and pathological neovascularisation
• Alternative splicing of ADAM15 can lead to proline-concentrated regions to interact with SH3 domain
• Early stage cancer with ADAM15B isoform correlated with a worse prognosis

  
• ADAM15B


  Cleaves FGFG2iiib (tumour suppressor in breast cancer) upon phosphroylation by Src

  
• Splicing affects focal adhesion turnover

  Long focal adhesion points - mature adhesions that are stable which are not processed > if not broken down, cancer cells have very poor motility

  
• Bypass signalling
• MEK kinase inhibitor anti-tumour therapy

  1. MEK kinase inhibitor treatment to suppress tumour growth results in bypass signalling to regain proliferation and drug resistance

  2. MEKi leads to a global reduction of ectodomain including shedding across a wide range of substrates

  3. ADAM blocked by TIMP1 at the cell surface

  4. High AXL-levels at the cell membrane

  5. AXL activated (phosphorylated)

  6. JNKAkt-cJun activated

  7. Unregulated TIMP1 MMPi) synthesis

  
• AXL and MAPK inhibitor treatment

  1. Reduces tumour growth and metastasis

  2. MEKi induces ADAM10 driven cell surface TIMPI accumulation

  3. AXL inhibitor blocks evasive signalling

  4. MAPK inhibitor can be replaced with MMP inhibitors only in the presence of AXL inhibitor

  
• MAPK Inhibition and blocking TIMP1 with Ab

  1. Prevents drug resistance and promotes tumour killing

  2. ADAM10 cleaves AXL from the cell membrane

  3. Loss of bypass signalling via AXL

  
• What kills most patients in cancer?
• The metastatic Cascade


  1. Clonal expansion, growth, diversification, angiogenesis

  2. Metastatic subclone

  3. Adhesion to and invasion of basement membrane

  4. Passage through extracellular matrix

  5. Intravasation

  6. Interaction with host lymphoid cells

  7. Tumor cell embolus

  8. Adhesion to basement membrane

  9. Extravasation

  10. Metastatic deposit

  11. Angiogenesis

  12. Growth

  
• ECM


  • A three-dimentional, non-cellular structure, provides physical support for tissue integrity and elasticity.
  • It is dynamic structure that is constantly remodelled to control tissue homeostasis.
• ECM types


  • The basement membrane: separates the epithelium from the surrounding stroma.
  • Interstitial connective tissue matrix: surrounds cells and provides structural scaffolding for tissues
• Early trials- stop cancer metastasis and angiogenesis by inhibiting MMPs

  
• Only three MMPs were known then: MMP1, MMP2, MMP3
• Focus on A category of proteases → metalloproteases