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Integrin-Mediated Cell-Matrix Interaction in Physiological and Pathological Blood Vessel Formation

Integrin-Mediated Cell-Matrix Interaction in Physiological and Pathological Blood Vessel Formation Hindawi Publishing Corporation Journal of Oncology Volume 2012, Article ID 125278, 25 pages doi:10.1155/2012/125278 Review Article Integrin-Mediated Cell-Matrix Interaction in Physiological and Pathological Blood Vessel Formation Stephan Niland and Johannes A. Eble Center for Molecular Medicine, Department of Vascular Matrix Biology, Excellence Cluster Cardio-Pulmonary System, J. W. Goethe University Hospital, Theodor-Stern-Kai 7, Building 9 b, 60590 Frankfurt, Germany Correspondence should be addressed to Johannes A. Eble, eble@med.uni-frankfurt.de Received 25 May 2011; Accepted 15 July 2011 Academic Editor: Debabrata Mukhopadhyay Copyright © 2012 S. Niland and J. A. Eble. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Physiological as well as pathological blood vessel formation are fundamentally dependent on cell-matrix interaction. Integrins, a family of major cell adhesion receptors, play a pivotal role in development, maintenance, and remodeling of the vasculature. Cell migration, invasion, and remodeling of the extracellular matrix (ECM) are integrin-regulated processes, and the expression of certain integrins also correlates with tumor progression. Recent advances in the understanding of how integrins are involved in the regulation of blood vessel formation and remodeling during tumor progression are highlighted. The increasing knowledge of integrin function at the molecular level, together with the growing repertoire of integrin inhibitors which allow their selective pharmacological manipulation, makes integrins suited as potential diagnostic markers and therapeutic targets. 1. Introduction growth, the tumor needs to hook up to the vascular system by forming neovessels. Invasive cancer is among the leading causes of death world- During tumor progression, an angiogenic switch is acti- wide, and rates are still increasing, due to ageing and vated causing a continuous neovessel formation emanating changes in lifestyle [1]. Cancer is a collective term for many from the normally quiescent vasculature, which sustains diseases, rather than a single disease, with the common tumor growth [6]. This process called tumor angiogenesis characteristic that tissue growth goes haywire [2]. Patients is a collective term that is generally used for all types of who have undergone cancer treatment show an increased risk tumor neovascularization. In addition to vessel co-option of developing a second tumor, mainly due to the same risk and to endothelial cell (EC) sprouting, tumor vessels can also factors that were responsible for the first tumor but also in develop by intussusceptive or glomerular angiogenesis, or, part due to the treatment of the first tumor with mutagenic in a way of vascular mimicry, even tumor cells themselves chemotherapeutics or radiation [3]. Therefore, new strate- can form vessel-like hollow structures. These types of vessel gies for cancer treatment with as little as possible adverse formation can occur in parallel, and also gradual transitions side effects are needed that effectively eradicate the primary are possible. Vessel formation by the latter types requires less tumor and also do not increase the risk of recurrence. energy than sprouting angiogenesis, is thus carried out faster, A tumor initially grows without any connection to the and usually can be observed in, for example, gliosarcoma vasculature until it reaches a critical size of about two mm multiforme, melanoma, and breast and colon cancer [7]. in diameter. Then it remains in a dormant state, in which For neovessel formation, ECs need to migrate into a pre- proliferation and apoptosis due to lack of oxygen, are in a viously avascular region and to extensively remodel the dynamic equilibrium unless it develops in a well-vascularized extracellular matrix (ECM). In this process, integrins, which region or is able to recruit its own vasculature. Hanahan are cell adhesion receptors for various ECM proteins and Weinberg have proposed six hallmarks of cancer, one of and immunoglobulin superfamily molecules, are the most them being the induction of angiogenesis [4, 5]. For further important matrix receptors [8, 9]. Therefore, integrins are 2 Journal of Oncology appealing targets for cancer therapy using a variety of inte- In different vessel types, that is, arteries, arterioles, cap- grin-specific antagonists, ranging from endogenous antago- illaries, venules, and veins, this general blueprint is mod- nists over humanized or chimeric antibodies to peptides and ified corresponding to the respective functional require- small nonpeptidic compounds [10–12]. ments. For example, endothelia, which are continuous in most instances, can become fenestrated, as in exocrine or In this paper, based on the general assembly of blood endocrine gland tissues, or even discontinuous, as in liver, vessels, the specific organization of tumor vasculature will be spleen, or bone marrow, in order to facilitate the exchange described, as well as the dynamic sequence of events by which of hormones or metabolites. Elastic and muscular arteries a tumor gains access to the body’s vasculature. In this context, illustrate other examples for a modification of this general the role of integrins and possibilities of their pharmacological blueprint. In order to even the pulsatile blood flow coming manipulation are explored. from the heart, the proteins elastin and fibrillin are abundant in the tunica media ECM of elastic arteries, which is the 2. The Static Picture: The Extracellular Matrix direct cause for the vessel wall’s elastic properties. Muscular of Blood Vessels arteries possess numerous concentric sheaths of smooth muscle cells. By means of vasoconstriction and vasodilation, The tissue’s ECM is a structure-shaping molecular scaffold they can distribute and direct the blood to different organs. and also a repository for cytokines and other growth factors [13]. Cells embedded in this matrix need to be supplied 2.2. Extracellular Matrix in the Vessel Wall. The ECM of with oxygen and nutrients, signaling molecules need to be blood vessels together with their resident cells contributes to received and emitted, and metabolic waste products need essentially all physiological functions of blood vessels and has to be disposed of. These tasks are optimally fulfilled by the been reviewed recently [15]. cardiovascular system with its intricate and dynamic network The subendothelial basement membrane (BM) com- of blood vessels. Depending on their functions, different partmentalizes the vessel’s single-layered endothelium from types of blood vessels show special histological and molecular the vascular connective tissue. The molecular architecture adaptations. The heart, as a double-acting pump, drives of BMs has recently been reviewed [16–18]. Fibronectin, the blood circulation within the vasculature via the aorta incorporated between endothelial and perivascular cells, is through arteries and arterioles into capillaries, from where essential for blood vessel morphogenesis [19]. The presence the blood flows back through venules and veins. Due to of von Willebrand factor (vWF) is characteristic for the the prevailing pressure conditions, the body fluid is forced subendothelial BM, where also other BM proteins, such as through the vessel wall to form the lymph, which then is the network-forming collagens IV and XVIII can be found, drained by lymph vessels back to the blood circulation. together with laminins, nidogens, and perlecan. Thirteen Additionally, the vasculature serves as “highway” system different collagens are present in the vascular wall [20, 21]. for leukocytes to patrol the body during immunological The network-forming collagen IV [22] plays a key role for surveillance and to quickly reach sites of inflammation. The the mechanical stability of the BM [23], which, especially in vascular wall is capable of self-sealing upon smaller injuries, arterial regions of the circulatory system, has to withstand a and leukocytes are able to penetrate the blood vessel wall considerable blood pressure. in a complex interplay without any obvious vessel leakage. In the tunica media of elastic and muscular arteries, Pathologically, tumor cells capitalize the blood vessel system covalently crosslinked supramolecular aggregates of elastin to disseminate from a primary tumor and to colonize distant form concentric lamellae and fibers in a proportion of up organs where they develop metastases. to 50% of the vessel’s dry weight and confer resilience to pulsatile blood flow [24–26]. Regions of the ECM that 2.1. General Organization of the Vessel Wall. Histologically, consist mostly of elastin are confined by EMILINs, that is, the walls of blood vessels comprise three concentric layers, homotrimeric elastin microfibril interphase-located proteins that is, tunica intima, tunica media, and tunica adventitia [27]. Anchored to microfibrillar bridges of fibrillin-1 and [14], which are separated by two sheet-like structures of fibulin-5 between these concentric elastin lamellae, vascular ECM proteins. The membrana limitans interna and externa smooth muscle cells (VSMCs) are sandwiched in a fishbone- establish a border between tunica media and tunica interna like pattern and thus can effectively regulate the vessel’s and adventitia, respectively. These ECM sheaths tightly caliber [25, 28–31]. Dependent on the vessel type, distinct connect the cell layers of the vessel wall to form a functional fibulins are involved in the assembly of the ECM. While unit, which becomes evident when too weak cell-matrix fibulin-1 is widespread and occurs in the BMs of all blood interactions lead to life-threatening aneurysms. vessels, heart valves and septa, fibulin-3, and fibulin-4 occur The tunica intima comprises a single layer of squamous in the walls of capillaries and larger blood vessels [32]. The ECs and lines the inner surface of all blood vessels. The innermost and outermost elastic lamellae are referred to as tunica media, which is usually the thickest layer in arteries, membrana limitans interna and membrana limitans externa, is composed of mural cells, which are smooth muscle cells respectively. Between the elastic lamellae, type I and III in larger blood vessels and pericytes in capillaries. The collagens are deposited that bear tensile forces exerted on tunica adventitia finally interconnects the blood vessel with the vessels and limit their elastic dilatability. In contrast, in the surrounding connective tissue, and it is usually most the interstitial connective tissue between the subendothelial prominent in veins. membrane and the membrana limitans interna, type VI and Journal of Oncology 3 type VIII collagens are found [21, 33]. The connection of the osteopontin receptor α8β1, and integrin αvβ3 is also the membrana limitans