Mediadores de la angiogénesis

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Biochimica et Biophysica Acta 1654 (2004) 51–67

Review

Endothelial cell integrins and COX-2:

mediators and therapeutic targets of tumor angiogenesis

Curzio Ruegga,b,*, Olivier Dormonda,1, Agnese Mariottia

aCentre Pluridisciplinaire d’Oncologie (CePO), University of Lausanne Medical School, CH-1011 Lausanne, SwitzerlandbNCCR Molecular Oncology, Laboratory of the CePO, Swiss Institute for Experimental Cancer Research (ISREC), 155 Chemin des Boveresses,

CH-1066 Epalinges s/Lausanne, Switzerland

Received 16 April 2003; received in revised form 3 September 2003; accepted 3 September 2003

Abstract

Vascular integrins are essential regulators and mediators of physiological and pathological angiogenesis, including tumor angiogenesis.

Integrins provide the physical interaction with the extracellular matrix (ECM) necessary for cell adhesion, migration and positioning, and

induce signaling events essential for cell survival, proliferation and differentiation. Integrins preferentially expressed on neovascular

endothelial cells, such as aVh3 and a5h1, are considered as relevant targets for anti-angiogenic therapies. Anti-integrin antibodies and small

molecular integrin inhibitors suppress angiogenesis and tumor progression in many animal models, and are currently tested in clinical trials as

anti-angiogenic agents. Cyclooxygense-2 (COX-2), a key enzyme in the synthesis of prostaglandins and thromboxans, is highly up-regulated

in tumor cells, stromal cells and angiogenic endothelial cells during tumor progression. Recent experiments have demonstrated that COX-2

promotes tumor angiogenesis. Chronic intake of nonsteroidal anti-inflammatory drugs and COX-2 inhibitors significantly reduces the risk of

cancer development, and this effect may be due, at least in part, to the inhibition of tumor angiogenesis. Endothelial cell COX-2 promotes

integrin aVh3-mediated endothelial cell adhesion, spreading, migration and angiogenesis through the prostaglandin-cAMP-PKA-dependent

activation of the small GTPase Rac.

In this article, we review the role of integrins and COX-2 in angiogenesis, their cross talk, and discuss implications relevant to their

targeting to suppress tumor angiogenesis.

D 2004 Elsevier B.V. All rights reserved.

Keywords: Integrin; Cell adhesion; Signaling; Angiogenesis; Cancer; Therapy

1. Introduction ing antibodies efficiently suppressed angiogenesis and

Vascular biology and angiogenesis is one of the fastest

developing areas in biomedical research. A main reason for

the enormous interest in angiogenesis research is due to the

potential for therapeutic interventions in angiogenesis-de-

pendent diseases, such as cancer, chronic inflammation and

proliferative retinopathies. Vascular integrins emerged as

critical mediators and regulators of developmental and

postnatal angiogenesis. In preclinical studies, low molecular

weight integrin inhibitors and anti-integrin function-block-

0304-419X/$ - see front matter D 2004 Elsevier B.V. All rights reserved.

doi:10.1016/j.bbcan.2003.09.003

* Corresponding author. NCCR Molecular Oncology, Laboratory of the

CePO, Swiss Institute for Experimental Cancer Research (ISREC), 155

Chemin des Boveresses, CH-1066 Epalinges, s/Lausanne, Switzerland.

Tel.: +41-21-692-5853; fax: +41-21-692-5872.

E-mail address: [email protected] (C. Ruegg).1 Current address: Department of Internal Medicine, Centre Hospitalier

Universitaire Vaudois (CHUV), CH-1011 Lausanne, Switzerland.

inhibited cancer progression. aVh3 was the first vascular

integrin targeted to suppress tumor and ischemia-induced

angiogenesis. Antagonists of integrin aVh3 are currently

tested in clinical trials as anti-angiogenic agents. Recent

experiments, however, have provided evidence that the role

of aVh3 integrin in regulating angiogenesis may be more

complex than previously appreciated. Besides mediating

physical cell adhesion, integrins also initiate signaling

events, which, alone or in combination with growth factor

receptor-mediated signals, promote essential cellular func-

tions, such as cell migration, proliferation, survival and

differentiation. Intracellular signaling events mediated by

integrins may therefore represent valid alternative targets the

therapeutic inhibition of angiogenesis. In the light of new

developments in the field, a critical reevaluation of the role

of integrins in angiogenesis and of the strategies aimed at

their inhibition is necessary.

C. Ruegg et al. / Biochimica et Biophysica Acta 1654 (2004) 51–6752

2. Tumor progression depends on tumor angiogenesis

Malignant cell transformation and tumor progression

result from the accumulation of multiple mutations in the

cell DNA, leading to the disruption of critical cellular

signaling pathways and molecules involved in the regulation

of normal vital cellular functions [1]. Cellular transforma-

tion alone, however, is not sufficient to give rise to a

clinically relevant tumor. In order to survive and grow, a

cancer cell largely depends on the ‘support’ from the

surrounding normal tissue (the so-called ‘tumor stroma’).

The formation of a tumor vasculature is an essential stromal

reaction supporting tumor growth [2–5]. In the absence of

tumor angiogenesis, the tumor mass stabilizes at a volume

of a few cubic millimeters (equivalent to approximately 105

to 106 cells) as a result of a balance between cell prolifer-

ation and apoptosis [6]. In addition, tumor vessels favor the

escape of tumor cells into the blood circulation, which

constitutes the initial step of metastatic spreading [7,8].

3. Cellular and molecular mechanisms of blood vessel

formation

Over the past two decades, thanks to contributions from

developmental biology, mouse genetics, cell and molecular

biology, experimental pathology and drug discovery, we

have gained a detailed insight into many of the cellular and

molecular events that mediate and regulate blood vessel

formation. During embryonic development the differentia-

tion of vascular progenitor cells (i.e. the hemangioblast and

the angioblast) into mature endothelial cells gives rise to a

primitive but nevertheless functioning vascular plexus. This

process is called vasculogenesis [9–11]. The primitive

vascular plexus is then remodeled into a mature vascular

system through sprouting, trimming, intussusception and

hierarchical branching by a process globally referred to as

angiogenesis [9,12]. This vascular maturation process also

involves the recruitment of perivascular cells, also referred

to as pericytes [13], which confer mechanical stability and

contractility to the vessel wall, and provide essential factors

for endothelial cell survival [14,15]. During postnatal

angiogenesis, quiescent vascular endothelial cells are in-

duced to proliferate through the combined action of differ-

ent factors: the de novo expression of angiopoietin-2, an

endogenous antagonist of angiopoietin-1, which ‘releases’

the endothelial cells from the stabilizing effects of the

vessel wall, the local secretion or activation of vascular

growth factors (e.g. VEGF), and the inactivation or sup-

pression of endogenous anti-angiogenic molecules (e.g.

thrombospondins) [10,16–18]. In contrast to physiological

angiogenesis, tumor vessels do not undergo full matura-

tion. They frequently display incomplete endothelial cell

lining and tumor cells are often in direct contact with the

circulation [19], they show disorganized recruitment of

pericytes [20], remain highly permeable and unstable,

and fail to generate a hierarchically branched vascular

network [21].

