Angiogenesis Flashcards
Angiogenesis
New blood vessel formation
Sprouting angiogenesis
- tip/stalk cell selection
- tip cell navigation and stalk cell proliferation
- branching coordination
- stalk elongation, tip cell fusion and lumen formation
- perfusion and vessel maturation
How to make a blood vessel
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Regulators of angiogenesis: Activators
GROWTH FACTORS -VEGF family -FGF family -TGF-beta -PDGF SOLUBLE FACTORS -IL-6 -Factor XIII -TNF-alpha CELL SURFACE RECEPTORS -Alpha-V beta-3
Regulators of angiogenesis: Inhibitors
EXTRACELLULAR MATRIX -Thrombospondin-1 -Angiostatin -Endostatin SOLUBLE FACTORS -sVEGF-R -IL-10 -IL-12 -TNF-alpha CELL SURFACE RECEPTORS -Alpha-V beta-3
Regulators of angiogenesis: Maturation and Integrity
- VE-Cadherin (Junctions)
- Angiopoietin/Tie2
- Notch pathway
- ERG pathway
- Platelets
Hypoxia
A lower-than-normal concentration of oxygen in arterial blood
- HIF (hypoxia-inducible transcription factor=controls regulation of gene expression by oxygen)
- pVHL (Von-Hippel-Lindau tumour suppressor gene=controls levels of HIF)
Vascular Endothelial Growth Factor (VEGF) and its receptors
- Family of 5 members: VEGF-A, VEGF-B, VEGF-C, VEGF-D, and placental growth factor (PlGF)
- Three tyrosine kinase receptors: VEGF receptor (VEGFR)-1, VEGFR-2, and VEGFR-3; and co-receptors neuropilin (Nrp1 and Nrp2)
- VEGFR-2 is the major mediator of VEGF-dependent angiogenesis, activating signalling pathways that regulate endothelial cell migration, survival, proliferation
Tip cells and sprouting angiogenesis
-specialised endothelial tip cells lead the outgrowth of blood-vessel sprouts towards gradients of VEGF
Canonical Notch signalling pathway
- Notch receptors and ligands=membrane-bound proteins that associate through their extracellular domains.
- The intracellular domain of Notch (NICD) translocates to the nucleus and binds to the transcription factor RBP-J
Selection of tip cells: VEGF/Notch signalling
- In stable blood vessels, Dll4 and Notch signalling maintain quiescence
- VEGF activation increases expression of Dll4
- Dll4 drives Notch signalling, which inhibits expression of VEGFR2 in the adjacent cell
- Dll4-expressing tip cells acquire a motile, invasive and sprouting phenotype
- Adjacent cells (Stalk cells) form the base of the emerging sprout, proliferate to support sprout elongation
Sprout outgrowth and guidance
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Macrophage participation in vessel anastomosis
- Macrophages play a significant role in both physiological and pathological angiogenesis
- Macrophages carve out tunnels in the extra cellular matrix (ECM), providing avenues for capillary infiltration
- Tissue-resident macrophages can be associated with angiogenic tip cells during anastomosis
Platelet role in angiogenesis
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Stabilisation and quiescence
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Tight junctions and adherens junctions in endothelial cells
- Constitutively expressed at junctions
- Homophilic interaction mediates adhesion between endothelial cells and intracellular signalling
- Controls contact inhibition of cell growth
- Promotes survival of EC
Mural cells (pericytes) in stabilising neovessels
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Signalling pathways controlling stability: The Angiopoietin-Tie2 ligand-receptor system
- Ang-1 and Ang-2 are antagonistic ligands of the Tie2 receptor
- Ang-1 binding to Tie2 promotes vessel stability and inhibits inflammatory gene expression
- Ang-2 antagonises Ang-1 signalling, promotes vascular instability and VEGF-dependent angiogenesis
Increase in Ang-2 plasma levels during:
- Congestive heart failure
- Sepsis
- Chronic kidney disease
Tumour angiogenesis and neovasculature
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The angiogenic switch
-Discrete step in tumour development that can occur at different stages in the tumour-progression pathway, depending on the nature of the tumour and its microenvironment
Tumour blood vessels
-irregularly shaped, dilated, tortuous
not organized into definitive venules, arterioles and capillaries
-leaky and haemorrhagic, partly due to the overproduction of VEGF
-perivascular cells often become loosely associated
-some tumours may recruit endothelial progenitor cells from the bone marrow
Tumour neovasculature: comparative tortuosity (twisted) and disorganisation