interna to the subendothelial BM by expressed on glial cells [49]. type XVIII collagen is assumed [34]. Also type XVI collagen, As EC-derived tumors, angiosarcomas express the inte- which is produced by VSMCs and found close to both grins α1β1, α2β1, α3β1, α5β1, and α6β1, and in benign and elastic microfibrils and fibrillar type I and type III collagens, malignant mesenchymal tumors as well as in the desmoplas- may contribute to the connection between the elastic and tic stroma of carcinomas, integrins α1β1and α5β1are widely collagenous phases of the ECM [35, 36], especially, as type distributed [50]. Integrins α1β1and α2β1 bind to the same XVI collagen contains a binding site for the major collagen ligand in the ECM and are VEGF-dependently upregulated receptor on VSMCs, integrin α1β1[37, 38]. on migrating ECs, and antagonists against both integrins The ECM of the tunica media is synthesized by VSMCs, inhibit VEGF-mediated angiogenesis without affecting the which are all encapsulated by an (incomplete) BM containing existing vasculature [51, 52]. Therefore, and against the the usual BM proteins, type IV collagen and laminins background of gene ablation studies, they are believed to dif- [33, 39]. Depending on microenvironmental cues, VSMCs ferentially regulate angiogenesis [49]. Important coreceptors can reversibly acquire distinct phenotypes, which can be for integrin α2β1 are the syndecans-1 and -4, which weaken characterized as either (i) contractile and differentiated or the invasiveness of tumor cells into a collagenous matrix [53]. (ii) secretory, migratory, and less differentiated [37, 39]. Cells bind to fibronectin and vitronectin preferentially Under physiological conditions, the contractile phenotype via the RGD-dependent integrins αvβ3and α5β1[54]. prevails, at which the VSMCs transduce forces on the Fibronectin can also be bound by the leukocyte-specific pericellular matrix especially by the collagen-binding inte- integrins α4β1and α4β7[55]. Cell-fibronectin interactions grin α1β1, by the laminin-binding integrin α7β1and by are modulated by proteoglycans, glycoproteins of the ECM, dystroglycan [37]. In contrast, in the secretory, proliferatory, and the coreceptors syndecans [56]. and migratory phenotype, the integrin equipment of the Integrin αvβ3 was identified as a marker for angiogenic VSMCs predominantly consists of the fibronectin receptor, vascular tissue [57]. In contrast to quiescent ECs, integrin α5β1, and the integrins α4β1and α9β1. Consistently, in the αvβ3 is highly expressed on activated ECs during tumor proximity of secretory VSMCs, the fibronectin splice variants angiogenesis,aswellasonsometumor cells[58, 59]. In the V (IIICS) and EIIIA with binding sites for the integrins α4β1, tumor microenvironment, angiogenic ECs can interact due α5β1, and α9β1 are abundant [39]. In capillaries, scattered to their increased levels of the integrins αvβ3and αvβ5 pericytes, each encapsulated by an own BM, stabilize the with provisional matrix proteins, such as vitronectin, fib- endothelium and its subendothelial BM [40–42]. rinogen, vWF, osteopontin, and fibronectin. Also, partially The fibroelastic connective tissue of the tunica adventitia proteolyzed collagen in the tumor exposes RGD sites and is connects the blood vessel with the perivascular connective a further ligand for integrin αvβ3[60]. Thus, the ECM of tissue. It is rich in versican, a glycoprotein, which can interact the tumor microenvironment both provides survival signals with fibrillin-1 [43], fibulin-1 [44], and fibulin-2 [45], as well and facilitates invasion. Integrin-αvβ3-mediated adhesion as with other ECM molecules. to platelets protects malignant cells from clearance through the immune system, and moreover, αvβ3 integrin also helps tumor cells to adhere to the vessel endothelium and to spread 2.3. Receptors for ECM Molecules. To interact with their microenvironment and to spatiotemporally regulate their into adjacent tissues [61]. differentiation state, morphology, metabolism, and survival, The pharmacological inhibition of integrin-αvβ3- mediated cell-matrix interaction impedes tumor angiog- cells are equipped with a variety of receptors for all the ECM molecules [13]. Integrins are the largest family of these enesis and growth [62], as does a replacement of the β3 receptors, and they mediate adhesion to collagens, laminins, subunit with a mutated nonphosphorylatable subunit in a and fibronectin. In addition, there are other receptors and murine model [63], which provides evidence for a proan- coreceptors, such as the syndecans [46]. giogenic role of integrin αvβ3, in contrast to integrin αvβ5, Binding to a wide variety of different ECM molecules which does not seem to play an essential role in angiogenesis and transmitting signals bi-directionally in an outside-in [64]. Interestingly, the analysis of αv-knock-out mice and inside-out manner, integrins constitute functional hubs, revealed that, despite being embryonic or perinatally lethal, the vascular endothelium was not impaired in the absence which, according to an interesting concept in network theory and systems biology, integrate networks of angiogenic sig- of the αv subunit, whereas the primary cause of death was naling cues that orchestrate the behavior of ECs and VSMCs brain hemorrhage [65–67]. Also endothelial Tie-2-specific knockout of the αv subunit did not result in any vascular during angiogenesis [47, 48]. Thus, therapeutically targeting integrins as the operationally important circuit-integrating or angiogenesis defect [67]. Moreover, in an integrin hubs rather than single pathways of the complex system may subunit β3- and also β5-deficient mouse model, pathologic result in a more pronounced inhibition of angiogenesis [47]. angiogenesis and tumor growth are increased [68]. A ECs express the vitronectin receptors αvβ3and αvβ5; possible cause for these seemingly contradictory phenomena could be a relief of a transdominant inhibition by αvβ3on moreover, on ECs and pericytes the following integrins are expressed: the collagen receptors α1β1and α2β1, the laminin other integrins or other molecules, which would enhance receptors α3β1, α3β6, and α6β4, the osteopontin receptor their proangiogenic function [69, 70]. Likewise, there could be a compensatory role of other integrins with overlapping α9β1, and the fibronectin receptors α4β1and α5β1[49]. Pericytes additionally express the laminin receptor α7β1, and function [49]. Moreover, inhibition could also stabilize the 4 Journal of Oncology integrin αvβ3 in its unligated conformation and thus induce interaction with a multitude of proteins, such as MMPs, apoptosis by triggering an integrin-mediated death program uPA/uPAR, tissue inhibitor of matrixmetalloproteinase-2 [71]. (TIMP-2), vWF, TSP-1, osteopontin, syndecan-1, insulin- Integrin αvβ8isimportant forvasculardevelopment in receptor substrate-1 (IRS-1), cytohesin-1, integrin cytoplas- the embryonic brain and in the yolk sac [72]. It is expressed mic domain-associated protein-1 (ICAP-1), integrin-linked on astrocytes but not on ECs or pericytes, nevertheless plays kinase (ILK), calcium- and integrin-binding protein (CIB), an important role in angiogenesis, as it binds in addition to β3-endotoxin, talin, actinin, tensin, nischarin, and the Ras- several ECM proteins also to the latency-associated peptide related protein Rab 25 [9]. (LAP) of TGFβ1, which in cooperation with the membrane- The subendothelial BM of the tunica intima serves as type metalloproteinase MT1-MMP/MMP14 results in acti- a mechanical support to which ECs are anchored by vari- vation of TGFβ and triggering of its downstream signal ous adhesion molecules, especially integrins [46, 105–108]. cascades [73–75]. Additionally, the subendothelial BM provides microenviron- Collagen IV, an essential component of BMs, is bound mental information that regulate the metabolic activity of by integrin α1β1, whichisexpressedonmesenchymal cells attached ECs, such as their production of leukocyte adhesion and can also bind to other collagens [76, 77]. Further molecules [107] or antithrombotic prostacyclins [109], as collagen-binding integrins are α2β1, the main receptor for well as other properties, for example, the tightness of inter- fibrillar collagens, which is expressed on epithelial and some cellular contacts [108]. Therefore, angiogenesis is regulated mesenchymal cells as well as on thrombocytes [78], α10β1in not least by integrins which are adhesion receptors for cartilage [79], and α11β1, a key receptor for fibrillar collagen matricellular proteins, ECM proteins, and immunoglobulin on fibroblasts [80]. The integrins α1β1and α2β1are involved superfamily molecules, on nearly all cells including ECs in the regulation of collagen and MMP synthesis and thus [8, 58]. of special importance for ECM turnover [81–83]. Discoidin In addition to their mechanical function [110], integrins domain receptors DDR1 on epithelial cells and DDR2 on also assist growth factor receptors and play important roles mesenchymal cells are further collagen receptors with tyro- in signaling processes, in particular as soluble growth factors, sine kinase function and are relevant for cancer [84]. and other signaling molecules are bound by integrins as well Other collagen receptors are glycoprotein GPIV on platelets [111]. For example, the proangiogenic VEGF-A is bound [85], the leukocyte-associated immunoglobulin-like receptor by integrins αvβ3and a3β1[112] and also by the tenascin- LAIR-1/CD305 [86], and the urokinase-type plasmino- C- and osteopontin-receptor integrin α9β1[113]. The latter gen activator receptor-associated protein uPARAP/Endo180, integrin, furthermore, binds the lymphangiogenic growth which is involved in matrix turnover during malignancy [87]. factors VEGF-C and VEGF-D [114]. Angiopoietins-1 and Laminin, as a further integral component of BMs, is -2 are bound by integrin α5β1[115]. Integrin α6β1isa bound by the integrins α3β1, α6β1, α6β4, and α7β1[88– receptor for the proangiogenic CCN-family member CYR61, 91] and also by α-dystroglycan [92, 93] and by the 67 kDa and is involved in in vivo in tube formation [116, 117]. The laminin receptor 67LR [94]. 67LR is increased in various fibronectin receptor integrin αvβ3, which is the best-studied tumors and correlates with their metastatic potential [95, integrin in relation to angiogenesis and is upregulated during 96]. The different laminin receptors may also act coopera- wound healing and retinal vascularization and especially on tively in laminin binding, for example, laminin-binding β1 tumor blood vessels, also binds to fibroblast growth factor integrins and 67LR [97] or integrin α6β4 and syndecan 1 FGF-1 [118]. Semaphorin 7A binding is also reported for the [98]. collagen receptor integrin α1β1[119]. Integrin α3β1, which in the vascular wall binds to lam- Stimulated by PDGF, vascular smooth muscle cells inins-411 (laminin 8) and-511 (laminin 10), thrombospon- express the laminin receptor integrin α7β1, which plays an din (TSP), TIMP2, tetraspanin CD151, and to the C-termi- important role in recruitment and differentiation of VSMCs nal domain of the collagen IV α3 chain, is controversially [120, 121]. ascribed either a positive or a negative role in angiogenesis Integrin α9β1 is not only involved in lymphangiogenesis (cf. [99]). [114] but also plays a role in EC adhesion [122]. While There is controversy whether the hemidesmosomal inte- binding of TSP-1 to integrin α9β1 promotes angiogenesis grin α6β4, whichisexpressedonasubset of ECs[100]and on [123], VEGF-A is another ligand of integrin α9β1[113]. tumor ECs [101], aggravates pathological angiogenesis [101] or whether it is a negative regulator of angiogenesis that is downregulated at its onset [102]. 