Besides sprouting angiogenesis, intussusception (i.e.

the internal division of a capillary plexus) and bone

marrow-derived endothelial cell precursors (ECP) were

shown to contribute to developmental and tumor angio-

genesis [22–25].

Many molecules mediating and modulating physiologi-

cal and pathological angiogenesis have been identified and

characterized in recent years. A comprehensive review of

these molecules and mechanisms is beyond the scope of this

article and some excellent review articles have been recently

published [10,16,18,26]. Here we briefly summarize the

major classes of molecules involved in angiogenesis.

x Growth factors and growth factor receptors, such as

Vascular Endothelial Growth Factors (VEGF A, B, C, D

and E) and VEGF-Receptor (R) 1, 2, and 3; Fibroblast

Growth Factors (FGFs) and FGF-Rs [27,28];

x Adhesion molecules of the integrin and cadherin families,

such as aVh3, a5h1 and VE-cadherin [29,30];

x Extracellular matrix (ECM) proteins, such as fibronectin,

collagens, and laminins [29,31,32];

x Remodeling and morphogenic molecules and their

receptors, in particular Angiopoietins/Ties and Eph/

Ephrins [33,34];

x Proteinases such as matrix metalloproteinase (MMP) -2

and -9, plasminogen activators (u-PA and t-PA) and their

inhibitors (TIMPs and PAIs);

x Intracellular signaling molecules, most notably protein

kinases (e.g. Raf and Mitogen Activated Protein Kinase

(MAPK), Protein Kinase A (PKA), Protein Kinase B

(PKB/Akt)) and GTPases (e.g. Ras and Rho families)

[35–38];

x Transcription factors and regulators, including Hypoxia

Inducible Factor (HIF) -1a, Inhibitors of differentiation

(Id1/3), Nuclear Factor (NF)-nB and homeobox gene

products (e.g. Hox D3 and B3) [39–43].

4. Integrins as adhesion molecules

Integrins are heterodimeric cell surface adhesion recep-

tors formed by two noncovalently associated subunits, a

and h. There are 18 a and 8 h subunits which associate to

form 24 different heterodimers [44,45] (Fig. 1). a subunits

are 1000–1200 amino acids long, while h subunits are

slightly shorter (around 750 amino acids long). Integrin

subunits consist of a large extracellular domain, a single

transmembrane domain and a short cytoplasmic tail, except

for the h4 subunit which contains over 1000 intracellular

residues. Integrins are the most important receptors for

ECM proteins, such as fibronectin, laminins, collagens or

vitronectin. Integrins are promiscuous and redundant recep-

tors: i.e. one integrin can bind several different ligands, and

many different integrins can bind to the same ligand. The

Fig. 1. The integrin family of adhesion receptors and their main ligands. There are 18 a and 8 h mammalian subunits which assemble to form 24 different

heterodimers. Although integrins are the main receptors for ECM proteins, they can also bind cellular counter receptors (e.g. ICAMs, VCAM), soluble

molecules (fibrinogen, complement components) and pathogens (e.g. viruses and bacteria). Based on sequence alignments, functional properties or expression

patterns, one can define different integrin subfamilies: the RGD-binding integrins (blue), the laminin-binding integrins (red), the leukocyte integrins (black), the

I-domain-containing integrins (green), and the a4 sub-family integrins.

C. Ruegg et al. / Biochimica et Biophysica Acta 1654 (2004) 51–67 53

most promiscuous integrin is aVh3 which can bind to over

a dozen of different ligands, including fibronectin, vitronec-

tin, thrombospondin, tenascin-C, fibrin, von Willenbrand

Factor, osteopontin and denatured collagen I [30,44,46]

(Table 1). The reason and significance for integrin redun-

dancy and promiscuity are not completely clear.

Promiscuity may be of advantage when the cellular

responses needed (e.g. survival or migration) in a particular

context are more relevant than the nature of the ECM

protein eliciting them. This may be the case during would

healing or tissue remodeling, where resident and infiltrating

cells have to rapidly react and adapt to changes in the

composition of the ECM. Several plasma proteins (e.g.

fibrinogen-fibrin, vitronectin) extravasate and are deposited

following an increase in vascular permeability, while other

ECM proteins (e.g. tenascin-C, thrombospondin) are syn-

thesized in situ by activated resident cells and infiltrating

inflammatory cells. In addition, preexisting proteins (e.g.

collagens, laminins) are proteolytically processed by protei-

nases. It may therefore be more efficient to ‘catch’ all these

new ligands with one single integrin with broad specificity,

than expressing several different integrins, each specific for

one or a few ligands. Indeed, aVh3 is found expressed in

cells that migrate through different tissues, such as neutro-

phils, monocytes and lymphocytes [44]. aVh3 expression

in resident cells is up-regulated during tissue remodeling,

such as in cancer and inflammation, and many aVh3ligands are abundant in tissues undergoing remodeling

[44–47]. Redundancy may reflect the need to transduce

different signaling and cellular responses from the same

ECM. For example, integrin a5h1 binding to fibronectin

suppresses cell migration [48], while aVh6-mediated fibro-

nectin binding promotes it [49]. This notion is also consis-

tent with the observation that constitutive deletion in

embryonic stem cells of individual integrins that bind to a

common ligand (i.e. a3h1, a4h1, a5h1, a8h1, aVh1,aVh3, aVh5, aVh6, a4h7, aVh8—fibronectin) gives rise

to highly divergent and non-overlapping developmental

phenotypes. For example, constitutive deletion of the fibro-

nectin gene results in early embryonic lethality during

development [50,51]. The phenotypes observed in mice

lacking any of the individual fibronectin receptors are

highly different, and none of them alone reproduce the

fibronectin-null phenotype [52]. One should also consider

the possibility that integrin redundancy in cells at a given

time point may be restricted due to the differential spatio-

temporal patterns of expression of individual integrins.

