The angiogenesis process, the factors regulating it, different assays for it, a little about tumour angiogenesis, the drugs and new therapeutic approaches towards inhibiting or augmenting the process.

1. ANGIOGENESIS
IN
HEALTH AND DISEASE
DR DIBYAJYOTI PRUSTY
1ST Year PG
Moderator:
Dr Pranati Mohanty
Asst. Professor
Department of Pathology
SCB Medical College, Cuttack

2. Cross section of blood vessels
2

3. OVERVIEW OF VESSEL FORMATION

5. TYPES OF ANGIOGENESIS

6. The process of angiogenesis occurs as an orderly series of
events :
1. Vasodilation and increased permeability
2. Separation of pericytes
3. Migration of endothelial cells
4. Proloferation of endothelial cells behind the leading
‘tip’ cell
5. Remodelling in to capillary tubes
6. Recruitment of periendothelial cells
7. Suppression of endothelial proliferation and deposition
of basement membrane
Folkman J, D’Amore PA. Blood vessel formation: what is its molecular basis?
Cell 1996;87:1153-1155.

7. Angiogenic
stimulators

8. REGULATION OF ANGIOGENESIS
METABOLIC FACTORS:
a. Capillary growth is proportional to metabolic activity
b. Increase in metabolic activity stimulates blood vessel growth
c. Decrease in metabolic activity causes vascular regression ;
d. Over oxygenation often leads to capillary rarefaction in sedentary
muscles by auto regulatory vasoconstriction of arterioles.
e. Long term increase in BP leads to vascular rarefaction by auto
regulatory vasoconstriction mechanism.
f. OXYGEN is the master signal in growth regulation of vascular
system. Chronic exposure to hypoxia leads to increase in arterial
diameter .
g. Role of Adenosine:
Vasodialatory property- restores balance between O2 demand and
supply
Serve as a negative signal to maintain tissue oxygenation in normal
range.

9. MECHANICAL FACTORS:
1. PHYSICAL FORCES acting on the wall of blood
vessels
Shear stress is sensed by the endothelium. This stress acts
as a proangiogenic factor.
REGULATION OF ANGIOGENESIS

10. 2.MECHANOSENSORY MECHANISMS
 EpNaC Proteins : Localised in ECs, Smooth muscle cell
membranes.
 Both these cells express alha-, beta-, and gamma-
subunits of EpNaC.
 Mechanosensory complex formed by
- endothelium & smooth muscle cell(cytoskeletons and
EpNaCs)
- extra cellula matrix
 It plays a critical role in angiogenic process- as
mechanosensor for migration of endothelial cells and
smooth muscle cells.
 Specific EpNaC inhibitor used- Benzanil- to prevent
angiogenesis.

11. ROLE OF PERICYTES
 Pericytes are single layer of periendothelial smooth
muscle cells that modulate endothelial cell function.
 Regulate vascular function:
-vascular diameter
-vascular permeability
- endothelial survival

12. FACTORS AND RECEPTORS

13.  VASCULAR ENDOTHELIAL GROWTH
FACTOR
 ANGIOPOETINS
 BASIC FIBROBLAST GROWTH FACTOR
 PLATELET DERIVED GROWTH FACTOR
 EPIDERMAL GROWTH FACTOR
 TRANSFORMING GROWTH FACTOR-
ALPHA
ACTIVATORS OF ANGIOGENESIS

14. VASCULAR ENDOTHELIAL GROWTH FACTOR
 Glycoproteins consisting of A-, B-, C-, D-, E- forms and
Placental Growth Factor (PlGF)
 Within the six subtypes multiple isoforms exists
 Loss of even a single VEGF-A allele results in embryonic
lethality
 Angiogenesis is primarily mediated through interaction of
VEGF-A with VEGFR-2
VEGF RECEPTORS
3 types of receptors- VEGFR-1, VEGFR-2 (KDR, Flk-1), VEGFR-3
Tyrosine kinases

16. VEGF production is under control of :
hypoxia-inducible transcription factors(HIFs)
 VEGF receptor expression is up-regulated under:
hypoxic or ischemic conditions;
(30-fold within minutes)

17. VEGF is a major player in angiogenesis initiation:
it cause:
1. Endothelial cell survival
2. Stimulates endothelial division, induce
locomotion/migration
3. Induce the expression of proteases and receptors
4. Prevent endothelial cell apoptosis
5. Increase in vascular permeability by upregulating
second messengers such as NO

19. Fibroblast growth factor
 Basic Fibroblast growth factor (bFGF) family are also
potent inducers of angiogenesis. The effects of FGFs are
mediated via high-affinity tyrosine kinase receptors.
 Cellular responses mediated by FGFs include
 cell migration
 proliferation
 differentiation
Platelet-derived growth factor
 The platelet-derived growth factor (PDGF) regulates
 the recruitment of PERICYTES and
 smooth muscle cells
required for further stabilization of the new capillaries

20. ENDOSTATIN
 Produced by proteolytic cleavage of collagen
ANGIOSTATIN
 Produced by proteolytic cleavage of plasminogen,
THROMBOSPONDIN-1(TSP-1)
Adhesive glycoprotein in matrix.
P53 up regulates TSP-1
INHIBITORS OF ANGIOGENESIS

21. CELL ADHESION MOLECULES(CAM)
1. Integrins, cadherins
2.Vascular cell adhesion molecule-1,
3.P-selectin and E-selectin
Integrins are expressed at high levels in :
tumor vasculature and wound-healing tissues ,
but at extremely low levels in normal blood
vessels. 21