2.4. Vascular-Relevant Integrin-Deficient Mouse Models. The Thus, many molecules of the ECM scaffold, for example, crucial involvement of integrins in EC biology has been laminins, collagens, fibronectin, and vitronectin, are ligands elucidated substantially by the examination of genetic knock- for integrins that link the cell’s cytoskeleton to the ECM. Loss out studies [124]. By ablation of the respective genes, the of this matrix-integrin contact triggers apoptotic cell death EC integrins α1β1, α2β1, α4β1, α5β1, α6β1, α6β4, α9β1, [103]. Picking up signals from the cell’s microenvironment, αvβ3, and αvβ5 and also the VSMC integrin α7β1 and the integrins functionally sense, interpret, and distribute infor- glial cell integrin αvβ8havebeenimplicatedinregulation mation, which allows the cell to modulate its proliferation, of cell growth, survival, and migration during angiogenesis differentiation, migration, and shape [104]. The modulatory (for recent reviews of the findings from knock-out mice cf. and regulating function of integrins is emphasized by direct [8, 10]). However, due to redundancy and compensatory Journal of Oncology 5 mechanisms, the interpretation of knock-out results is often angiogenesis [101], but die of severe skin defects [100]. In difficult. neovascularization, the endothelial expression of integrin Itgb1−/− mice die at E5.5 before they start to develop α6β1 is downregulated [102]. While it is not required for EC their vasculature [125, 126]. Mice with a conditional knock- proliferation and survival, it promotes tumor angiogenesis out in Tie-2-positive ECs survive until E9.5–E10.5, and [101]. In contrast, genetic ablation of α7β1, which is they are capable of vasculogenesis, but their angiogenesis is expressed on VSMCs but not on ECs, leads to incomplete disturbed showing defects in sprouting and branching [127– cerebral vascularization and hemorrhage and also to pla- 129]. Another endothelial-specific knockout of the integrin cental vascular defects, which results in partial embryonic β1 subunit is mediated via VE-cadherin-Cre recombinase lethality and demonstrates that integrin α7β1isimportant and becomes manifest later in embryogenesis resulting in for recruitment and survival of VSMCs [121, 137]. lethality between E13.5 and E17.5 [130]. In this mouse Deletion of Itga8 resulting in lack of integrin α8β1, model, loss of β1 integrin leads to a decreased expression a receptor for fibronectin and tenascin, results in partial of the cell polarity gene PAR3 and thus to disruption of EC embryonic lethality, but no defects in vascular development polarity and lumen formation [130]. (Mul ¨ ler and Reichardt, cited in [138]). Itga1−/− mice, deficient for the collagen-binding inte- Itga9−/− mice lacking integrin α9β1, which is the grin α1β1, show a normal vascular development and a receptor for tenascin-C, osteopontin, VCAM-1, and also for reduced tumor angiogenesis in adulthood, which has been VEGF-A, -C, and -D [113, 114], have defects in large lym- attributed to increased MMP activity [131], while α2β1- phatic vessels and die postnatally at P8-12 from a bilateral deficient Itga2−/− mice show an enhanced tumor angio- chylothorax [139]. genesis in adulthood, but an otherwise normal vascular Ablation of Itgav, resulting in simultaneous loss of the development [131, 132], and integrin α2β1isinvolvedin two integrins αvβ5, a receptor for vitronectin, osteopontin, the PlGF-dependent regulation of VEGFR-1 [132]. Although and Del-1 (developmental locus 1), and αvβ3, a recep- integrin α1β1and α2β1 bind to the same ligand in the tor for a variety of ECM proteins, such as fibronectin, ECM, their differential knockout results in opposing effects vitronectin, laminins, fibrinogen, fibrin, TSP, tenascin-C, on angiogenesis, suggesting a regulatory role for this pair of vWF, denatured collagen, osteopontin, MMP-2, Del-1, bone integrins. sialoprotein, FGF-2, thrombin, and CCN1 (cystein-rich Da Silva and coworkers generated EC-specific condi- protein 61), leads to 80% embryonic lethality at E9.5, and tional α3 integrin knock-out mice and showed that these the other 20% die at P0 with brain hemorrhage [65]. On mice, in contrast to a global ablation, are viable and fertile the other hand, Itgb3−/− mice, which are just integrin-αvβ3 but display enhanced tumor growth, elevated hypoxia- deficient, show 50% embryonic and early postnatal lethality induced retinal angiogenesis and tumor angiogenesis, and and an enhanced angiogenesis in surviving adult animals, increased VEGF-mediated neovascularization [99]. The indicating that this integrin is not strictly required for vascu- authors also could show that α3β1 is a positive regulator of lar development [140]. Surprisingly, animals with an intact EC-derived VEGF, which again represses VEGFR2 expres- but nonfunctional β3 integrin subunit develop normally but sion. Their data demonstrated that endothelial α3β1nega- show defects in angiogenesis in adulthood [63]. In contrast, tively regulates pathological angiogenesis and implicated an Itgb5−/− animals lacking integrin αvβ5develop normally unexpected role for low levels of EC-derived VEGF as an and angiogenesis is not significantly affected, indicating that activator of neovascularization. this integrin is not mandatory for vascular development [64]. Itga4−/− mice, deficient for fibronectin- and VCAM1- Integrins β3and β5 doubly deficient mice show enhanced binding integrin α4β1, are embryonic lethal with 50% dying tumor growth and angiogenesis. This strongly suggests that at E9.5–10.5 due to failure of chorion-allantois fusion and these integrins are not required for vascular development or 50% dying at E11.5 due to cardiovascular defects [55]. for pathological angiogenesis, pointing out that the mode of Mice, which by ablation of Itga5 are deficient for the action of αvβ3 antagonists and antiangiogenic therapeutics is fibronectin receptor integrin α5β1, show normal vasculo- still insufficiently understood [68]. Ablation of Itgb8 leads to genesis but no angiogenesis, which results in embryonic the loss of integrin αvβ8 on glial cells and thus to disrupted lethality at E10-11 due to defects in posterior somites, blood vessel formation in the brain, thereby demonstrating yolk sac, and embryonic vessels [133, 134]. This demon- that this integrin is mandatory for brain’s blood vessel strates the requirement of the integrin α5 subunit during development [72]. Moreover, the phenotype of β8-deficient embryonic development of early blood vessels and other mice resembles that of αv-deficient mice, which provides tissues. Accordingly, integrin α5β1, which is poorly expressed evidence that most defects in αv-deficient mice are due to on normal quiescent ECs, is markedly upregulated during the loss of integrin αvβ8[72]. tumor angiogenesis [135]. Among the laminin-binding integrins, integrin α6isnot 2.5. Integrin Structure. The family of integrins contains 24 essentially required for vascular development, although α6- structurally related N-glycosylated heterodimeric proteins deficiency is lethal with skin blistering defects resembling assembled noncovalently from 18 α-subunits and eight epidermolysis bullosa [136]. In line with the α6 knock- β-subunits. Each subunit comprises a large extracellular out mice, Itgb4−/− mice, lacking a functional laminin- domain, a single transmembrane domain, and with the binding integrin α6β4 by deletion of its signaling domain, exception of the β4 integrin subunit, a short noncatalytic show normal vascular development, although with reduced cytoplasmic tail [141]. Integrins are of special importance as α β 6 Journal of Oncology Collagen α β α β α ββ α α β 13 2 Integrin signaling Figure 1: Integrin activation. Integrins are a family of heterodimeric transmembrane adhesion receptors that bidirectionally relay signals with the extracellular matrix (ECM) and also with other cells. When activated, a conformational change increases the affinity, and clustering increases the avidity towards the ligand. (1) By inside-out signaling, integrins can reversibly undergo a conformational change from a bent inactive to an upright activated conformation with intermediate ligand affinity, at which the cytoplasmic domains are still close together. (2) Upon ligand binding, the integrin adopts a high-affinity conformation with a concomitant parting of the legs and a separation of the cytosolic α-and β-tails that unlocks docking sites for cytosolic molecules. (3) Clustering of ligand-occupied and activated integrins establishes a mechanical link between ECM and cytoskeleton and leads to the recruitment of scaffolding molecules and kinases. (4) The assembly of focal adhesions triggers intracellular signaling cascades. Details can be found in the text. they mediate cell matrix crosstalk via both outside-in and adopts an activated upright conformation [106, 151]. This inside-out signaling [54, 142]. Moreover, the 24 different conformational change is conveyed through the transmem- integrins possess promiscuous and redundant ligand speci- brane domains towards the cytoplasmic tails [54, 105, 152], ficities, which is of importance when distinct signals are to be where cytoskeletal proteins and signaling molecules relay the transduced or when in a particular context a defined cellular incoming signal intracellularly [153]. In inside-out signaling, response is elicited, as is discussed by Ruegg ¨ and Alghisi [11]. the binding of intracellular molecules, such as talin or Integrin structure and function have been studied in kindlins [154, 155], to the cytoplasmic integrin tails leads via detail at the molecular level [143, 144]. The extracellular a separation of the transmembrane domains [156]toaswitch headpiece is formed by a disk-like propeller domain of the blade-like erection of the extracellular domains [147, 157, α subunit and globular domains of the β subunit [145, 146]. 