5. Integrins as signaling molecules

Integrin-mediated signaling events are essential for stable

cell adhesion, cell spreading, migration, survival, prolifera-

tion and differentiation [53,54]. Integrins, however, have no

intrinsic enzymatic activity, and to transduce signals they

depend on the recruitment of cytoplasmic structural (e.g. a-

actinin, talin, vinculin) and signaling (e.g. focal adhesion

kinase (FAK), paxillin and Src family kinases) proteins and

the assembly of characteristic structures named focal con-

tacts and focal adhesions [55]. Many signaling pathways

activated by integrins are also activated by growth factor

receptors. The cross talk between integrins and growth factor

receptors is believed to provide enhanced specificity and

control over many cellular events, compared to signaling

Table 1

Vascular integrins in angiogenesis

Integrin Major ECM ligands Gene

deleted

Vascular phenotype in mice

with constitutive gene deletion

Suppression of tumor angiogenesis

by pharmacological inhibitors

a1h1 CO, LM a1 No defects in vascular development;

reduced tumor angiogenesis

Yes; antibodies

a2h1 CO, LM a2 No defects in vascular development Yes; antibodies,

inhibitor of expression

a3h1 LM, TSP a3 Perinatal lethality; defects in kidney,

lung, skin but no vascular defects

?

a4h1 FN a4 Lethal at E 11–14; placenta fusion

defect and coronary arteries defect

No/?

a5h1 FN, Fibrin a5 Lethal at E 10; vasculogenesis

but no maturation/angiogenesis

Yes; antibodies, small

molecular inhibitors

a6h1 LM a6 Lethal at birth; no defects

in vascular development

?

a8h1 FN, TN-C a8 Partial embryonic lethality; no

defects in vascular development

?

a9h1 TN-C a9 Lethal at birth; defects in

large lymphatic vessels

?

aVh1 FN, VN (1)

h1 Lethal at E 5.5; failure of

organizing the embryonic inner mass

aVh3 FN, VN, LM, FB, Fibrin, TSP,

TN-C, vWF, dCO, OPN, MMP-2,

Del-1, BSP, FGF-2, thrombin

h3 No defects in vascular development;

enhanced postnatal angiogenesis

Yes; antibodies, small


aVh5 VN, Del-1 h5 No defects in vascular development;

enhanced postnatal angiogenesis?

Yes, but often targeted

together with aVh3 small


a6h4 LM h4 Lethal at birth; no vascular

defects in development

?

aVh8 VN h8 Lethal at E 12-perinatal; vascular

defects in the placenta and brain

?

aV Lethal at E 12-perinatal; vascular

defects in the placenta, brain and intestine

Vascular integrins in developmental and postnatal angiogenesis. Endothelial cells express at least 13 different integrins. Disruption of the a5 gene causes severe

vascular defects during early development. Disruption of the av or h8 gene causes mild defect in vascular development, and most mice undergo extensive

vasculogenesis and angiogenesis. Mice lacking the a4 gene die of cardiac and coronary malformations. Deletion of any of the other a subunits causes no or

only minor vascular defects (e.g. a3). Mice deficient for the h3 (Fh5) subunit have enhanced postnatal angiogenesis and tumor growth, while a1 subunit-null

mice have reduced tumor angiogenesis. The integrins a1h1, a2h1, a5h1 and aVh3 are up-regulated during tumor angiogenesis. Abbreviations: CO, collagen;

Del-1, Developmental locus-1 LM, laminin; VN, vitronectin; TNC, Tenascin-C; TSP, thrombospondin; EL, elastin; FN, fibronectin; OPN, osteopontin; FB,

fibrinogen; vWF, von Willenbrand factor; MMP, matrix metalloproteinase; dCO, denatured collagen. BSP, Bone Sialo Protein. (1), testing of aVh1 function is

not possible by gene deletion experiment. ?, not known. The list of the integrin ligands is limited to the major and best characterized ones.


from one receptor class only. Here we will briefly review the

four main signaling pathways activated by integrins which

are relevant to angiogenesis (Fig. 2). For a comprehensive

overview of integrin-mediated signaling, we refer the reader

to a number of recent reviews [46,53,54,56–59].

� MAPK. Stimulation of ERK by endothelial-cell integ-

rins is mediated by either FAK, or the Src-family kinases

Fyn and Yes. In the first case, recruited FAK

autophosphorylates at 397Y and associates with c-Src

or Fyn, which further phosphorylates it at 925Y.

Phosphorylated FAK binds to the Grb2–Sos complex

and activates the Ras–ERK cascade [60,61]. In the

second case, Fyn or Yes, activated by some h1 and aV

integrins, recruit Shc through their SH3 domain and

phosphorylates it, thus promoting Shc association with

the Grb2–Sos complex [62,63]. Activated ERK enters

the nucleus to promote transcription [64]. The MAPK

pathway is critically involved in controlling prolifer-

ation, survival, and migration of endothelial cells.

Angiogenesis in the chorio-allantoid membrane (CAM)

model requires two waves of ERK activation, the first

one being dependent on FGF-2, and the second one on

aVh3 integrin ligation [65].

� PI-3K-PKB/Akt. Integrins can activate the serine/

threonine kinase PKB/Akt in a PI-3K-dependent

manner through FAK and integrin-linked kinase (ILK)

[66,67]. For activation, PKB/Akt requires phosphatidyl

inositol-3 phosphate-dependent translocation to the cell

membrane and phosphorylation at two sites, 308T and473S [68]. Activation of PKB/Akt by VEGF [69],

insulin [70], or angiopoietin-1 [71] only occurs under

conditions of integrin-dependent adhesion [72]. PKB/

Akt-dependent signaling controls many essential cel-

lular processes such as cellular metabolism, tran-

scription, cell proliferation, migration and survival

βα

Fig. 2. Signaling pathways activated by integrins. Integrin ligation activates four major signaling pathways relevant to angiogenesis: the Rac GTPase, the

MAPK, PI-3K-PKB/Akt and the NF-nB pathways. These pathways generate essential signals for endothelial cell migration, proliferation, and survival (see text).


[37,73,74]. PKB/Akt is believed to promote endothelial

cell survival and angiogenesis via the phosphorylation

and inactivation of the pro-apoptotic molecules Bad,

Bax, caspase 9, and the Forkhead (FRKH) transcription

factor [75], the stimulation of nitric oxide (NO)

production [76,77], and the inhibition of the stress-

activated kinases p38 and JNK [78]. Regulation of cell

migration appears to implicate a cross talk with Rho-

family proteins. In migrating cells, PKB/Akt localizes

to the leading edge [79], a site of high Rac activity

[80]. In endothelial cells PKB/Akt has been reported to

be both upstream and downstream of Rac and Cdc42

[81–83]. Endothelial cell migration induced by VEGF-

R2-stimulation involves integrin activation via a PI-3K

and PKB/Akt-dependent signaling pathway [84].

