6. Neoplasia VI- Angiogenesis And Metastasis

Question 1

Describe the four major steps and the factors involved in the formation of new capillaries during the angiogenic process (objective)

Answer 1

Answer later

Question 2

Describe the role of angiogenesis/neovascularization in the growth and metastasis of tumor cells (objective)

Answer 2

Answer later

Question 3

Describe the concept of the “angiogenic switch” in a tumor and the factors that control it (objective)

Answer 3

Answer later

Question 4

Describe the key events in the epithelial to mesenchyme transition that induce metastasis potential to a tumor (objective)

Answer 4

Answer later

Question 5

Describe the process of metastasis and how tumors are targeted to specific organs (objective)

Answer 5

Answer later

Question 6

Interesting Facts

Answer 6

1cm3 of tissue is 10^9 cells
1cm3 of tumor is 10^11
Time of detection depends on primary site
Signs/symptoms of cancer: physical pressure, organ dysfunction
Death from cancer: primary tumor (10%), metastases (90%)
Metastatic tropism (tumors originating from given organ preferentially seed in particular tissue)

Question 7

Angiogenesis and Invasion Video

Answer 7

When cells don’t have enough oxygen, VEGF is secreted, leading to TIP cell formation (filopenol) and stalk cells
Recruit pericytes to stabilize structure

Tumor cells can have poorly assembled vessels, continues secreting VEGF and spreads

Question 8

Three Types of Capillaries

Answer 8

Continuous

Fenestrated

Sinusoid

Question 9

Continuous capillaries

Answer 9

Have tight occluding junctions that seal the spaces between endothelial cells. All transport must take place across the membranes.

Question 10

Fenestrated capillaries

Answer 10

Have perforations (fenestrations) through the endothelial cells that allow exchange of small molecules with blood (ie endocrine organs, intestinal wall)

Holes in cytoplasm

Question 11

Sinusoid capillaries

Answer 11

Have wide spaces between endothelial cells, large fenestrations, and a discontinuous basement membrane that allow for exchange of macromolecules and cells with tissues and blood (ie bone marrow, liver and spleen

Basement membrane not continuous (like tumors)

Question 12

Endothelial cell, basement membrane and pericyte (schematic)

Answer 12

Pericyte on outside to stabilize capillary

Basement membrane on outside

Endothelial cell on inside

Question 13

Tumors require access to circulation in order to grow and survive and shed waste products and carbon dioxide

Answer 13

Cancer cells grow preferentially around blood vessels
-tumor cells more than 0.2mm away from vessels were non-growing
-those even further were dying
0.2mm=distance oxygen can diffuse through living tissues
Hypoxia can lead to necrosis

Question 14

Vessel 1- Anoxia- Vessel 2 (diagram)

Answer 14

PH is lowest (most acidic) in between vessels
Glucose/ATP/Oxygen is lowest in between vessels
Lactate is highest between vessels in anoxia zone

Question 15

Developing vasculature through angiogenesis is essential for tissue survival and is seen in:

Answer 15

Embryonic development
Implantation of the placenta
Wound healing
Many disease processes
Tumorigenesis

Question 16

VEGF and bFGF

Answer 16

Receptors display on the surfaces of endothelial cells

Stimulate endothelial cell proliferation

EGF molecules bind to tyrosine kinase which have dimerized, which then phosphorylate each other after binding

Question 17

Production of VEGF is governed by availability of oxygen

Answer 17

HIF-1a has two paths based on oxygen
Normoxia: enzyme proline hydroxylase brings OH for degradation, recognition and binding by pVHL and other proteins, polyubiquitylation, degradation in proteasome
Hypoxia: angiogenesis- HIF-1a/b activate VEGF gene in nucleus

Question 18

Endothelial cell nuclei join to form capillary lumen

Answer 18

Slide 16

Question 19

As tumor growth continues, capillaries are increasing

Answer 19

• Circulating endothelial cells from bone marrow are recruited to settle in the tumor stroma and differentiate
• Capillaries are also being assembled from endothelial cells present within tumor stroma
• Other key players: TGF-beta, IL-8, angiopoietin, angiogenin 1/2.

Question 20

The chaotic organization of tumor-associated vasculature

Answer 20

Normal tissue very organized

Tumor tissue is chaotic and more permeable (can get more nutrients and dispose of waste)

Question 21

Steps in formation of new capillaries (4 steps)

Answer 21

1. Stimulation of endothelial cells by angiogenic growth factors (VEGF)
2. Degradation of the parental vessel basal lamina by activated endothelial cells to facilitate the formation of a capillary sprout
3. Endothelial cell migration/proliferation
4. Maturation of endothelial cells involving formation of capillary tubes, reformation of basal lamina and recruiting pericytes

Question 22

Angiogenesis in Tumors

Answer 22

• Cells of the vasculature (endothelial cells, pericytes, and smooth muscle cells)
• Nonvascular cells (neoplastic cells, supporting cells of the stroma)

Question 23

Tumors resemble wound healing sites

Answer 23

Slide 22

Question 24

Step 1: Stimulation of Endothelial Cells by Angiogenic Growth Factors

Answer 24

1. Basic Fibroblast Growth Factor (bFGF)
2. Vascular Endothelial Growth Factor (VEGF)

Tumor expresses hypoxia, growth factors bind to endothelial

Question 25

Step 2: Degradation of the Capillary Basal Lamina by Activated Endothelial Cells

Answer 25

Secreted Proteases destroy basement membrane: 1. Matrix metalloproteinases (MMPs) 2. Plasminogen Activator Urokinase (uPA) *MMPs secreted to allow cells to migrate through the ECM