22. PROTEASES
Matrix metalloproteases Plasminogen activator(PA) /
(MMPs) plasmin system
PAs activate the plasminogen
degrade different into plasmin, which degrades
protein types several components of
extracellular matrix (ECM)
 Both PAs and MMPs are secreted together with their
inhibitors.
It ensures a stringent control of local proteolytic activity.
22

24. Why Tumors require Angiogenesis?
A, Tumors less than 1 mm3
receive oxygen and nutrients by
diffusion from host vasculature.
B, Larger tumors require new
vessel network. Tumor secretes
angiogenic factors that
stimulate migration,
proliferation, and neovessel
formation by endothelial cells
in adjacent established vessels.
C, Newly vascularized tumor
no longer relies solely on
diffusion from host vasculature,
facilitating progressive growth.

25. STRUCTURE AND FUNCTION OF TUMOR VESSELS:
Chaotic architecture and heterogeneous blood flow that leads
to abnormal growth
 Excessively dilated blood vessels
 Extreme corkscrew like tortuosities
 Lack of pericyte support or abnormal pericytic function:
Permeability strongly increased
-fenestrae
-enlarged Junctions
 No functional lymphatics inside the tumor
-enlarged in surrounding,
-increases metastasis

26. CHAOTIC ORGANIZATION OF TUMOR-ASSOCIATED
VASCULATURE

27. ANGIOGENESIS ASSAYS
 1. In-vitro assays
-Proliferation assay
-Migration assay
-Tube formation assay
-Rat and mouse aortic ring assay
 2. In-vivo assays
-Corneal angiogenesis assay
-Chick choreoallantoic membrane angiogenesis assay
-Matrigel Plug assay
The difficulties faced:
-ECs are heterogeneous
-In-vitro conditions rarely reflects in-vivo environment

28. IN HEALTH AND DISEASE

29. ANGIOGENESIS:
ABNORMAL OR EXCESSIVE
ORGANS INVOLVED DISEASES
Numerous Organs Cancers
Infectious diseases
Autoimmune disorders
Blood vessels Vascular malformations,
DiGorge syndrome
Atherosclerosis,
Adipose tissue Obesity
Skin Psoriasis
Scar keloids, Pyogenic granuloma

30. ORGANS INVOLVED DISEASES
Eye Diabetic retinopathy
Retinopathy of prematurity
Choroidal neovascularisation
Lung Primary pulmonary hypertension
Asthma , Nasal polyps
Gastro-Intestinal
Bone, joints
Reproductive
system
Inflammatory bowel disease,
Ascitis,
Synovitis , Osteomyelitis
Osteophyte formation
Endometriosis , Uterine bleeding,
Ovarian cyst

31. ANGIOGENESIS:
INSUFFICIENCY OR VESSEL REGRESSION
ORGANS INVOLVED DISEASES
Blood vessels Atherosclerosis, Hypertension,
Diabetic ischemic limbs
Nervous system Alzheimer’s disease
Amyotrophic lateral sclerosis
Diabetic neuropathy
Stroke
Gastrointestinal Gastric or oral ulcerations
Chrohn’s disease

32. R
ORGANS INVOLVED DISEASES
Skin Hair loss
Skin purpura, Telangeactasia
Reproductive system Pre-eclampsia, Menorrhagia
Lungs
Kidney
Bones & joints
Neonatal respiratory distress
Emphysema
Pulmonary fibrosis
Nephropathy
Osteoporosis
Impaired fracture healing

33. Heterotypic interactions as targets for therapeutic intervention

36. CURRENT ANGIOGENIC INHIBITORS IN CLINICAL
USE AND CLINICAL TRIALS
 Bevacizumab (Avastin™)
 Sunitinib (Sutent™)
 Sorafenib (Nexavar™)
 Cederanib (Recentin™ - AZD- 2171)
Many others in development

37. FUTURE DIRECTIONS-VEGF-TRAP
 Composite decoy receptor based on VEGFR-1 and
VEGFR-2 fused to a human Fc segment of IgG1 that
binds VEGF
 Decreases free VEGF to bind to receptors and prevent
vessel growth
 FDA approved for neovascular macular degeneration

38. THE CHALLENGES!!!
 Proangiogenic growth factor redundancy
 Selection of hypoxia-resistant cells
 Co-option of normal organ vasculature
 Vascular remodelling
 Angiogenesis contributing to growth of ‘liquid’
hematologic malignancies, not just solid tumors
 Circulating endothelial progenitor cells or
precursor cells
 Diverse array of molecular mediators of
angiogenesis

39. CONCLUSION
 Historically, angiogenesis is only implicated in cancer,
arthritis, psoriasis…etc. In recent years it is evident that
excessive, insufficient or abnormal angiogenesis
contributes many more disorders.
 Both pro- & antiangiogenic therapy with single
angiogenic molecule is more challenging than
anticipated
 Monotherapy with a single angiogenic inhibitor may not
suffice to combat the angiogenic factors produced by
cancer cells
 Understanding the complex interplay of molecular
signals in more integrated manner to develop efficient
and safe therapies.

40. INHIBITION OF BLOOD VESSEL GROWTH WITHIN A
TUMOR COULD PROLONG TUMOR DORMANCY
AND IMPROVE SURVIVAL OF PATIENTS WITH
MINIMAL TOXICITY.