� Rho family GTPases. The small GTPases Rho, Rac and

Cdc42 participate in the regulation of actin polymer-

ization necessary for the formation of stress fibers, focal

adhesions, lamellipodia and filopodia [85], and modu-

late cell proliferation and gene expression [86,87].

Integrin ligation activates Rho proteins by promoting

GTP loading and translocation from the cytosol to the

membrane [88]. Rac activity is essential for endothelial

cell migration and angiogenesis [89,90] and this effect

requires the Rac effector protein p21 activated kinase 1

(PAK1). Inhibition of Rac or PAK1 function by

dominant negative constructs in endothelial cells in

vivo inhibits growth factor-induced angiogenesis

[90,91]. Activated Rac promotes recruitment of high-

affinity integrins aVh3 to lamellipodia [92] and the

establishment of new adhesions at the leading edge,

which are essential for directional cell migration.

Furthermore, integrin-specific activation of Rac by

a5h1 promotes endothelial cell proliferation on fibro-

nectin through the induction of cyclin D1 synthesis via a

Shc-FAK-SOS-PI-3K-dependent signaling [93].

� The NF-nB. NF-nB is an essential transcription factor

for the regulation of the immune and inflammatory

responses [94,95]. NF-nB also promotes cell survival in

response to pro-apoptotic stimuli by inducing the

expression of anti-apoptotic molecules [94]. NF-nBactivity is required for angiogenesis [41]. In vivo

inhibition of NF-nB activation suppresses retinal neo-

vascularization in a murine model of ischemic retinop-

athy [96] and blocks FGF-2-induced angiogenesis in the

Matrigel plug assay [97]. Ligation of endothelial cell

integrin aVh3 by osteopontin protects endothelial cells

against serum withdrawal-induced apoptosis through the

activation of NF-nB [98] and the expression of

osteoprotogerin [99]. Integrin a5h1-mediated adhesion

to fibronectin activates a NF-nB-dependent gene

expression program important for angiogenesis [97].

While integrin signaling events have been classically

considered to occur at focal adhesion sites, recent evidence

suggests that some integrins can localize to lipid rafts and

activate specific signaling pathways. With the exception of

the integrin a6h4 that can be directly targeted to rafts by

palmitoylation of the h4 subunit [100], integrin transloca-

tion probably depends on their interaction with proteins with

higher affinity for rafts. The association of integrins with

raft membrane glycoproteins has been reported, and there is

evidence suggesting that these interactions are functionally

relevant. For example, aVh3 associates with CD47 in rafts


and binding of thrombospondin to CD47 modulates the

function of aVh3 in melanoma and endothelial cells,

resulting in enhanced integrin-mediated cell spreading or

migration through the activation of heterotrimeric G pro-

teins and decreased intracellular cAMP levels [101,102].

Also, integrin-dependent activation of ERK seems to require

integrin signaling from lipid microdomains. Some integrins

can form a complex with caveolin-1 and the membrane raft-

localized Fyn and Yes. Upon ligation, they stimulate Src-

family kinases, thus promoting tyrosine phosphorylation of

Shc and ERK activation [63]. The role of raft dynamics in

regulating integrin signaling in quiescent and angiogenic

endothelial cells, however, is not known.

6. Integrins as endothelial cell survival factors

Integrins are emerging as essential determinants of en-

dothelial cell survival. Loss of integrin-mediated adhesion

and impaired spreading results in rapid onset of endothelial

cell apoptosis [103–105]. The mechanism of cell detach-

ment-induced death, a phenomenon also referred to as

anoikis [106], is still a matter of debate [46,107]. Two

complementary events may be at play: endothelial cells

may die because they are deprived of integrin-mediated

survival signals, and/or because unligated integrins may

actively initiate death-inducing signaling events. Indeed,

there is experimental evidence supporting both possibilities.

For example, integrin ligation has been reported to induce

expression of Bcl-2 [108–110], phosphorylation of the Rb

tumor suppressor protein [111], inhibition of p53 function

[109], activation of MEKK-1/ERK [112,113], PKB/Akt

[114,115] and NF-nB [98,116], all of which provide pro-

survival or anti-apoptotic signals. On the other hand, matrix

detachment and appearance of unligated integrins was

shown to induce caspase-8 activation and apoptosis [117–

119]. This mechanism has been referred to as integrin-

mediated death (IMD) [120]. Loss of integrin-mediated

adhesion also induces rapid JNK activation [108] and

JNK-1 and -2 activations were shown to mediate stress-

induced apoptosis in fibroblasts [121]. Cell detachment was

also reported to cause an increased production of Fas-ligand

and a diminished expression of c-Flip (an endogenous

inhibitor of caspase 8) resulting in Fas-mediated death

[122]. Although overexpression of dominant negative Fas-

associated death domain (FADD) protein was shown to

suppress anoikis [117,118], the role of cell surface death

receptors in initiating cell-detachment induced death re-

mains controversial.

In addition, cell adhesion promotes survival through

mechanical and shape effects [123–125]. The intact cyto-

skeleton is essential for maintaining the normal distribution

and function of death receptors [126,127], for PKB/Akt

signaling and expression of Bcl-2 [128], for the membrane

translocation and activation of Rac and Rho GTPases

[38,129,130], and for the sequestration of proapoptotic

factors such as Bim and Bmf [131,132]. It is therefore

conceivable that a ‘collapse’ of the cytoskeleton following

detachment results in the disruption of pro-survival path-

ways and the release of pro-apoptotic stimuli eventually

leading to cell death.

Taken together, cell death induced by the lack of sub-

strate adhesion is likely to involve multiple mechanisms:

loss of integrin-dependent pro-survival signals, induction of

integrin-mediated death, and imbalance of anti/pro-apopto-

tic events associated with the disruption of the cytoskeleton.

