Mechanism of Cancer Invasion and Metastasis

Metastatic cascade: overview flowchart

Primary malignant tumor
        ↓
Local invasion through basement membrane and ECM
        ↓
Intravasation into lymphatic or blood vessel
        ↓
Survival in circulation as circulating tumor cells/tumor emboli
        ↓
Arrest and adhesion at distant capillary bed
        ↓
Extravasation through endothelium and vascular basement membrane
        ↓
Survival in foreign microenvironment (micrometastasis/dormancy)
        ↓
Colonization, angiogenesis and growth
        ↓
Clinically evident secondary metastatic tumor

1. Invasion of extracellular matrix (ECM)
                    +
2. Vascular dissemination, tissue homing and colonization

I. Local invasion of extracellular matrix

Epithelial basement membrane
        ↓
Interstitial connective tissue
        ↓
Vascular/lymphatic basement membrane
        ↓
Enter circulation

Steps of ECM invasion

1. Loosening of tumor cell-cell adhesion
                    ↓
2. Degradation of basement membrane and interstitial ECM
                    ↓
3. Attachment of tumor cells to remodeled ECM
                    ↓
4. Migration/locomotion through ECM
                    ↓
5. Intravasation

1. Loosening of tumor cell-cell adhesion

• Normal epithelial cells adhere to each other through E-cadherin, a transmembrane adhesion molecule.
• In many carcinomas, there is decreased E-cadherin function due to:
  • CDH1 mutation, for example in diffuse gastric carcinoma and lobular carcinoma breast
  • Transcriptional repression during epithelial-mesenchymal transition (EMT)
• Loss of E-cadherin causes detachment of malignant cells from the primary tumor.

Normal epithelial cells
E-cadherin-mediated adhesion
        ↓ loss of E-cadherin
Reduced cell-cell cohesion
        ↓
Tumor cells detach and become invasive

2. Degradation of ECM and basement membrane

Important proteases

• Tumor-associated fibroblasts and inflammatory cells also produce proteases.
• Reduced metalloproteinase inhibitors further favors matrix degradation.
• MMP-9 can release ECM-sequestered VEGF, promoting angiogenesis.
• ECM breakdown also produces fragments of collagen and laminin that promote tumor-cell migration.

Tumor cells/stromal cells
        ↓
MMPs, cathepsin D, uPA
        ↓
Basement membrane + interstitial ECM degradation
        ↓
Release of VEGF and chemotactic/growth-promoting fragments
        ↓
Facilitated invasion and angiogenesis

3. Attachment to remodeled ECM

• Tumor cells alter expression of integrins, which mediate cell-ECM attachment.
• Proteolysis of collagen IV and laminin exposes new binding sites for tumor-cell receptors.
• This promotes attachment and directed migration.
• Malignant cells resist anoikis, which is apoptosis induced by loss of attachment to ECM.

MMP-mediated cleavage of collagen IV/laminin
        ↓
New ECM binding sites exposed
        ↓
Altered integrin-mediated adhesion of tumor cell
        ↓
Survival despite detachment + migration

4. Migration and locomotion

Attachment at leading edge
        ↓
Actin cytoskeleton contraction
        ↓
Forward movement
        ↓
Detachment at trailing edge

• Tumor-derived chemokines and growth factors, including IGF
• Cleavage products of collagen and laminin
• Stromal cell-derived hepatocyte growth factor/scatter factor (HGF) acting through the MET receptor
• Signals from cancer-associated fibroblasts, inflammatory cells and endothelial cells

II. Epithelial-mesenchymal transition (EMT)

EMT flowchart

Epithelial carcinoma cell
E-cadherin+, cohesive, polarized, relatively immobile
        ↓
EMT-inducing signals
        ↓
SNAIL, TWIST, ZEB transcription-factor activation
        ↓
↓ E-cadherin and epithelial markers
↓ Cell polarity and intercellular junctions
        +
↑ Vimentin, smooth muscle actin and mesenchymal phenotype
        ↓
Spindle-shaped, motile, invasive tumor cell
        ↓
ECM invasion and intravasation

Molecular basis

• EMT is regulated by transcription factors SNAIL and TWIST.
• These suppress epithelial markers, especially E-cadherin.
• Simultaneously, cells increase expression of mesenchymal markers such as:
  • Vimentin
  • Smooth muscle actin
• The result is a pro-migratory, invasive phenotype.

Reverse process: MET

EMT at primary site → dissemination
MET at distant site → colonization and growth

III. Vascular dissemination

1. Intravasation

Invasive tumor cell
        ↓
Penetrates vessel basement membrane
        ↓
Passes through endothelial layer
        ↓
Intravasation
        ↓
Circulating tumor cell

2. Survival in circulation

• Mechanical shear stress
• Immune destruction
• Anoikis
• Failure to adapt to circulation

Tumor cells + platelets + fibrin
        ↓
Tumor embolus
        ↓
Protection from immune cells and shear stress
        ↓
Improved survival and arrest in capillary beds

• Coating them and reducing immune recognition
• Promoting aggregation
• Assisting arrest within capillary beds

IV. Homing, extravasation and colonization

1. Arrest and homing at a distant organ

Anatomic vascular drainage
        +
Tumor-cell organ tropism
        +
Favorable microenvironment ("seed and soil")
        +
Ability to escape dormancy

Examples

Basis of organ tropism

• Tumor-cell adhesion molecules bind preferentially to endothelium of target organs.
• CD44 expression may facilitate spread to lymph nodes and other sites.
• Chemokine receptors on tumor cells respond to chemokines in particular organs.
• The target organ must provide a favorable environment for growth.

Tumor cell = seed
Target-organ microenvironment = soil
        ↓
Metastasis occurs only when seed reaches suitable soil

2. Extravasation

Tumor embolus arrests in capillary bed
        ↓
Adhesion to endothelial cells
        ↓
Passage between endothelial cells
        ↓
Penetration of vascular basement membrane
        ↓
Entry into distant tissue

3. Colonization

• Extravasated cells must survive in an unfamiliar microenvironment.
• Many remain dormant or die.
• Successful cells proliferate, recruit blood vessels by angiogenesis, and form a clinically evident metastatic deposit.

Micrometastasis
        ↓
Adaptation to distant tissue
        ↓
Escape from dormancy
        ↓
Angiogenesis
        ↓
Progressive metastatic colony

Conclusion / high-yield summary

Loss of E-cadherin / EMT
        ↓
ECM degradation by MMPs and other proteases
        ↓
Attachment to remodeled ECM via integrins
        ↓
Migration and invasion
        ↓
Intravasation
        ↓
Survival as platelet-coated tumor emboli
        ↓
Arrest and homing in target organ
        ↓
Extravasation
        ↓
Colonization, angiogenesis and metastatic growth