158]. Likewise, in outside-in signaling, ECM ligand binding The joint globular head harbors the ligand-binding site [146, to the integrin headpiece also induces a conformational 147]. The crystal structure of the integrin-αvβ3-binding site change in the hybrid domain and thereby a separation of with an inserted RGD ligand [148] helped to map functional the integrin subunits’ legs [144]. This parting of the legs amino acid residues on other integrins [149]. Recently, separates the cytosolic tails and allows binding of cytosolic the binding pocket of integrin α5β1has been mapped by proteins and thus clustering of integrins and formation of swapping regions of zebrafish and human α5 subunit in a focal adhesion sites (Figure 1). gain-of-function approach [150]. By clustering into focal adhesions, integrins recruit talin, paxillin, α-actinin, tensin, and vinculin and thereby mechan- 2.6. Integrin Signaling. Depending on their activity, integrins ically couple the ECM scaffold to the actin cytoskeleton. adopt distinct conformations (Figure 1). In the inactive rest- Additionally, integrins bind scaffolding molecules, such as ing conformation, the headpiece of the heterodimer bends p130 CRK/BCAR1, and recruit and activate kinases, such as towards the plasma membrane, and the transmembrane focal adhesion kinases (FAKs), Src family kinases (SFKs), and domains of the α and β subunits are associated [146]. Upon integrin-linked kinase (ILK), the latter forming a complex ligand binding, the previously bent integrin ectodomain with the adapter molecules parvin and PINCH/LIMS1 [159]. Inactive integrin Activatedintegrin Ligand-occupied integrin Clustered integrin Journal of Oncology 7 In addition, tetraspanins can recruit integrins to mem- the activation of PI3K by Ras is important for lymphangio- brane microdomains, thus regulating integrin function genesis [190]. [160]. Thereby, the rather unstable nascent adhesions are In addition to a direct activation of ERK, integrins can transformed into focal complexes, focal adhesions, fibrillar also activate a Raf/MEK/ERK signaling cascade in ECs [189, adhesions, or podosomes. This clustering of integrins leads 191, 192]. Raf-deficient and MEK-deficient mice have severe to a reorganization of the plasma membrane around the focal vascular defects [193, 194]. Growth-factor-mediated ERK adhesion into caveolin-containing lipid rafts, to which also signaling is linked with integrin-mediated signaling via FAK growth factor receptors often localize, and to the assembly [195]. Integrin-mediated ERK signaling is important for cell of adhesion signaling complexes [161–163]. This allows a proliferation and migration of ECs [191, 196]. Integrin α1β1 regulation of growth factor signals by integrin-mediated is unique among the collagen-binding integrins because it caveolae trafficking [164, 165]. In the assembly of such promotes cell proliferation by activating the Ras-Shc-MAPK integrin adhesions, up to 156 distinct molecules, amongst pathway, and cell cycle progression is regulated via FAK, other adaptor proteins, kinases, and phosphatases, are Rac, and cyclin D by integrin-mediated adhesion and matrix involved [48, 163]. Membrane lipid-protein interactions that stiffness [197–199]. modulate the homo- or heterotypic association of receptor Integrins can also activate the NF-κB pathway in ECs and molecules in the cell surface, or between adjacent cells, protect them from apoptosis [200–202]. Additionally, NF-κB have been reviewed recently [166]. From the focal adhesion signaling regulates the expression of cyclooxgenase-2 (COX- sites signal pathways diverge that regulate diverse cellular 2), which again is involved in EC spreading and migration programs, such as adhesion, migration, proliferation, and and in the induction of VEGF and FGF-2 [177, 203, survival. To provide an overview, integrins generally relay 204]. However, inhibition of the NF-κB pathway increases their signals via the FAK, ERK, and NF-κBpathways[153]. angiogenesis pathologically [205]. In most cases, in mechanosensory signaling FAK, Src, Integrins alone are not oncogenic, but some oncogenes and SH2, domains containing protein tyrosine phosphatase may depend on integrin signaling for tumor growth and 2(SHP2)are involved [167]. Upon integrin binding, FAK invasion. For example, integrin-triggered FAK signaling is autophosphorylates and binds to Src, which further phos- essential for Ras- and PI3K-mediated oncogenesis [206, phorylates FAK and several downstream binding partners, 207]. Also the expression of the cancer stem cell marker amongst others, JNK and Rho [168–170]. CD44 is integrin-regulated, and it can be speculated that Activated FAK also recruits PI3K, which mediates the integrin-relayed signals are needed to maintain a cancer activation of AKT and procures integrin-mediated cell sur- stem cell population [12, 208]. On the other hand, there vival, and likewise the antiapoptotic AKT can be activated via is evidence that the collagen receptor integrin α2β1has a Ang-1 [171]. Moreover, signals relayed via integrins and Src tumor-suppressing function [209, 210]. can be integrated by FAK with growth factor receptor-relayed Ligated integrins promote survival, whereas unligated signals via Ras, MEK, and MAPK [172]. Growth factors integrins recruit caspase-8 to the plasma membrane and can activate Ras signaling independently from integrin- promote apoptosis in a process termed integrin-mediated relayed adhesion signals. Nevertheless, MEK1 and Raf1 are death [71, 211], which differs from anoikis induced by loss important interfaces between integrin-relayed and growth- of cell adhesion to the ECM [103, 212]. Loss of caspase-8 factor-relayed signaling, because both MEK1 and Raf1 need confers resistance to integrin-mediated death of tumor cells, to be activated via adhesion-mediated activation of Src and and unligated integrin αvβ3 promotes the malignancy of FAK in order to activate MAPK [173, 174]. such tumors [213, 214]. Cell survival is promoted by integrin An endothelial-specific ablation of FAK results in im- ligation-dependent upregulation of BCL2 and FLIP/CFLAR, paired blood vessel development and embryonic lethality activation of the PI3K-AKT pathway, NF-κB signaling, [175] Downstream of FAK, Src couples integrin-mediated and p53 inactivation [176, 202, 215–217]. Survival is also and VEGF-receptor-mediated proangiogenic signaling in promoted by crosstalk between integrins and growth factor ECs [176–178]. However, endostatin can also activate Src receptors, for example, αvβ3 and FGFR or αvβ5 and VEGFR2 via integrin α5β1 and thereby disassemble actin stress fibers [195, 218]. and focal adhesions and thus inhibit cell migration, which is In various steps of angiogenesis and tumor progression, regulated by integrins via the Ras/ERK pathway [179–181]. crosstalk between integrins and growth factor receptors Important for adhesion and migration of endothelial and on tumor cells and also on host cells is important. This VSMCs are also p130Cas and PLC-γ, which can interact with crosstalk can consist in either an activation of a latent growth FAK [182–185]. factor, a regulation of common pathways for signaling or PI3K is of pivotal importance for angiogenesis, because internalization and recycling, a collaborative or a direct its deletion results in embryonic lethality E9.5 to E10.5, activation, or also a negative regulation [111]. The outcome when angiogenesis is important for vascular development. of a growth factor signal in a particular context is often PI3K deletion also causes decreased Tie-2 expression and determined by a synergistic and reciprocal interaction of thus creates a phenotype resembling Tie-2 deficiency [186, integrins with growth factor receptors, such as tyrosine 187]. Moreover, EC-specific deletion of the PI3K isoform kinase receptors like VEGFRs and Tie-2, Met, and FGFR, and p110α impairs angiogenesis [188]. In ECs, adhesion via semaphorins regulate integrin function as well [111, 219– integrins elicits a survival signal via FAK/PI3K/mTOR/4E- 221]. A complex of VEGF with the fibronectin heparin II BP1 and Cap-dependent translation [189]. Furthermore, domain increases, upon cell binding via integrin α5β1and 8 Journal of Oncology the signaling via VEGFR2 synergistically [222]. Expression of intensively studied one [237]. Mediated by HIF-1, VEGF-A integrin α11β1 on tumor-associated fibroblasts has a tumor- synergizes with FGF-2. VEGF is upregulated under hypoxic promoting effect, because it upregulates the expression and hypoglycemic conditions prevailing within tumor tissue of insulin-like growth factor 2 (IGF2), which is another [230]. example of integrin-regulated growth factor signaling [223]. The role of chemokines in tumor angiogenesis and neo- Beside binding ECM proteins and thus regulating adhe- vascularization has been reviewed recently [238]. Tumor sion and migration, integrins can also directly interact with cells express CCL2/MCP-1 (C-C-motif ligand 2/mono- pro- and antiangiogenic factors [221]. Integrin α5β1can cyte chemotactic protein-1), and thus, tumor-associated bind to matrix-bound VEGFR-1 [224]. In addition, integrin macrophages (TAMs) are recruited, resulting in an inflam- α9β1 can directly interact with VEGF-A, -C, and -D and matory response. These TAMs are again a source for also with hepatocyte growth factor (HGF) [113, 114, 225]. angiogenic growth factors, such as, VEGF and FGF-2 [239, Moreover, integrin α3β1and αvβ3 bind VEGF-A and 240]. MCP-1 also mediates the recruitment of mural cells in VEGF-A [112]. FGF is directly bound by integrin αvβ3 an Ang-1-dependent manner in an ex vivo model [241]. [226]. Angiopoietins also can directly interact with many Multiple sequential steps are required for angiogenesis integrins [115, 221, 227, 228]. to be successful and in all steps of this angiogenic cascade In the context of a hypoxic tumor microenvironment, it integrins, which mediate interactions of cells with surround- is especially interesting that the expression of integrins α1β1 ing insoluble ECM proteins, in addition to soluble growth and α2β1 is upregulated by VEGF [51]. factors, play an important role [15]. In a first step, the BM of an existing vessel is degraded by MMPs that are expressed by ECs, such as MMP-1, MMP-2, MMP-9, and 3. The Dynamic Process: Connection of MT1-MMP/MMP14 [242–244], at which MMP-9 is required a Tumor to the Host Vasculature for tumor vasculogenesis rather than angiogenesis [245]. Angiogenesis is an important step in the metastatic cascade, Subsequently, cell-matrix contact influences the outgrowth of tip cells and the proliferation of stalk cells that thereupon which not only provides the tumor with nutrients but also form endothelial tubes [246]. A new BM is assembled by is a route for dissemination. An important trigger for this is hypoxia [229]. newly synthesized BM proteins. Finally, the newly generated capillaries undergo maturation, pruning, and expansion. 