7. Integrins in vascular development

Endothelial cells have been shown to express at least 13

different integrins, depending on their state of development,

differentiation and function: a1h1, a2h1, a3h1, a4h1,a5h1, a6h1, a6h4, a8h1, a9h1, aVh1, aVh3, aVh5and aVh8 [133–139] (Table 1). Several of these integrins

(e.g. a1h1, a2h1, a3h1, a6h1) bind to ECM proteins

present in the basal membrane of mature vessels (e.g.

laminins or collagens), while other integrins (i.e. a5h1,aVh1, aVh3 and aVh5) bind to ECM proteins present at

sites of angiogenesis (e.g. fibronectin or vitronectin) [30,46]

(Table 1). Mouse genetic experiments in which integrin

genes were disrupted in embryonic stem cells and experi-

ments based on the use of integrin antagonists have pro-

vided compelling evidence that integrins are required for

embryonic vascular development and postnatal angiogene-

sis [52,140] (Table 1). Mice deficient for integrin a5h1have pronounced defects in posterior trunk and yolk sac

mesodermal structures, including the developing vascula-

ture, and die early during embryonic development [141].

a5h1-null embryos develop the extraembryonic and em-

bryonic vasculature but the overall blood vessel pattern

complexity is reduced, and blood vessels themselves are

greatly distended. This vascular phenotype correlates with a

decreased perivascular deposition of fibronectin, the main

ligand of a5h1 integrin [142]. Moreover, in embryonic

bodies formed from embryonic stem cells lacking a5h1integrin, there is a significant reduction in early capillary

plexus formation and maturation [142]. Taken together,

these findings point toward a critical role of a5h1 in

developmental angiogenesis. Embryos constitutively lack-

ing fibronectin, the main ligand of a5h1, die at E9-10 due

to defects in the mesoderm, neural tube and vascular

development [50,51]. Mice deficient for the integrin aV

subunits (thus lacking expression of aVh1, aVh3, aVh5,aVh6 and aVh8 heterodimers) have extensive vasculo-

genesis and angiogenesis and develop normally up to E9.5.

About 80% of the mice, however, die in mid gestation in

association with placental defects. The remaining 20% of

the mice are born alive with cerebral and intestinal malfor-

mations and cleft palates, exhibit intracerebral and intestinal

hemorrhages and die perinatally or shortly after [143].

These observations clearly indicate that aV integrins are


dispensable for most vascular development, while they

seem to be important in mediating vessels–parenchyma

interactions in the brain [144]. Mice lacking integrin

aVh3, aVh5 or both are viable and have no detectable

developmental defects [145–147]. The dispensable role of

aVh3 in developmental angiogenesis is consistent with the

observation that humans with a nonfunctional h3 subunit

suffer of severe bleeding due to impaired platelet aggrega-

tion, a condition known as Glanzmanns’s thrombasthenia,

but have normal angiogenesis [148,149]. Mice deficient for

the h8 subunit have very similar vascular defects as aV-

deficient mice [143,150], while mice lacking h1, h3, h5,and h6 integrin subunits have very distinct phenotypes

[147,151–153] suggesting that the phenotype observed in

the aV-null mice is due to the absence of the aVh8heterodimer. The integrin a9 subunit is expressed in lym-

phatic endothelial cells but not in vascular endothelial cells

[139]. Mice lacking a9h1 fail to generate large lymphatic

vessels, have edema of the chest wall and die perinatally

due to the accumulation of lipid-rich lymph in the pleural

cavity [154]. This phenotype is consistent with a role for

a9h1 in the normal development or homeostasis of lym-

phatic vessels, including the thoracic duct. The role of a9h1in postnatal lymphangiogenesis is not known.

8. Vascular integrins in tumor angiogenesis

The first integrin shown to be associated with postnatal

angiogenesis, including tumor angiogenesis, was aVh3(reviewed in Refs. [30,155,156]. aVh3 is highly expressed

in angiogenic endothelial cells in wound granulation tissue

and in malignant tumors but not, or to a much lower extent,

on quiescent endothelial cells [157–161]. Pharmacological

antagonists of aVh3 are potent inhibitors of angiogenesis.

An anti-aVh3 function-blocking monoclonal antibody

(LM609) [162] or a aVh3/aVh5-specific antagonistic cy-

clic peptide (EMD121974) [163] suppressed cornea vascu-

larization [164], hypoxia-induced retinal neovascularization

[165] and tumor angiogenesis in the mouse [158,166–168].

Quiescent and preexisting vessels were not perturbed by

treatment with aVh3 antagonists. Based on these observa-

tions, aVh3 was considered as an angiogenesis-promoting

integrin [29,155]. Small molecular integrin antagonists (e.g.

EMD121974 or Cilengitide) and a humanized LM609

blocking antibody (Vitaxin) are currently tested in clinical

trials for their therapeutic efficacy in human cancer. Phase I

trials demonstrated that these drugs are safe and well

tolerated [169].

The generation and analysis of mice lacking one or more

of the aV-containing integrins, however, raised some basic

questions on the role of aV integrins in angiogenesis and on

the interpretation of the experimental results obtained with

the pharmacological antagonists. aV-deficient mice lacking

all five aV-containing integrins undergo extensive develop-

mental vasculogenesis and angiogenesis [143], although

they die peri-/post-natally with brain and intestinal hemor-

rhages. Mice lacking aVh3, aVh5, or aVh3/aVh5 are

viable and display enhanced postnatal angiogenesis in

response to hypoxia, VEGF and tumors [147]. While these

discordant results (i.e. normal developmental angiogenesis

and excessive postnatal angiogenesis in aVh3-deficientmice vs. anti-angiogenic effects of pharmacological aVh3antagonists) are now accepted in the field, we are still

lacking a unifying understanding of the underlying mecha-

nisms involved. It was recently proposed that aVh3 may

have both positive and negative regulatory functions. The

net outcome (i.e. angiogenesis, vs. anti-angiogenesis) may

depend on the ligation state, the nature of the ligand, and the

presence or absence of additional extracellular signals [170].

In conclusion, it appears that the role of aVh3 in regulating

angiogenesis is more complex than previously appreciated

(see next paragraph).

There is mounting evidence that additional integrins, in

particular a5h1, contribute to tumor angiogenesis. a5h1 is

up-regulated in angiogenesis and blocking anti-a5 antibod-

ies suppressed VEGF-induced and tumor angiogenesis in

both chick embryo and murine models [171–174]. a5h1-blocking peptides have been reported [174,175], but their

use as anti-angiogenic agents has remained limited. Com-

bined administration of ATN-161, a a5h1 antagonist, and 5-

FU in a mouse model of tumor metastasis reduced liver

metastasis, consistent with the notion that a5h1 is a relevant

target for the inhibition of tumor angiogenesis [176].

