Unit 6- Tumor Angiogenesis & Anti-Angiogenic Therapy

Question 1

Although previously believed that tumors result from the homogenous proliferation of tumur cells (false), what is true?

Answer 1

• tumor microenvironment
  > tumors are complex heterogenous multi-cellular structures

Question 2

What is angiogenesis?

Answer 2

• formation of new blood vessels from preexisting vasculature
• involved in physiological/ pathological processes

Question 3

What are the layers (anatomy) of a blood vessel?

Answer 3

lumen > endothelial cell lining > basal lamina > elastin fibers > smooth muscle > loose connective tissue

Question 4

What are the layers (anatomy) of a capillary?

Answer 4

lumen > endothelial cell lining > basal lamina
- pericytes

Question 5

What are the 4 steps in angiogenesis?

Answer 5

1. Disruption of basal lamina of existing vessel
2. Migration of endothelial cells toward a chemotactic signal
3. Proliferation of endothelial cells
4. Formation of tubes/ maturation of new vessel

Question 6

What are the 2 main types of angiogenesis?

Answer 6

Sprouting Angiogenesis
- endothelial cells activated/ protease degradation basal lamina
- EC proliferation/ migration > vessel stabilization

Intussusceptive Angiogenesis
- faster/ no extensive proliferation of endothelial cells
- ECs make transluminal bridge > reorganization of vessel
- bifurcation/ vessel splitting due to ↑ pressure

Question 7

What are examples of physiological angiogenesis?

Answer 7

• embryo development
• wound healing
• female reproductive system

Question 8

What is tumor/ pathologic angiogenesis?

Answer 8

• tumors must recruit own blood supply > growth/ metastasis
  > get nutrients from blood/ for metastatic spread

Question 9

What stimulates tumor angiogenesis?

Answer 9

“Angiogenic Switch”
- when tissue ↑ secretion of pro-angiogenic growth factors
- usually concurrent with ↓ expression of anti-angiogenic factors

Question 10

What is Folkman’s hypothesis?

Answer 10

• tumors smaller than 2 mm3 can exist with limited blood supply
• once beyond 2mm3 they must recruit new blood vessels
  (O2 can passively diffuse from vessel > tissue under 2mm)

Question 11

What is the most potent pro-angiogenic factor?

Answer 11

VEGF = vascular endothelial growth factor

Question 12

How does VEGF mediate angiogenesis?

Answer 12

1. Upstream activators stimulate VEGF production
2. VEGF binds to specific receptors on endothelial cells
3. Angiogenesis is primarily mediated through interaction of VEGF-A with VEGFR-2
4. Other variants of VEGF ligand/ receptor have a secondary role in this process

Question 13

How is angiogenesis primarily mediated?

Answer 13

• primarily through interaction of VEGF-A/ VEGFR-2

Question 14

What kind of receptor is VEGFR?

Answer 14

Tyrosine kinase

Question 15

What is the role of the ECM in tumor angiogenesis?

Answer 15

• in initial phase of angiogenesis > proteases degrade basement membrane/ destabilize ECM
• allows migration of endothelial cells toward chemotactic signal/ tumor
• integrin helps pull the sprouting new blood vessel forward

Question 16

After endothelial cell migration/ proliferation to tumor, what helps stabilize the new blood vessel?

Answer 16

• integrin molecules pull the sprouting new blood vessel forward
• endothelial cells deposit a new basement membrane/ secrete growth factors like PDGF (attract supporting cells to stabilize the new vessel)

Question 17

What is the role of hypoxia in tumor angiogenesis?

Answer 17

• cells closest to vasculature have sufficient blood supply
• O2 supply diminishes further from blood vessels
• as O2 supply ↓ cells become hypoxic/ necrotic
• tumor is not homogenously perfused (some areas high/low O2)

Question 18

How do hypoxic signals influence O2/nutrient delivery?

Answer 18

Hypoxia > HIF-1 activated > blood vessel growth
- hypoxic signals elicit mechanisms to ↑ nutrient/ O2 delivery

Question 19

What is the role of HIF-1/ when is it active?