3.1. An Angiogenic Switch Triggers the Angiogenic Cascade. In avascular tissue regions, an oxygen diffusion limit of about 3.2. Tumor Vessels Can Arise by Different Types of Vessel 150 μm restricts tumor growth to just a few millimeters in Formation. During embryonic morphogenesis, endothelial diameter. Thus, in this prevascular phase of tumor dor- precursor cells called angioblasts initiate the body’s vascu- mancy, there is a dynamic equilibrium between proliferation lature by forming tubes in a process called vasculogenesis. and hypoxia-induced apoptosis [230]. The dormant phase This is subsequently accompanied by sprouting (angio- ceases when a tumor recruits its own vasculature by the genesis) of new vessels from already existing ones. Once secretion of angiogenic factors into its environment [231], morphogenesis is completed, the adult vasculature is largely a process denoted as angiogenic switch [2, 6]. After this quiescent, except for transient events, such as wound healing angiogenic switch is thrown, the tumor hooks up to the or menorrhea [247]. However, angiogenesis takes place body’s vascular system and thus resumes its growth. under many pathological conditions, such as atherosclerosis, In tumor development, the establishment of an angio- endometriosis, osteomyelitis, diabetic retinopathy, rheuma- genic phenotype is a crucial and general step [232–234]. toid arthritis, psoriasis, and tumor growth [230]. During Depending on tumor type and environment, this induction tumor progression, the quiescent vasculature becomes per- of new vessel sprouting can occur at different stages of manently activated to sprout new vessels that enable blood the tumor progression pathway, and it leads to exponential supply and thus help sustain tumor growth [5, 6]. Due to macroscopic tumor growth [2, 4, 6]. In addition, recent data its increased metabolic rate, tumor tissue requires blood indicate that angiogenesis also contributes to the microscopic supply for expansive growth, which is circumstantiated by premalignant phase of neoplastic progression [5]. the observation that tumor cells, which are p53 deficient Infiltration of bone-marrow-derived monocytes that and thus show a reduced apoptosis rate, die beyond an differentiate into macrophages can trigger this angiogenic oxygen diffusion limit in the range of 150 μm[248]. Tumor switch in spontaneous tumors by releasing both numerous cells proliferate around the continuously formed neovessels proangiogenic cytokines, for example, VEGF, TNFα,IL-8, which markedly differ from normal vessels in morphology and bFGF [235, 236] and MMPs (e.g., MMPs-2, -7, and and molecular composition [219, 249]. Tumor vasculature -9) together with elastase and uPA [236]. These matrix- generally appears highly tortuous, chaotic, and disorganized. degrading enzymes loosen the avascular ECM for the angio- The vessels themselves are leaky due to a discontinuous genic ingrowth of neovessels. endothelium, a poorly formed BM, and a lack of mural From the multitude of proangiogenic molecules, such as cells. In addition, tumor cells sometimes mimic ECs. This FGF-1 and -2, G-CSF, HGF, IL-8, PD-ECGF, PGE-1 and -2, poor quality of tumor-associated blood vessels compromises PlGF-1, and -2, TGF-α and -β,TNF-α, and VEGF-A through blood flow, impairs drug delivery, and facilitates tumor E, only the VEGFs and PlGFs are specific for ECs [230]. cell intravasation leading to hematogenous or lymphatic VEGF-A, which exists in five splice variants, is the most metastasis. In addition to histological vessel malformations, Journal of Oncology 9 tumor vessels show an anomalous composition of their ECM, for example, tenascin-C and –W, and the oncofetal fibronectin ED-B splice variants are associated with tumor vessels [250, 251]. ED-B fibronectin is synthesized by neo- plastic cells [252]. Melanoma and glioblastoma cells secrete tenascin-C as do cancer-associated fibroblasts (CAF) of most carcinomas [253]. Tenascin-C stimulates angiogenesis in ECs, mediates survival of tumor stem cells, enhances proliferation, invasiveness, and metastasis in tumor cells, and blocks immunosurveillance [250, 253]. Tenascin-W is more strictly associated with tumorigenesis and can be used as a tumor biomarker for breast and colon cancer, because it is undetectable in healthy stroma but overexpressed in the tumor stroma [254, 255]. Vascularization mechanisms in cancer have been re- viewed recently [256, 257]. New tumor blood vessels can either arise by vessel co-option or be formed by tumor angiogenesis, but there is also evidence for vasculogenesis or recruitment of circulating bone-marrow-derived endothelial progenitor cells that differentiate into ECs [230, 258–260] (Figure 2(A)). Depending on the tumor type, tumor blood F vessels build different and characteristic vascular beds, and, according to the function of the vascular bed and Endothelial cell (EC) Tip cell the osmotic pressure of the surrounding tissue, endothelia Endothelial progenitor cell (EPC) Tumor cell represent highly heterogeneous “vascular addresses” [230]. Tumor vessels constantly change their shape due to persis- EPC-derived cell tent growth, and about 30% of the vasculature comprise Figure 2: Diverse types of vessel formation. Tumor neovasculariza- arteriovenous shunts bypassing capillaries. The concomitant tion can take place by distinct types of vessel formation, which can poor perfusion leads to hypoxia of ECs, which consequently proceed simultaneously and also merge seamlessly. (A) Neovessel synthesize more proangiogenic molecules and thus crank formation by recruitment of bone-marrow-derived endothelial tumor angiogenesis [230]. progenitor cells. (B) Sprouting angiogenesis is initiated by the differentiation of an EC into a migratory but nonproliferating tip 3.2.1. Endothelial Sprouting. Endothelial sprouting can be cell. (C) Intussusceptive angiogenesis starts with the insertion of a triggered by hypoxia, hypoglycemia, and inflammatory or connective tissue pillar into a preexisting vessel, and the vessel is dis- mechanical stimuli, such as blood pressure, and is regulated placed as the pillar extends in size. (D) In glomeruloid angiogenesis, by many angiogenic growth factors, such as VEGF, and complex vascular aggregates of several closely associated vessels are matrix proteases. When neovessels sprout from capillaries, formed. (E) Vessel co-option is the acquisition of host capillaries by the tumor. (F) In vascular mimicry, tumor cells can partly assume pericytes are selectively lost, and upon receiving an angio- EC function and form vessel-like hollow structures. Arrows denote genic stimulus, select ECs differentiate into tip cells that consecutive stages of vessel formation. Tumor tissue is depicted dark invade the avascular ECM (Figure 2(B)). These tip cells gray. See text for details. migrate into the ECM following the stimulatory gradient. Behind the tip cells, other ECs begin to proliferate and, as stalk cells, form cord-like structures. These develop cell invasion [266]. In addition to VEGF, FGF, PDGF, and into endothelial tubes [130, 261, 262] that subsequently PlDGF are involved, and Ang-2/Tie-2 signaling regulates the anastomose and thus allow blood flow. Finally, pericytes and detachment of pericytes. Later, PDGF-BB recruits pericytes smooth muscle cells are recruited, a new BM is synthesized, andsmoothmusclecells to the newlyformedECtube, and the ECs become quiescent again. and TGF-β1 and Ang-1/Tie-2 stabilize the EC-mural cell The molecular background of capillary sprouting and interaction [231]. the key role of VEGF have been reviewed by Carmeliet [231]. Upon a hypoxic stimulus, VEGF is produced, and as a consequence the endothelium’s permeability is increased 3.2.2. Intussusceptive Angiogenesis. Another way of tumor and the BM loosened by the activity of MMPs [243, 263] neovascularization is intussusceptive angiogenesis, which and the urokinase plasminogen activator system [264]. The represents a nonproliferative and noninvasive mechanism MMP inducer EMMPRIN/CD147 also upregulates soluble for the enlargement of a capillary plexus by intussusceptive VEGF isoforms 121 and 165 and VEGFR-2 on ECs and growth, arborization, and remodeling [267](Figure 2(C)). thus promotes sprouting angiogenesis [265]. Integrin αvβ3 As this mode of vascularization is mostly independent from mediates migration into the fibrin-rich cancer stroma and EC proliferation and migration, as well as BM degradation, furthermore can associate with MMP-2, thus enabling ECs this process is more economical and, occurring within to maintain the BM in the sol state and to promote tumor hours or even minutes, is noticeably faster than sprouting 10 Journal of Oncology angiogenesis [268]. It begins with the formation of translu- of angiogenesis, and they can arise by two types of vascu- minal pillars from the EC walls. Their subsequent expansion logenic mimicry, designated the tubular and the patterned splits the preexisting vessel into two, thereby enhancing the matrix type [277]. These tubular vessel-like networks resem- vascular surface. In a subsequent process of arborization, ble the pattern of embryonic vascular networks, and, in the disorganized capillary network is remodeled into a their gene expression pattern, aggressive tumors that form functional tree-like structure by serial pillar formation. In such channels resemble endothelial, pericytes, and other a final remodeling step, the branching angles are modified, precursor stem cells, suggesting that tumor cells might and the capillary network is pruned. The formation of new disguise as embryonic stem-cell-like or other cell types capillaries is initiated by sprouting angiogenesis that is later [256]. Vasculogenic mimicry of the patterned matrix type accompanied or followed by intussusceptive angiogenesis, looks completely different and is characterized by a fluid- which increases the EC surface [269]. Intussusceptive angio- conducting meshwork of extravascular patterned depositions genesis is synergistically regulated by VEGF and Ang-1, of matrix proteins such as laminins, collagens IV and VI, and and it seems to be induced by laminar shear stress on the heparin sulfate proteoglycans that anastomose with blood vessel walls, whereas oscillating shear stress favors sprouting vessels [277–279]. Although it is not yet elucidated how such angiogenesis [269]. channels are connected to the vasculature, the latter type of vascular mimicry has been reported for many cancers, such as breast, ovarian, and prostate carcinoma, melanoma, 3.2.3. Glomeruloid Angiogenesis. In many aggressive tumors, soft tissue sarcomas, osteosarcoma, and phaeochromocy- glomeruloid angiogenesis gives rise to complex vascular toma [277, 280]. In aggressive melanoma, the expression structures termed glomeruloid bodies, in which several of tissue factor pathway-associated genes, such as tissue microvessels together are ensheathed by a BM of varying factor (TF), TF pathway inhibitor-1 (TFPI-1), and TFPI- thickness containing sparse pericytes [270](Figure 2(D)). 