Mice lacking integrin a1h1 develop normally, but have

reduced number and size of intra tumoral capillaries. This

effect was associated with an increased plasma level of

angiostatin due to enhanced MMP-7- and MMP-9-mediated

processing of circulating plasminogen [177]. Furthermore,

fibroblasts derived from a1-null mice have reduced MAPK

activation when plated on collagen, despite normal attach-

ment and spreading, due to deficient recruitment of the

adaptor protein Shc and Grb2 [177]. This observation raises

the possibility that a1-deficient endothelial cells may have a

decreased proliferation response in a collagenous environ-

ment. Strikingly, absence of a1h1 integrin causes a loss of

the feedback regulatory mechanism of collagen synthesis,

resulting in increased collagen deposition [178]. Mice

lacking integrin a2h1 develop normally, are fertile, and

have a decreased complexity in mammary gland branching

and deficient platelets adhesion to collagen, but otherwise

have no obvious anatomical abnormalities, including in the

vasculature [179,180]. These mice were not analyzed for

potential defects in postnatal tumor angiogenesis [181].

Integrins a1h1 and a2h1 are highly up-regulated by VEGF

in cultured endothelial cells, resulting in enhanced a1h1-and a2h1-dependent cell spreading on collagen I, and anti-

a1h1 and anti-a2h1 antibodies inhibited VEGF-driven

angiogenesis in vivo [182]. Moreover, combined adminis-

tration of anti-a1h1 and anti-a2h1 antibodies to mice

bearing squamous cell carcinoma xenografts resulted in

reduced tumor angiogenesis and tumor growth [183].


E-7820, an aromatic sulfonamide derivative, was reported

to block in vitro proliferation and tube formation of cultured

endothelial cells through the inhibition of a2 integrin

subunit expression. Oral administration of E-7820 inhibited

tumor-induced angiogenesis and human tumor growth in a

murine xenograft model [184].

In conclusion, genetic inactivation or pharmacological

inhibition experiments have identified integrins a1h1,a2h1, a5h1, aVh3/aVh5 as critical mediators or regulators

of angiogenesis. The characterization of the role of addi-

tional integrins in angiogenesis (e.g. a8h1, aVh8, or

a6h4), the dissection of the signaling pathways involved,

and the identification of the precise mechanism by which

pharmacological integrin antagonists inhibit angiogenesis

clearly require further experimental work.

9. Endothelial cell AVB3, a Janus integrin in

angiogenesis?

The discrepancy between the normal developmental

angiogenesis and excessive tumor angiogenesis observed

in AVB3-deficient mice on one hand and the suppression of

angiogenesis by pharmacological AVB3 antagonists seen in

wild-type mice on the other hand suggests that AVB3 may

have a dual role in regulating angiogenesis, i.e. it may

stimulate angiogenesis by promoting endothelial cell sur-

vival, as well as it may inhibit angiogenesis by inducing

endothelial cell death (Fig. 3).

A pro-angiogenic role of AVB3 is supported by a large

body of experimental evidence. Ligation of AVB3 promotes

endothelial cell proliferation by enhancing VEGF-R2 de-

pendent signaling [185], inducing sustained MAPK activity

[65], suppressing p53 activity and reducing p21WAF expres-

sion [109,186]. AVB3 ligation enhances cell survival

through the activation of NF-KB [98,99], the increase in

the Bcl-2/Bax ratio [109] and the suppression of caspase-

8 activation [119]. Furthermore, AVB3 stimulates endothe-

lial cell motility [187] and localizes MMP-2 at sites of cell

invasion [188]. VEGF induces affinity maturation of AVB3

resulting in enhanced ligand binding, cell adhesion and

migration [84]. An anti-angiogenic function of AVB3 is

supported by the observation that expression of unligated

integrin heterodimers causes apoptosis through the recruit-

ment and activation of caspase 8 [119]. This model proposes

that AVB3 ligation suppresses apoptosis by preventing

clustering and activation of caspase 8 and caspase 3 [120].

In the absence of AVB3 expression, cells would become

insensitive to integrin-mediated death [120], resulting in

enhanced survival and angiogenesis as observed in the

AVB3-deficient mice [147]. Given the strong presence of

AVB3 ligands in the tumor environment, however, it is hard

to imagine that AVB3 may actually exist without bound

ligands. Instead, it is conceivable that some ligands, in

particular soluble ones, may act as negative regulators of

angiogenesis. This hypothesis is consistent with the obser-

vation that some natural ligands of AVB3, such as throm-

bospondin-1 and -2 [189,190], the MMP-2 proteolytic

product PEX [191–193], and tumstatin, a proteolytic frag-

ment of collagen IV A3 chain, are known inhibitors of

angiogenesis [194,195].

Taken together, there is evidence to support the notion

that AVB3 may have both positive and negative functions

depending on the ligation state, the engaged substrate, the

presence of additional factors or cytokines, or the differen-

tiation state of the cell [57,170].

10. COX-2 and angiogenesis

Clinical and experimental evidence indicates that aspirin

and other cyclooxygenase inhibitors, collectively called

nonsteroidal anti-inflammatory drugs (NSAIDs), protect

against cancer [196]. Human epidemiological studies have

shown that chronic intake of NSAIDs significantly reduces

colon polyp formation and recurrence, resulting in a de-

creased risk of colon cancer development [197]. In animal

models, NSAIDs prevent tumor formation and slow tumor

progression [198]. Among the three known cyclooxygenase

(COX) isoforms (i.e. COX-1, COX-2 and COX-3), COX-2

has emerged as the one critically involved in cancer pro-

gression. COX-2 is absent from most normal tissues [196],

but its expression is strongly induced by inflammatory

cytokines [199] and by cellular transformation [200].

COX-2 overexpression is often observed in many human

cancers, including colon, breast, prostate and skin [201].

In a murine model of human familial adenomatous poly-

posis coli, genetic inactivation of COX-2 dramatically

reduced the number and size of intestinal polyps [202].

Transgenic overexpression of COX-2 in the skin and in the

breast promoted tumorigenesis and tumor progression, and

this effect was associated with enhanced angiogenesis

[203,204]. COX-2-specific inhibitors (COXIBs) suppress

tumor growth in animal models and reduce the risk of

developing polyps and colon cancer in humans [205]. In a

rat model of corneal angiogenesis, the COX-2-specific

inhibitor Celecoxib suppressed corneal blood vessel forma-

tion [206]. This effect was associated with a decrease in

prostaglandin production, an increase in apoptosis and a

decrease in proliferation of angiogenic, but not quiescent,

endothelial cells [207].