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Multicellular response promotes tumour angiogenesis
- Cancer-associated fibroblasts (CAFs) secrete extracellular matrix; pro-angiogenic growth factors, (VEGFA; FGF2; CXCL12; PDGFC)
- Pericytes are loosely associated with with tumour-associated blood vessels (TABVs), and this favours chronic leakage in tumours. This is enhanced by angiopoietin 2 (ANGPT2)
- Platelets release pro-angiogenic mediators and proteases that support the proliferation and activation of CAFs, such as PDGFB and TGFβ
Role of platelets in tumour angiogenesis
LINK BETWEEN CANCER PROGRESSION AND THROMBOCYTOSIS
Activated platelets are a source of:
- pro-angiogenic factors: VEGFA, platelet-derived growth factors (PDGFs), FGF2
- angiostatic molecules: thrombospondin 1, plasminogen activator inhibitor 1 (PAI1), endostatin
- Tumours cause platelet activation, aggregation and degranulation
- Disrupting platelet function does not obviously impair tumour angiogenesis, however the overall outcome of platelet activation in tumours appears to be pro-angiogenic
Therapeutic strategies to inhibit VEGF signalling
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VEGF inhibition by soluble VEGFR1 (Flt-1) reduces tumour growth
- Cells stably transfected with control or sFlt-1 plasmid to promote Flt-1 (VEGFR1) expression
- VEGFR1 binds to VEGF and “mops it up” preventing it from stimulating angiogenesis
- Flt-1 expression reduces tumor growth in vivo, without affecting tumor cell growth in vitro: effect on vasculature
Anti-VEGF humanised MAb (Avastin)
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Avastin side effects
- GI perforation
- Hypertension
- Proteinuria
- Venous thrombosis
- Haemorrhage
- Wound healing complications
Limited efficacy of Avastin
- No overall survival advantage over chemo alone
- No quality-of-life or survival advantage
- In some cases benefits are transitory, followed by a restoration of tumour growth and progression
- In other cases there is no objective benefit
Potential mechanisms of resistance to anti-VEGF therapy in cancer
- VEGF inhibition aggravates hypoxia increasing tumour’s production of other angiogenic factors or increases tumour invasiveness
- Tumours vessels maybe less sensitive to VEGF inhibition due to vessel lining by tumour cells or endothelial cells derived from tumours
- Tumour cells that recruit pericytes maybe less responsive to VEGF therapy
The future for anti-angiogenic therapy
-Anti-angiogenic therapy in combination with other anti-cancer therapies
-Resistance: combinatorial strategies involving angiogenesis inhibition & drugs targeting resistance mechanisms
-Novel non-VEGF targets – novel molecular mechanism
-Anti-angiogenic therapy in other diseases:
Retina vascularization (diabetic retinopathy, wet AMD)
Finding novel molecular mechanism cell by cell: single cell RNASeq of tumour endothelium
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Age-related macular degeneration (AMD)
Abnormal growth of choroidal blood vessels
- ‘Leaky’ vessels cause oedema
- Visual impairment
Anti-VEGF therapy for age-related macular degeneration (AMD): Lucentis
- AMD is the main cause of blindness
- Avastin not FDA approved for AMD, but used off-label
- Lucentis developed by Genentech from the parent molecule Avastin
- June 2006: FDA approval for Lucentis for AMD
- High efficacy of both treatments in maintaining or improving vision
- Many patients become refractory to treatment >2 years
- Ranibizumab (Lucentis)=$2,023 per dose (up to 12 injections per year)
- Bevacizumab (Avastin)=$55 per dose.
How to find better therapeutic strategies to inhibit angiogenesis in cancer
- Tumours are complex three-dimensional (3D) structures with their own unique microenvironments
- We lack good in-vitro models - our understanding of tumour behaviour in a complex 3D environment is limited and drug screens are often misleading
- Studies are performed on cell lines growing as two-dimensional (2D) monolayers, which do not mimic the complex interplay between tumour cells and their extracellular environment
- The phenotype of tumour cells when cultured in 2D vs 3D is different
- Crucially, tumours receive nutrients and therapeutics through the vasculature, which is not included in any in-vitro tumour models
‘tumour-on-a-chip’ platform
-develop a microphysiological system that incorporates human cells in a 3D extracellular matrix, supported by perfused human microvessels