2, is upregulated, suggesting an anticoagulation mechanism The frequency of occurrence of such glomeruloid bodies in the channel-forming tumor cells [281]. Fluid propelled is an indication for the tumor’s aggressiveness and the through these channels by a pressure gradient might facilitate patient’s survival [271]. The formation of such glomeruloid the supply with nutrients and oxygen, and, additionally, this bodies is rather a remodeling than true angiogenesis, because fluid-conducting network could substitute for a lymphatic proliferating and migrating tumor cells can actively pull vascular system and drain extravasated interstitial fluid capillaries of the surrounding host vasculature and adjacent in tumors that lack lymphatic vessels, for example, uveal capillary branching points into the tumor node. Thereby, melanoma [279, 280]. formed coiled vascular structures develop subsequently into glomeruloid bodies that are connected to the surrounding vasculature via numerous narrowed capillaries [256]. 4. Manipulation of Cell Matrix Interaction in Tumor Angiogenesis 3.2.4. Vessel Co-Option. Malignant cells can initially grow Cell-matrix interactions regulate signaling pathways that are in the vicinity and along pre-existing microvessels and thus intricately interconnected with cytokine-regulated pathways, use the host vasculature for their own benefit (Figure 2(E)). which complicates the analysis of their contribution to a This co-option of the host vasculature was originally believed particular step in angiogenesis [153]. ECM receptors can to be limited to the initial phase of tumorigenesis [272]. be manipulated with a wide variety of different compounds Meanwhile, however, there is evidence that vessel co-option ranging from endogenous compounds, such as matrikines, might persist during all stages of primary and metastatic over their synthetic analogues and peptides mimicking only growth of various tumors [256], for example, cutaneous integrin-binding sites to function-blocking antibodies and melanoma, which appears to grow by co-opting the vascular small molecules with integrin inhibitory function. Other plexus in its surrounding connective tissue, while there is no starting points for an antiangiogenic therapy are the inhibi- sign of directed vessel ingrowth [273]. tion of signaling cascades downstream of the ECM receptors Vessel co-option is regulated dependent on the tumor or cytokine receptors and as a new avenue the blocking type and the host environment, but the key regulators are of microRNAs with antisense RNAs in ECs [282, 283]. An again VEGF and angiopoietins [272, 274]. Ang-1 binds to efficient antivascular cancer therapy can target either the Tie-2 and thus triggers signaling cascades, assuring survival angiogenic signaling pathways or the vascularization mech- and quiescence of ECs, and thus causing tumor vessel anism [256]. A combination of conventional chemotherapy maintenance, whereas the nonsignaling Tie-2 ligand Ang-2 with angiosuppressive or vascular disrupting therapy is often acts as a negative regulator and destabilizes the capillary walls problematic and needs careful design [256]. by detachment of pericytes [272, 274]. Subsequently, VEGF via its receptor VEGFR-2 promotes both survival of ECs and growth of new vessels [237, 275]. 4.1. Pharmacological Intervention of Integrin-ECM Interac- tion. In addition to soluble growth factors, such as VEGF, 3.2.5. Vascular Mimicry. Aggressive melanomas can form there are several endogenous angiogenesis inhibitors, for fluid-filled vessel-like channels without any EC lining in example, endostatin, endorepellin, and tumstatin, which a nonangiogenic process termed vascular mimicry [276] share the common feature that they all are proteolytic frag- (Figure 2(F)). These channels allow perfusion independent ments of ECM molecules [284, 285]. In tumor angiogenesis Journal of Oncology 11 within a primary tumor, such ECM fragments are generated In a phase I clinical trial, endostatin, the C-terminal by the release of MMPs, in order to degrade the BM. This fragment of collagen XVIII, blocks the function of integrin results not only in labile and leaky tumor vessels but at α5β1[179, 304, 305] and also binds to heparin and with the same time keeps metastases from growing, as these lower affinities to other heparan sulfate proteoglycans that endogenous angiogenesis inhibitors are distributed via the are involved in growth factor signaling [306, 307]. Endo- blood stream [230]. Therefore, they are of pharmacological statin’s antiangiogenic activity can also be mimicked with interest with regard to their use as angiogenesis inhibitors. derived short non-RGD but arginine-rich peptides [308]. Intensive efforts have been directed towards the development Integrin α5β1 can also be blocked by the synthetic non- of integrin antagonists for the treatment of cancer and many RGD peptides PHSCN, named ATN-161, [309] and cyclic other diseases, ranging from autoimmune diseases over CRRETAWAC [310], as well as by the peptide mimetics SJ749 inflammatory to thrombotic diseases, and their applications [311] and JSM6427 [312], and it can be inhibited by the seem promising [11, 286]. Integrin-mediated interactions affinity-matured humanized chimeric monoclonal antibody of cells with their surrounding ECM can be manipulated M200/volociximab [313]. by antibodies, peptides, small nonpeptidic compounds, and Angiostatin is a proteolytic fragment of plasminogen that endogenous inhibitors (Figure 3). Integrin antagonists with effectively inhibits integrin αvβ3[314], and its antiangio- antiangiogenic activities have been reviewed recently with genic effect can also be achieved by its isolated kringle-5 special emphasis on drugs that are in clinical trials [11]. domain [315]. Kringle-1 to 3 show the same antiproliferative Spurred by the success in pharmacologically targeting effect as the whole angiostatin, but hardly inhibit migration, RGD-dependent integrins, there are also attempts to phar- whereas kringle-4 inhibits EC migration but shows only a macologically manipulate RGD-independent integrins, such marginal antiproliferative effect [316]. Other endogenous as the collagen- and laminin-binding integrins, as reviewed integrin αvβ3 inhibitors are the collagen XVIII fragment recently [287]. The collagen-binding subgroup of integrins endostatin [304], and the C-terminal fragment of the with their common A domain comprises interesting targets collagen IV α3-chain termed tumstatin [317], which also in the development of drugs against thrombosis, inflam- binds to integrin α6β1[318]. Tumstatin has two binding matory diseases, and cancer. TSPs-1 and -2 are naturally sites for integrin αvβ3. The N-terminal site mediates an occurring potent angiogenesis inhibitors, and their anti- antiangiogenic signal, whereas the C-terminal binding site angiogenic effects can be imitated by short-peptide mimetics is associated with the antitumor cellactivity [318, 319]. that among other targets bind to β1 integrins [288, 289]. Canstatin, the NC1 domain of the collagen IV α2 chain, An endogenous inhibitor, which blocks the interaction inhibits both integrins αvβ3and αvβ5[320] and seems of integrin α1β1 with collagen I and also binds to heparan to interact with integrin α3β1too [321]. A hemopexin- sulfate proteoglycans, is arresten, the C-terminal fragment like domain comprising C-terminal fragment of MMP-2, of the collagen IV α1 chain [290, 291]. Endorepellin, a termed PEX, also antagonizes integrin αvβ3 by preventing C-terminal fragment of perlecan specifically blocks the its binding to MMP-2 and thus inhibiting proteolytic activity function of integrin α2β1[292] and interestingly also on the cell surface, especially during vessel maturation [322, binds to endostatin, thus counteracting its antiangiogenic 323]. Fastatin and other FAS1 domains, which are present effect [293]. Additionally, integrin α1β1 can be specifically in the four human proteins periostin, FEEL1, FEEL2, and inhibited with obtustatin from the snake venom of Vipera βhig-h3, also function via integrin αvβ3asendogenous lebetina obtusa [294, 295]. The interaction of integrin α2β1 regulators of pathogenic angiogenesis [324]. Next to these with collagen can be specifically inhibited with the C- natural antagonists there is a variety of synthetic RGD— type lectin rhodocetin from the snake venom of Callose- containing peptide inhibitors that mimic a motif that occurs lasma rhodostoma [296, 297]. In addition, it can also be on many ECM molecules, such as fibronectin, vitronectin, selectively antagonized by the protein angiocidin, which fibrinogen, osteopontin, TSP, vWF, and partially degraded was first detected in lung carcinoma cells [298, 299]. The collagen. Most integrins of the αv subfamily and the integrins aromatic tetracyclic polyketides maggiemycin and anhydro- α5β1and αIIbβ3 bind to this motif. Therefore, adhesion and maggiemycin from Streptomyces, which have been described spreading of ECs to the ECM can be competitively inhib- as potential antitumor agents [300], inhibit collagen binding ited by RGD peptides, whereby anchorage-dependent ECs by blocking the A domain of the integrin subunits α1, undergo apoptosis [230]. To this group belong compounds, α2, α11, and to a lesser extent α10 while cell adhesion such as cilengitide/EMD121974 [325], S137 and S247 [326, to fibronectin, mediated by integrins α5β1, αvβ3, αvβ5, 327], the TSP-derived peptide TP508/chrysalin [328], and is unaffected [301]. Recently, the sulfonamide derivative several integrin αvβ3- and αvβ5-specific peptidomimetics, BTT-3016 has been described as a potent antithrombotic such as BCH-14661, which preferentially inhibits αvβ3and small-molecule inhibitor of integrin α2β1 with only slight BCH-15046, which blocks αvβ3, αvβ5, and α5β1[329], effect on other collagen-binding integrins and no effect on SCH221153 [330], and ST1646 [331]. Another inhibitor fibronectin- or vitronectin-binding integrins [302]. Another is the non-peptide antibiotic thiolutin, which intracellu- sulfonamide derivative, E7820, which does not interfere with larly blocks paxillin and thus, indirectly, integrin αvβ3- integrin-ligand interaction, reduces integrin α2 expression mediated adhesion to vitronectin [332]. Antibodies against on the mRNA level [303]. Angiogenesis can be inhibited with the β3 subunit inhibit contact of ECs to vitronectin and antibodies against the α subunits of the integrins α1β1and concomitantly VEGF-induced tyrosine phosphorylation of α2β1, whereas quiescent vessels are not affected [230]. VEGFR-2 in cell culture studies [333]. Moreover, integrin 12 Journal of Oncology ECM Matrikines Peptides Antibodies Small molecule inhibitors α β Adhesion Survival Migration Proliferation Figure 3: Options to manipulate integrin function. Essential cellular functions, such as adhesion, migration, proliferation, and survival, which all are regulated by integrins, can pharmacologically be manipulated with a panoply of matrikines, antibodies, peptides, and small molecule inhibitors, many of which are used as therapeutic tools in combination with conventional chemo- or radiotherapy to attack tumor cells and vasculature. Details are described in the text. αvβ3can be effectively antagonized with the monoclonal the RGD-containing inhibitors cilengitide and S 36578 alter antibody LM609/MEDI-552 and its humanized derivative the trafficking of integrins and VEGFR2 in tumor ECs, thus abegrin/etaracizumab/vitaxin [57, 334–337]. In contrast, stimulating