11. COX-2 activity promotes AVB3-mediated Rac

activation and angiogenesis

The mechanism by which COX-2 stimulates angiogen-

esis is a matter of intense research [208]. A first link was

established between COX-2 activity and VEGF production

and action. Disruption of the COX-2 gene in mice dramat-

ically suppressed VEGF production in fibroblasts [209] and

inhibition of COX-2 decreased VEGF production in fibro-

Fig. 3. The proposed dual role of integrin aVh3 in regulating angiogenesis. Among endothelial cell integrins, aVh3 appears to be able to both promote and

inhibit angiogenesis. To reconcile these two opposing effects, a model has been proposed in which ligated aVh3 activates survival pathways and suppresses

latent pro-apoptotic signals. In contrast, unligated aVh3 integrins, or integrins occupied by soluble ligands, actively promote apoptosis by recruiting and

activating caspase 8, and by altering the balance of anti/pro-apoptotic signals. Cell death induced by unligated integrins is referred to as integrin-mediated death

(IMD) [46,119,120]. In contrast to anoikis, IMD is initiated in attached cells.


blasts and tumor cells [210]. Furthermore, COX-2 inhib-

itors prevented VEGF-induced MAPK activation in endo-

thelial cells [211]. Our laboratory uncovered a link between

COX-2 and integrin AVB3-mediated endothelial cell mi-

gration and angiogenesis (Fig. 4). Inhibition of COX-2

activity in endothelial cells by NSAIDs and COXIBs

suppressed AVB3-dependent endothelial cell spreading

and migration in vitro and FGF-2-induced angiogenesis

in vivo [90]. Exogenous PGE2 rescued endothelial cell

spreading and migration in the presence of COX-2 inhib-

itors [90,212]. The COXIBs and NSAIDs effect was due to

the inhibition of AVB3-dependent activation Cdc42 and

Rac, two members of the Rho family of GTPases that

regulate cytoskeletal organization and cell migration. Be-

sides promoting Rac activation and cell spreading, the

COX-2 metabolite PGE2 also accelerates AVB3-mediated

endothelial cell adhesion [212]. Prostaglandins modulate

AVB3-dependent adhesion, spreading and migration

through the prostane-receptors EP2- and EP4-mediated

activation of adenylate cyclase, rise in cAMP level, acti-

vation of the cAMP-dependent protein kinase PKA and

Rac activation. These results demonstrate that inhibition of

AVB3-mediated Rac activity is an important mechanism by

which NSAIDs suppress angiogenesis and establish a novel

functional link between inflammation and angiogenesis.

The important role of Rac in angiogenesis was also

demonstrated by the observations that endothelial cell

chemotaxis induced by VEGF required Rac activation

[89,213] and that inhibition of the Rac-effector p21-acti-

vated kinase (PAK) -1, suppressed endothelial cell tube

formation in vitro and angiogenesis in the chick CAM

assay in vivo [91,214].

12. Endothelial cell cAMP and PKA regulate integrin

function and angiogenesis

The above results imply that cAMP and PKA may be

important regulators of integrin function and angiogenesis.

Indeed, there is further evidence that cAMP and PKA

positively and negatively regulate integrin function. cAMP

rise inhibited T cell adhesion on fibronectin [215] and

caused a rapid down regulation of integrin AIIbB3 function

in platelets exposed to platelet-derived growth factor [216].

On the other hand, cAMP enhanced integrin-dependent B

lymphocyte aggregation [217], increased the binding of

immature thymocytes to fibronectin [218] and promoted

endothelial cell adhesion on vitronectin [174]. In endothe-

lial cells, agents that increased cAMP level inhibited AVB3

but not A5B1-mediated focal adhesion formation and mi-

gration through the inhibition of Rho [174,219]. A rapid

increase in cAMP levels was shown to block Rho activity

through a transient activation of PKA [220]. Parathyroid

hormone-related peptide, a peptide hormone that regulates

bone metabolism and vascular tone, induced sustained

activation of PKA, and inhibition of Rac resulting in

diminished endothelial cell migration in vitro and angio-

genesis in vivo [221]. PKA was also implicated in the

suppression of endothelial cell apoptosis. Ligation of integ-

rin A5B1 was shown to suppress sustained PKA activation

and to promote survival, while A5B1 antagonists induced

caspase-8-dependent endothelial cell apoptosis [173]. Thus

cAMP and PKA appear to exert a biphasic action on

endothelial cell integrin function depending on their levels

and kinetics of activity: transient increases in cAMP levels

and PKA activity stimulate AVB3-mediated adhesion,

Fig. 4. A model for the cross talk between COX-2 and integrin aVh3. aVh3-dependent Rac activation, spreading and migration of endothelial cells is promoted

by COX-2 activity and prostaglandin (PGE2) production. PGE2 binds to E prostane receptors 2 and 4 (EPR2/4) and activates the G protein-coupled adenylate

cyclase (AC), causing a transient cAMP raise, protein kinase A (PKA) and Rac activation, and accelerated adhesion, spreading and migration [90,212].

COXIBs inhibit angiogenesis by suppressing VEGF production by tumor and stromal cells, and aVh3-dependent Rac activation and VEGF-R2 signaling in

endothelial cells resulting in decreased endothelial cell migration, reduced proliferation and increased endothelial cell death. a5h1 ligation induces a rapid

increase and a long-lasting suppression of PKA activity [173]. Suppressed PKA activity prevents caspase-8 activation and promotes endothelial cell survival.


spreading and migration, while sustained rises have the

opposite effect.

Taken together, these observations demonstrate the im-

portant role of the cAMP/PKA pathway in regulating integ-

rin function, Rac activity and caspase activation relevant to

angiogenesis. Many questions, however, remain open at this

point. For example, the exact relationship between the

kinetics of cAMP/PKA activity in response to integrin

ligation (early-transient vs. late-long lasting) or the absolute

level of activity (high–intermediate–low) and the net effects

on endothelial cell spreading, migration and angiogenesis

(stimulation vs. inhibition) require further investigations.

13. Therapeutic perspectives

While it is clear that integrins are essential mediators and

regulators of physiological and pathological angiogenesis, it

is less clear how they can be best inhibited in order to most

effectively suppress angiogenesis. Preclinical experiments

have shown that extracellular integrin antagonists inhibit

angiogenesis and progression of angiogenesis-dependent

conditions such as chronic inflammation, proliferative reti-

nopathy and tumor growth. Recent advances in the under-

standing of the structure, function and signaling of integrins,

however, suggest that there may be alternative strategies to

interfere with integrin-mediated functions in endothelial

cells.

x Alternative targets. AVB3 has been identified as the first

integrin target to inhibit tumor angiogenesis. Recent

findings suggest that additional integrins may also be

valuable targets, in particular A1B1, A2B1 and A5B1.