Question 26

Step 3: Capillary Sprout Formation and Migration of Endothelial Cells

Answer 26

Regulation of Cell Migration: 1. Integrins 2. Extracellular Proteinases (continue degradation)

Question 27

Step 4: New Vessel Maturation

Answer 27

1. Angiopoietin 1 (Ang 1)- tell to stop growing | 2. Platelet Derived Growth Factor (PDGF)- binds to pericyte precursor so that pericyte stabilizes vessel

Question 28

Tripping the "angiogenic switch" is essential for tumor expansion

Answer 28

Many tumor cell populations initially lack the ability to attract blood vessels. Not all tumors grow the same

Question 29

Angiogenic Switch Diagram

Answer 29

Hyperplastic islet can recruit signals for the bone marrow to get mass cells and macrophages which secrete MMPs which activate VEGF to activate the switch. Hyperplastic islet can undergo regular angiogenic switch to ultimately become angiogenic islet

Question 30

The Angiogenic Switch

Answer 30

-The induction of angiogenesis is a component of the tumor phenotype that is activated during the early pre-neoplastic stages in the development of the tumor -Most tumors arise without angiogenic activity, exist in this dormant stage (carcinoma in situ, CIS) without vascularization for long periods of time, and become vascularized when a subset of cells within the tumor switches to the angiogenic phenotype Dormant lesion: proliferation equals apoptosis, angiogenic inducers low, inhibitors high . Metastatic lesion: proliferation greater than apoptosis, angiogenic inducers high, inhibitors low

Question 31

Angiogenic Switch: Prevascular Phase

Answer 31

-Angiogenic activity absent/low -Tumors remain small with volumes in a few cubic mm (proliferation=apoptosis) -Tumors are generally thin or flat, stable, asymptomatic and rarely metastatic -Micrometastases may have similar pre-vascular phase Hyperplasia remains above basement membrane

Question 32

Angiogenic Switch: Vascular Phase

Answer 32

-Characterized by tumors which have entered phase of rapid growth, intensified invasion and increased metastatic potential -A bi-directional paracrine relationship between the tumor cells and endothelial cells is established. Endothelial-derived growth factors stimulate the grow of tumor cells -Hypoxic areas of tumor stimulate VEGF production and subsequent endothelial cell growth -Associated with increased appearance of symptoms. Bleeding, local edema, inappropriate hormonal activities, hypercoagulation states and cachexias Normal to Hyperplasia to Dysplasia (angiogenesis already started but membrane intact; try to catch this) to Carcinoma

Question 33

Angiogenic Switch: Heterogeneity of Angiogenic Activity in Tumors

Answer 33

- Neovascularization of tumor usually originates in a subset of cells due to presence of a mixture of angiogenic and non-angiogenic cells - Metastases derived from the angiogenic portion of tumor are already angiogenic upon arrival at the target organ and therefore have an increased chance of becoming a detectable metastasis

Question 34

Anti-Angiogenic Therapy

Answer 34

- Specificity have less side effects - Angiogenesis in adults limited to reproductive tissues and wound healing - Target accessible vessel endothelial cells and not traditional tumor vasculature which is a barrier to normal drug delivery - Endothelial cells are pretty stable with low mutation rates, so resistance is reduced - In theory, can work on all solid tumors regardless of origin, since endothelial cells do not vary from one type of tumor to another.

Question 35

Metastasis is Angiogenesis-Dependent

Answer 35

Slide 39

Question 36

Sequence of Events in Tumor Cell Metastasis

Answer 36

- Initial transforming event - Proliferation of transformed cells - Compromised nutritional supply to the tumor mass requiring release of tumor angiogenesis factors - Endothelial cell expansion and reorganization - Local invasion and destruction of ECM and parenchymal cells leading to migration of tumor cells away from the primary tumor mass - Penetration of cancers cells through the blood vessel wall (intravasation) - Arrest of cancer cells in the lumen of small blood vessels or lymphatics - Reverse penetration of blood vessels - Organ colonization resulting in the formation of the secondary tumor (metastases)

Question 37

Tumor Cells Can Secrete Enzymes that Degrade the Adhesion Proteins Between Cells of the Vasculature

Answer 37

A. Loosening of intercellular junctions B. Degradation of ECM C. Migration and Invasion

Question 38

Epithelial-Mesenchymal Transitions in Cancer

Answer 38

Cytoskeleton reorganization Membrane reorganization Increased growth Increased motility

Question 39

Epithelial markers

Answer 39

E-cadherin

Question 40

Mesenchymal markers

Answer 40

Vimentin Fibronectin N-cadherin

Question 41

Seed and Soil Hypothesis

Answer 41

- Colonization of metastasized tumor cells is inefficient - Micrometastases often seed organs, but most fail to grow into clinically observed tumors - Organs must provide a hospitable environment/soil for the metastasized tumors to grow

Question 42

Factors that Affect the Location and Efficiency of Metastasis

Answer 42

- Chemokines in the specific tissue that attract the tumor cells to the site - The differentiation, developmental and growth conditions of the tissue (mitogens and growth factors) - The nature of the trafficking circulation (blood and lymphatic) systems in the specific organ