angiogenesis and tumor growth [342]. the humanized anti-αv antibody CNTO95 targets both Current tumor therapy aims at vessel eradication in order integrins αvβ3and αvβ5[338]. The humanized Fab fragment to disrupt the connection of the tumor to the vascular 17E6/abciximab/ReoPro of the monoclonal antibody c7E3 system and thus cut off the supply of nutrients and oxygen. inhibits the integrins αvβ3 and also αMβ2/Mac-1 [339, This can be done with compounds that preferentially affect 340], whereas the human-specific monoclonal antibody tumor endothelia rather than normal cells, that is, (i) specific 17E6 targets all αv integrins [341]. Currently, humanized or angiogenesis inhibitors, (ii) tumor vessel toxins that attack chimeric integrin antibody antagonists of αvβ3, αvβ5, and inherent weaknesses in static tumor vessel endothelia and α5β1, and peptide inhibitors of these integrins are in clinical associated vascular structures, and (iii) dual-action com- trials as antiangiogenic agents [180]. pounds [343]. However, within the last years, a paradigm shift has taken place [344, 345]. Vessel normalization by pruning immature vessels and increasing pericytes and 5. Applications and Outlook BM coverage of the remaining vessels comes to the fore, Integrins and their binding partners are of special interest rather than vessel eradication, because mere antiangiogenic treatment can worsen malignancy [346]. A malformed as potential therapeutic targets, and several are already in clinical trials. However, the results fall short of the initial tumor vasculature creates and aggravates a hypoxic and expectations, pointing out that monotherapy with a single acidic milieu which hampers drug delivery and perfusion [347–349], and, due to its leaky endothelium, it promotes angiogenesis inhibitor is not sufficient to counteract the numerous angiogenic factors involved in tumor progres- tumor cell dissemination [346]. Therefore, chemotherapeu- sion [231]. Moreover, there are some caveats in aiming tic efficacy can be ameliorated by a concomitant vessel normalization therapy which improves delivery and efficacy at integrins as therapeutic targets. Obviously, integrins are expressed on virtually all cells under physiological as of cytotoxic drugs and also sensitizes the tumor cells to radiation [345, 350]. well as pathological conditions, and it is a major chal- lenge to target exclusively integrins on tumor or tumor- In vessel normalization, the interaction of cells with associated cells. Another problem is that low concentrations their surrounding ECM via integrins is of special impor- tance. However, many antiangiogenic compounds, for exam- of antagonists alter the signaling of integrins and other receptors. When administered in nanomolar concentrations, ple, ATN-161, endostatin, and integrin inhibitors, show Journal of Oncology 13 hormetic, that is, bell- or U-shaped, dose-response curves Additionally, integrins can be used as biomarkers to non- and thus present a challenge for clinical translation [351]. invasively assess the efficacy of chemotherapeutic and radio- Nanomolar concentrations of RGD-mimetic αvβ3and αvβ5 therapeutic drugs [12]. Integrin-targeted probes can be inhibitors (S 36578 and cilengitide) can paradoxically used to visualize tumor angiogenesis and the response to stimulate tumor growth and angiogenesis by altering the chemo- and radiotherapy by various imaging methods, trafficking of αvβ3 integrin and VEGFR2. Thus, they such as magnetic resonance imaging (MRI), positron emis- promote the migration of ECs towards VEGF, which has sion tomography (PET), and ultrasonography [360–362]. important implications for the use of RGD mimetics in Moreover, fluorescence labeling of integrin ligands allows tumor therapy [342]. Thus, depending on tumor type, dose, intraoperative fluorescence imaging, thus providing a tool and manner of application, the currently available-integrin to intraoperatively detect and remove metastases of sub- targeting compounds can act either anti- or proangiogenic. millimeter size [363]. A promising approach may be a combination therapy that In summary, the above data illustrate the importance blocks simultaneously angiogenic integrin αvβ3 and VEGFR of integrins and integrin-binding and signaling proteins in activities [352–355]. both physiological and pathological blood vessel formation. To circumvent these problems, instead of targeting the Thus, they may be potential targets for antiangiogenic tumor integrins, which are in principle present on both normal and therapy. Although our knowledge concerning this matter has malignant cells, another strategy aims at tumor-promoting increased remarkably within the last years, the understanding integrin ligands, such as ED-B fibronectin, tenascin-C, and is far from complete. tenascin W [252, 253, 255]. Invasive tumor cells partially degrade and denature their surrounding ECM, and the thereby released cryptic collagen IV epitope HU177 may also Abbreviations be a potential target for antiangiogenic and tumor-selective drug delivery [356]. Ang: Angiopoietin In comparison to a systemic administration of a chemo- BM: Basement membrane therapeutic agent, its therapeutic index can be increased CRDGK, Amino acid sequences in single letter by selectively targeting integrins that are overexpressed on CRGDKGPDC, code tumor cells [357]. Chemotherapeutic small molecules, pep- CRRETAWAC, tides, and proteins as well as nanoparticle-carried chemo- PHSCN, RGD: therapeutics, which are conjugated to ligands of integrins ECM: Extracellular matrix that are overexpressed on angiogenic ECs or tumor cells, can EC: Endothelial cell be selectively internalized after integrin binding [357]. Espe- FAK: Focal adhesion kinase cially nanoparticles, such as micelles, liposomes, polymeric FGF: Fibroblast growth factor nanospheres, and polymersomes loaded with chemothera- G-CSF: Granulocyte colony-stimulating peutic or radiotherapeutic drugs and equipped with multi- factor valent integrin ligands show decreased systemic toxicity, pro- HGF: Hepatocyte growth factor longed half-life and passive retention in the tumor, improved HIF: Hypoxia-inducible factor binding affinity, and facilitated internalization, thus resulting IL: Interleukin in increased drug delivery [12, 357, 358]. A therapeutic MAPK: Mitogen-activated protein kinase strategy that targets several integrins and receptors by such MEK: MAPK/ERK kinase chemo-, radio-, and possibly gene therapeutic approaches MMP: Matrix metalloproteinase maybemoreeffective than a monotherapy [231, 357]. NC: Noncollagenous Coadministration of the αv integrin-targeting cyclic pep- NF-κB: Nuclear factor κ-light-chain tide iRGD (CRGDKGPDC), or structurally closely related enhancer of activated B cells peptides, with anticancer drugs considerably enhances their NRP-1: Neuropilin-1 efficacy and selectivity [359]. Upon binding to αv integrin- PDGF: Platelet-derived growth factor expressing tumor ECs, iRGD is proteolytically processed to PD-ECGF: Platelet-derived endothelial cell CRDGK with a much weaker integrin affinity, whereas this growth factor truncated peptide shows an increased affinity to neuropilin- PGE: Prostaglandin E 1 (NRP-1), thus increasing vascular and tissue perme- PI3K: Phosphatidylinositol-3 kinase ability in a tumor-specific and NRP-1-dependent manner PLC: Phospholipase C [359]. Interestingly, this coadministration does not require PlGF: Placenta-derived growth factor chemical conjugation of the drug with the iRGD peptide; Ras: Rat sarcoma protein that is, approved drugs could be used unmodified [359]. Src: Sarcoma oncogene Coadministration of such a tumor-penetrating peptide with TGF: Transforming growth factor either small molecules, such as doxorubicin, antibodies, such Tie: Tyrosine kinase with as trastuzumab, or nanoparticles, such as Nab-paclitaxel immunoglobulin-like and EGF-like (abraxane) or doxorubicin-loaded liposomes, resulted in domain equivalent or increased delivery and efficacy, and it improved TIMP: Tissue inhibitor of their therapeutic index by lowering the effective dose [359]. metalloproteinases 14 Journal of Oncology [15] J. A. Eble and S. Niland, “The extracellular matrix of blood vessels,” Current Pharmaceutical Design, vol. 15, no. 12, pp. TNF: Tumor necrosis factor 1385–1400, 2009. uPA(R): Urokinase-type plasminogen activator [16] V. S. LeBleu, B. MacDonald, and R. Kalluri, “Structure and (receptor) function of basement membranes,” Experimental Biology and VCAM: Vascular cell adhesion molecule Medicine, vol. 232, no. 9, pp. 1121–1129, 2007. VEGF(R): Vascular endothelial growth factor [17] R. V. Iozzo, J. J. Zoeller, and A. Nystrom, ¨ “Basement mem- (receptor) brane proteoglycans: modulators Par excellence of cancer VSMC: Vascular smooth muscle cell growth and angiogenesis,” Molecules and Cells, vol. 27, no. 5, vWF: Von Willebrand factor pp. 503–513, 2009. TSP: Thrombospondin. [18] P. D. Yurchenco, “Basement membranes: cell scaffoldings and signaling platforms,” Cold Spring Harbor Perspectives in Biology, vol. 3, no. 2, 2011. Acknowledgments [19] S. Astrof and R. O. Hynes, “Fibronectins in vascular morpho- genesis,” Angiogenesis, vol. 12, no. 2, pp. 165–175, 2009. The authors thank the Deutsche Forschungsgemeinschaft [20] S. Katsuda and T. Kaji, “Atherosclerosis and extracellular for financial support (SFB/TR23 A8 Eble). They sincerely matrix,” Journal of Atherosclerosis and Thrombosis, vol. 10, no. apologize to authors of important work not cited here for 5, pp. 267–274, 2003. reasons of space limitation. [21] G. A. M. Plenz, M. C. Deng,H.Robenek,and W. Volk ¨ er, “Vascular collagens: spotlight on the role of type VIII References collagen in atherogenesis,” Atherosclerosis, vol. 166, no. 1, pp. 1–11, 2003. [1] K. Kinzler and B. Vogelstein, The Genetic Basis of Human [22] K. Kuhn, ¨ “Basement membrane (type IV) collagen,” Matrix Cancer, McGraw-Hill, Medical Pub. Division, 2nd edition, Biology, vol. 14, no. 6, pp. 439–445, 1995. [23] E. Posc ¨ hl, U. Schlotz ¨ er-Schrehardt, B. Brachvogel, K. Saito, [2] G. Bergers and L. E. Benjamin, “Tumorigenesis and the Y. Ninomiya, and U. Mayer, “Collagen IV is essential for angiogenic switch,” Nature Reviews Cancer,vol. 3, no.6,pp. basement membrane stability but dispensable for initiation 401–410, 2003. of its assembly during early development,” Development, vol. [3] S. Rheingold, A. Neugut, and A. Meadows, “Secondary can- 131, no. 7, pp. 1619–1628, 2004. cers: incidence, risk factors, and management,” in Holland- [24] S. M. Mithieux and A. S. Weiss, “Elastin,” Advances in Protein Frei Cancer Medicine, D. Kufe, R. Pollock, and R. Weichsel- Chemistry, vol. 70, pp. 437–461, 2005. baum, Eds., p. 2399, B. C. Decker, Hamilton, Ont, Canada, [25] C. M. Kielty, “Elastic fibres in health and disease,” Expert Reviews in Molecular Medicine, vol. 8, no. 19, pp. 1–23, 2006. [4] D. Hanahan and R. A. Weinberg, “The hallmarks of cancer,” [26] A. Patel, B. Fine, M. Sandig, and K. Mequanint, “Elastin Cell, vol. 100, no. 1, pp. 57–70, 2000. biosynthesis: the missing link in tissue-engineered blood [5] D. Hanahan and R. A. Weinberg, “Hallmarks of cancer: the vessels,” Cardiovascular Research, vol. 71, no. 1, pp. 40–49, next generation,” Cell, vol. 144, no. 5, pp. 646–674, 2011. [6] D. Hanahan and J. Folkman, “Patterns and emerging mech- [27] A. Colombatti, R. Doliana, S. Bot et al., “The EMILIN protein anisms of the angiogenic switch during tumorigenesis,” Cell, family,” Matrix Biology, vol. 19, no. 4, pp. 289–301, 2000. vol. 86, no. 3, pp. 353–364, 1996. [28] B. S. Brooke, S. K. Karnik, and D. Y. Li, “Extracellular [7] B. Nico, E. Crivellato, D. Guidolin et al., “Intussusceptive matrix in vascular morphogenesis and disease: structure microvascular growth in human glioma,” Clinical and Exper- versus signal,” Trends in Cell Biology, vol. 13, no. 1, pp. 51– imental Medicine, vol. 10, no. 2, pp. 93–98, 2010. 