Combined administration of anti-A1B1 and A2B1 anti-

body reduced tumor growth and angiogenesis in mice

[183]. Similarly, the drug E-7820, a sulfonamide

derivative that suppressed A2 expression, inhibited in

vivo angiogenesis and tumor growth in mice [184].

Integrin A5B1 is up-regulated in angiogenesis and a

blocking anti-A5 mAb suppressed VEGF-induced angio-

genesis in a chick embryo and murine models. Additional

in vivo experiments are needed to confirm that A5B1

antagonists lead to consistent and significant inhibition of

angiogenesis. In a murine model of tumor metastasis, a

combined administration of ATN-161, an A5B1 antago-

nist, and 5-FU reduced liver metastases formation and

improved mice survival [176]. A recent report has

demonstrated that A9B1 is expressed on lymphatic but

not on vascular endothelial cells, raising the possibility

that A9B1 may be targeted to suppress lymphangio-

genesis in cancer [139]. Thus, these integrins should be

considered as potential alternative or complementary

targets to suppress tumor angiogenesis and lymphangio-

genesis in human cancer. Development of potent,

selective and nontoxic antagonists of these integrins,

however, is a prerequisite for entering clinical trials.

x Targeting the cytoplasmic domain. Interfering with the

integrin subunit cytoplasmic tail has been originally

shown to perturb integrin function in fibroblasts

[222,223]. We have shown that expression of an

insolated integrin B subunit cytoplasmic tail in confluent

endothelial cells through adenovirus-mediated delivery


results in endothelial cell detachment and induction of

apoptosis [224]. Infection of rat carotid arteries with the

same adenovirus resulted in endothelial cell rounding,

detachment and apoptosis, while cells infected with a

control construct retained a flattened morphology and did

not detach (G. Vassalli et al., manuscript submitted). This

approach is very effective, but so far it does not allow

selective targeting of one specific integrin because of the

intrinsic transdominant negative activity of these con-

structs. Using shorter regions of the cytoplasmic domains

may help to generate selective compounds. A peptide

carrying the very carboxyl-terminal residues (i.e. 747–

762) of the integrin B3 cytoplasmic domain was reported

to inhibit B3-mediated cell adhesion without affecting B1

function [225]. Alternatively, one may take advantage of

the fact that some cytoplasmic proteins specifically bind

to the cytoplasmic domain of one particular integrin (e.g.

B3-endonexin/B3 cytoplasmic domain) [226] to develop

antagonistic drugs specific for B3 integrin.

x Targeting signaling pathways:

Rac and COX-2 inhibitors. Rac and it downstream

target, PAK-1, have emerged as critical mediators of

angiogenesis. Inhibition of Rac or PAK-1 function in

angiogenic endothelial cells suppresses cell spreading,

migration [212,227], proliferation [93], and angiogenesis

[90]. Rac is expressed in virtually every cell and controls

essential cellular functions, such as cell polarity,

proliferation, migration, and vesicular trafficking

[228,229]. The indiscriminate pharmacological inhibition

of Rac is therefore expected to have severe toxic effects.

The observation that NSAIDs prevent Rac activation

induced by aVh3, but not by a5h1, suggests the

existence of a specific and NSAIDs-sensitive aVh3-dependent signaling pathway leading to Rac activation.

The notion of integrin-specific pathways leading to Rho-

family GTPases is consistent with the report that

overexpression of h3 integrins in CHO cells activates

Rho but not Rac, while overexpression of h1 integrin

leads to activation of Rac but not Rho [230]. The

identification of the molecules that mediate aVh3-dependent and COX-2-sensitive Rac activation may lead

the way to the design of drugs that selectively suppress

Rac activation in angiogenic endothelial cells.

The PKB/Akt pathway. The PI-3 kinase, PKB/Akt

pathway mediates cell motility and survival in response

to integrin ligation [66,231]. Tumstatin, an anti-angio-

genic proteolytic fragment of collagen IV [194], was

shown to suppress aVh3-dependent activation of the

FAK-PI-3K-PKB/Akt-mTOR pathway resulting in di-

minished endothelial cell protein synthesis and prolifer-

ation [195]. The pathways activated by these kinases may

therefore represent relevant pharmacological targets

downstream of integrins [232]. Rapamycin-mediated

inhibition of mTOR, a direct downstream target of

PKB/Akt, resulted in the suppression of tumor angio-

genesis and tumor growth in animal models [233].

Rapamycin derivatives are currently tested in clinical

trials as anti-angiogenic drugs. Direct inhibition of PKB/

Akt may be even more potent then Rapamycin in

inhibiting endothelial cell function and angiogenesis,

since PKB/Akt has many additional targets potentially

relevant to angiogenesis, such as Bad, eNOS, p21,

MDM2, FRKH and p38 stress-activated kinase

[78,234,235]. To date, however, there are no direct

pharmacological inhibitors of PKB/Akt. The recent

progresses in understanding PKB/Akt activation and the

resolution of three-dimensional structure of active PKB/

Akt will likely boost the design and development of

specific PKB/Akt inhibitors [236,237].

14. Conclusive remarks

Integrin-dependent endothelial cell adhesion, migration

and signaling events are essential for developmental and

tumor angiogenesis. Several integrin-dependent signaling

pathways contributing to angiogenesis were deciphered

and integrin antagonists with anti-angiogenic activity were

generated. While on the one side the potential efficacy of

integrin antagonists to suppress angiogenesis in human

cancer is already tested in clinical trials, we still face many

basic questions about the mechanism by which integrin

promote angiogenesis and integrins antagonists suppress it.

A better understanding of the exact role of individual

endothelial cell integrins in promoting endothelial cell mi-

gration, survival and proliferation and the dissection of the

associated signaling events will be necessary to improve the

therapeutic targeting of integrins to suppress angiogenesis.

Acknowledgements

The authors wish to thank Dr. F.J. Lejeune for continuous

support and R. Driscoll for critical reading of the manu-

script. Work in our laboratory was supported by Grants from

the Swiss National Science Foundation, the Molecular

Oncology Program of the National Center for Competence

in Research, the Swiss Cancer League/Oncosuisse, the

Gertrud-Hagmann Stiftung, fur Malignomforschung the

Leenards Foundation and the Fondation de la Banque

Cantonale Vaudoise and by a CORE award of Pharmacia

Oncology. We apologize to those colleagues whose work

could not be cited due to space limitations.

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