Answer 19

non-hypoxic (normal) conditions > VHL tumor suppressor targets HIF-1 for rapid ubiquitination (degraded as not in use)

hypoxic conditions > HIF-1 recognizes/ binds to genes that mediate angiogenesis like VEGF

Question 20

What are the characteristics of tumor blood vessels?

Answer 20

• very different than normal blood vessels (chaos)
• hyperactivated VEGF > uncontrolled perfusion > malformed vessels
• blind ends/ vessel breaks/ AV shunts ext
• ↑ permeability/ immaturity

Question 21

Dr. Petriks lab uses what model of tumor angiogenesis?

Answer 21

EOC = Epithelial ovarian cancer
- VEGF is ↑ in tumor-bearing mice

Question 22

What are microstructural changes in tumor vessels?

Answer 22

• disrupted basal membrane
• absence of pericytes/ smooth muscle cells
• fenestrated endothelial cells (spaces between them)

Question 23

Why is poor vasculature in tumors a problem?

Answer 23

• a lot of fluid leakage/ accumulation between cells > ↑ pressure
• poor perfusion > hypoxia > necrosis
• leaky/ poor blood flow > hard to get drugs into tumors

Question 24

What is anti-angiogenic therapy?

Answer 24

• use of “angiogenesis inhibitors” to destroy tumor blood vessels/ starve the tumor

Question 25

What are 3 anti-angiogenic strategies?

Answer 25

1. Drugs that stop new blood vessels from sprouting - true angiogenesis inhibitors 2. Drugs that attack a tumors established blood supply - vascular disrupting agents 3. Drugs with a dual effect (attack cancer and blood vessel cells)

Question 26

What is the difference between tumor vascular disrupting agents/ anti-angiogenic agents?

Answer 26

Vascular Disrupting Agent > attack a tumors established blood supply - cause extensive tumor necrosis - activity primarily at tumor core of large tumor masses Anti-Angiogenic Agent > stop new blood vessel sprouting - interfere with new blood vessel formation - prevent tumor growth/ limit metastatic potential - activity primarily at tumor periphery of small tumor masses

Question 27

What are the basic strategies of angiogenesis inhibition?

Answer 27

- use anti-angiogenic compounds - inhibit pro-angiogenic factors

Question 28

What are some specific angiogenesis inhibitors?

Answer 28

Avastin/ Celebrex/ Angiostatin/ Endostatin

Question 29

What is Avastin?

Answer 29

- humanized antibody that binds VEGF, neutralizing its angiogenic effects (preventing interaction with VEGFR) - inhibits VEGF, but VEGF is needed to make blood vessels! side effects > GI perforation (↓ blood flow) > necrosis > sepsis - impaired wound healing/ hemorrhage/ expensive

Question 30

What is Celebrex?

Answer 30

- non-steroidal/ anti-inflammatory drug for osteoarthritis = COX2 inhibitor - overexpression of COX2 induces VEGF production - COX2 ↑ vascular permeability/ ↑ endothelial cell proliferation - through COX2 inhibition, Celebrex inhibits formation of new vessels - ↓ expression of survivin (tumor survival protein)

Question 31

What is angiostatin?

Answer 31

- endogenous angiogenesis inhibitor - produced by cleavage of plasminogen - inhibits endothelial cell migration/ proliferation/ initiates apoptosis

Question 32

How do VEGF-R2 inhibitors work?

Answer 32

- inhibit receptor directly (not VEGF-a ligand)

Question 33

What is vascular targeting? (VTA's = vascular targeting agents)

Answer 33

- vascular targeting attacks existing tumor vessels (unlike anti-angiogenic approaches that inhibit further vessel development)

Question 34

What is Thrombospondin-1?

Answer 34

TSP-1 = endogenous inhibitor of angiogenesis - causes endothelial cell apoptosis/ ↓ migration/ inhibited chemotaxis - also directly inhibits VEGF expression

Question 35

How is Jim's lab researching Thrombospondin-1?

Answer 35

TSP-1- has 5 domains/ too big to be a therapeutic compound 1 main domain > inhibits VEGF-induced angiogenesis/ EC migration - 3TSR induces ovarian tumor/ endothelial cell death BUT... - induces tumor regression/ vessel normalization/ ↑ tumor perfusion - exploit this to ↑ chemo uptake/ improve drug therapy > combination therapy