56, 2003. [8] C. J. Avraamides, B. Garmy-Susini, and J. A. Varner, “Inte- [29] R. Timpl, T. Sasaki, G. Kostka, and M. L. Chu, “Fibulins: grins in angiogenesis and lymphangiogenesis,” Nature Re- a versatile family of extracellular matrix proteins,” Nature views Cancer, vol. 8, no. 8, pp. 604–617, 2008. Reviews Molecular Cell Biology, vol. 4, no. 6, pp. 479–489, [9] R. Rathinam and S. K. Alahari, “Important role of integrins in the cancer biology,” Cancer and Metastasis Reviews, vol. 29, [30] T. Nakamura, P. R. Lozano, Y. Ikeda et al., “Fibulin-5/DANCE no. 1, pp. 223–237, 2010. is essential for elastogenesis in vivo,” Nature, vol. 415, no. [10] G. Alghisi and C. Ruegg ¨ , “Vascular integrins in tumor angio- 6868, pp. 171–175, 2002. genesis: mediators and therapeutic targets,” Endothelium, vol. [31] M. Hirai, T. Ohbayashi, M. Horiguchi et al., “Fibulin- 13, no. 2, pp. 113–135, 2006. 5/DANCE has an elastogenic organizer activity that is [11] C. Ruegg ¨ and G. C. Alghisi, “Vascular integrins: therapeutic abrogated by proteolytic cleavage in vivo,” Journal of Cell and imaging targets of tumor angiogenesis,” Recent Results in Biology, vol. 176, no. 7, pp. 1061–1071, 2007. Cancer Research, vol. 180, pp. 83–101, 2010. [12] J. S. Desgrosellier and D. A. Cheresh, “Integrins in can- [32] R. Giltay, R. Timpl, and G. Kostka, “Sequence, recombinant expression and tissue localization of two novel extracellular cer: biological implications and therapeutic opportunities,” Nature Reviews Cancer, vol. 10, no. 1, pp. 9–22, 2010. matrix proteins, fibulin-3 and fibulin-4,” Matrix Biology, vol. 18, no. 5, pp. 469–480, 1999. [13] S.-H. Kim, J. Turnbull, and S. Guimond, “Extracellular matrix and cell signalling: the dynamic cooperation of [33] K. P. Dingemans, P. Teeling, J. H. Lagendijk, and A. E. integrin, proteoglycan and growth factor receptor,” Journal Becker, “Extracellular matrix of the human aortic media: of Endocrinology, vol. 209, no. 2, pp. 139–151, 2011. an ultrastructural histochemical and immunohistochemical [14] L. Gartner and J. Hiat, Color Atlas of Histology,Williams& study of the adult aortic media,” Anatomical Record, vol. 258, Wilkins, Baltimore, Md, USA, 1994. no. 1, pp. 1–14, 2000. Journal of Oncology 15 [34] A. G. Marneros and B. R. Olsen, “Physiological role of sarcomas,” American Journal of Pathology, vol. 142, no. 4, pp. collagen XVIII and endostatin,” FASEB Journal, vol. 19, no. 1009–1018, 1993. 7, pp. 716–728, 2005. [51] D. R. Senger, K. P. Claffey,J.E.Benes,C.A.Perruzzi, ¨ ¨ ¨ A. P. Sergiou, and M. Detmar, “Angiogenesis promoted by [35] S. Grassel, C. Unsold,H.Schacke, L. Bruckner-Tuderman, vascular endothelial growth factor: regulation through α1β1 and P. Bruckner, “Collagen XVI is expressed by human dermal fibroblasts and keratinocytes and is associated with and α2β1 integrins,” Proceedings of the National Academy of Sciences of the United States of America, vol. 94, no. 25, pp. the microfibrillar apparatus in the upper papillary dermis,” Matrix Biology, vol. 18, no. 3, pp. 309–317, 1999. 13612–13617, 1997. [52] D. R. Senger, C. A. Perruzzi, M. Streit, V. E. Koteliansky, A. [36] A. Kassner, U. Hansen, N. Miosge et al., “Discrete integration R. De Fougerolles, and M. Detmar, “The α1β1and α2β1 of collagen XVI into tissue-specific collagen fibrils or beaded integrins provide critical support for vascular endothelial microfibrils,” Matrix Biology, vol. 22, no. 2, pp. 131–143, growth factor signaling, endothelial cell migration, and tumor angiogenesis,” American Journal of Pathology, vol. 160, [37] E. P. Moiseeva, “Adhesion receptors of vascular smooth no. 1, pp. 195–204, 2002. muscle cells and their functions,” Cardiovascular Research, [53] K. Vuoriluoto, G. Hog ¨ nas, ¨ P. Meller, K. Lehti, and J. Ivaska, vol. 52, no. 3, pp. 372–386, 2001. “Syndecan-1 and -4 differentially regulate oncogenic K-ras [38] J. A. Eble, A. Kassner, S. Niland, M. Morgelin, ¨ J. Grifka, and S. dependent cell invasion into collagen through α2β1 integrin Grassel, ¨ “Collagen XVI harbors an integrin α1β1 recognition and MT1-MMP,” Matrix Biology, vol. 30, no. 3, pp. 207–217, site in its C-terminal domains,” Journal of Biological Chem- istry, vol. 281, no. 35, pp. 25745–25756, 2006. [54] R. O. Hynes, “Integrins: bidirectional, allosteric signaling [39] J. Thyberg, K. Blomgren, J. Roy, P. K. Tran, and U. Hedin, machines,” Cell, vol. 110, no. 6, pp. 673–687, 2002. “Phenotypic modulation of smooth muscle cells after arterial [55] J. T. Yang, H. Rayburn, and R. O. Hynes, “Cell adhesion injury is associated with changes in the distribution of events mediated by α4 integrins are essential in placental and laminin and fibronectin,” Journal of Histochemistry and cardiac development,” Development, vol. 121, no. 2, pp. 549– Cytochemistry, vol. 45, no. 6, pp. 837–846, 1997. 560, 1995. [40] A. P. Hall, “Review of the pericyte during angiogenesis [56] M. R. Morgan, M. J. Humphries, and M. D. Bass, “Synergistic and its role in cancer and diabetic retinopathy,” Toxicologic control of cell adhesion by integrins and syndecans,” Nature Pathology, vol. 34, no. 6, pp. 763–775, 2006. Reviews Molecular Cell Biology, vol. 8, no. 12, pp. 957–969, [41] R. H. Adams and K. Alitalo, “Molecular regulation of angio- genesis and lymphangiogenesis,” Nature Reviews Molecular [57] P. C. Brooks,R.A.F.Clark,and D. A. Cheresh, “Requirement Cell Biology, vol. 8, no. 6, pp. 464–478, 2007. of vascular integrin α(v)β3 for angiogenesis,” Science, vol. [42] H. Gerhardt and H. Semb, “Pericytes: gatekeepers in tumour 264, no. 5158, pp. 569–571, 1994. cell metastasis?” Journal of Molecular Medicine, vol. 86, no. 2, [58] R. O. Hynes, “Cell-matrix adhesion in vascular develop- pp. 135–144, 2008. ment,” Journal of Thrombosis and Haemostasis,vol. 5, no.1, [43] Z. Isogai, A. Aspberg, D. R. Keene, R. N. Ono, D. P. Reinhardt, pp. 32–40, 2007. and L. Y. Sakai, “Versican interacts with fibrillin-1 and [59] C. N. Landen, T. J. Kim, Y. G. Lin et al., “Tumor-selective links extracellular microfibrils to other connective tissue response to antibody-mediated targeting of αvβ3 integrin in networks,” Journal of Biological Chemistry, vol. 277, no. 6, pp. ovarian cancer,” Neoplasia, vol. 10, no. 11, pp. 1259–1267, 4565–4572, 2002. [44] A. Aspberg, S. Adam, G. Kostka, R. Timpl, and D. Heinegar ˚ d, [60] G. E. Davis, “Affinity of integrins for damaged extracellular “Fibulin-1 is a ligand for the C-type lectin domains of matrix: α(v)β3 binds to denatured collagen type I through aggrecan and versican,” Journal of Biological Chemistry, vol. RGD sites,” Biochemical and Biophysical Research Communi- 274, no. 29, pp. 20444–20449, 1999. cations, vol. 182, no. 3, pp. 1025–1031, 1992. [45] A. I. Olin, M. Morgelin, ¨ T. Sasaki, R. Timpl, D. Heinegar ˚ d, [61] B. Nieswandt, M. Hafner, B. Echtenacher, and D. N. Mannel, ¨ and A. Aspberg, “The proteoglycans aggrecan and versican “Lysis of tumor cells by natural killer cells in mice is impeded form networks with fibulin-2 through their lectin domain by platelets,” Cancer Research, vol. 59, no. 6, pp. 1295–1300, binding,” Journal of Biological Chemistry, vol. 276, no. 2, pp. 1253–1261, 2001. [62] P. C. Brooks,S.Stromblad,R.Klemke, D. Visscher, F. H. [46] J. Heino and J. Kap ¨ yla, ¨ “Cellular receptors of extracellular Sarkar, and D. A. Cheresh, “Antiintegrin αvβ3blockshuman matrix molecules,” Current Pharmaceutical Design, vol. 15, breast cancer growth and angiogenesis in human skin,” no. 12, pp. 1309–1317, 2009. Journal of Clinical Investigation, vol. 96, no. 4, pp. 1815–1822, [47] L. Contois, A. Akalu, and P. C. Brooks, “Integrins as “func- tional hubs” in the regulation of pathological angiogenesis,” [63] G. H. Mahabeleshwar, W. Feng, D. R. Phillips, and T. Seminars in Cancer Biology, vol. 19, no. 5, pp. 318–328, 2009. V. Byzova, “Integrin signaling is critical for pathological [48] R. Zaidel-Bar, S. Itzkovitz, A. Ma’ayan, R. Iyengar, and B. angiogenesis,” Journal of Experimental Medicine, vol. 203, no. Geiger, “Functional atlas of the integrin adhesome,” Nature 11, pp. 2495–2507, 2006. Cell Biology, vol. 9, no. 8, pp. 858–867, 2007. [64] X. Huang, M. Griffiths, J. Wu, R. V. Farese, and D. Sheppard, [49] R. Silva, G. D’Amico, K. M. Hodivala-Dilke, and L. E. “Normal development, wound healing, and adenovirus Reynolds, “Integrins: the keys to unlocking angiogenesis,” susceptibility in β5- deficient mice,” Molecular and Cellular Arteriosclerosis, Thrombosis, and Vascular Biology, vol. 28, no. Biology, vol. 20, no. 3, pp. 755–759, 2000. 10, pp. 1703–1713, 2008. [65] B. L. Bader, H. Rayburn, D. Crowley, and R. O. Hynes, “Ex- [50] M. Miettinen, R. Castello, E. Wayner, and R. Schwarting, tensive vasculogenesis, angiogenesis, and organogenesis pre- “Distribution of VLA integrins in solid tumors: emergence cede lethality in mice lacking all αv integrins,” Cell, vol. 95, of tumor-type- related expression patterns in carcinomas and no. 4, pp. 507–519, 1998. 16 Journal of Oncology [66] J. H. McCarty, R. A. Monahan-Earley, L. F. Brown et al., of collagenase (MMP-1) and collagen α1(I) gene expression,” “Defective associations between blood vessels and brain Journal of Biological Chemistry, vol. 270, no. 22, pp. 13548– parenchyma lead to cerebral hemorrhage in mice lacking αv 13552, 1995. integrins,” Molecular and Cellular Biology, vol. 22, no. 21, pp. [82] O. Langholz, D. Roc ¨ kel, C. Mauch et al., “Collagen and colla- 7667–7677, 2002. genase gene expression in three-dimensional collagen lattices [67] J. H. McCarty, A. Lacy-Hulbert, A. Charest et al., “Selective are differentially regulated by α1β1and α2β1 integrins,” ablation of αv integrins in the central nervous system leads Journal of Cell Biology, vol. 131, no. 6, pp. 1903–1915, 1995. to cerebral hemorrhage, seizures, axonal degeneration and [83] H. Gardner, A. Broberg, A. Pozzi, M. Laato, and J. Heino, premature death,” Development, vol. 132, no. 1, pp. 165–176, “Absence of integrin α1β1 in the mouse causes loss of 2005. feedback regulation of collagen synthesis in normal and [68] L. E. Reynolds, L. Wyder, J. C. Lively et al., “Enhanced patho- wounded dermis,” Journal of Cell Science, vol. 112, no. 3, pp. logical angiogenesis in mice lacking β3 integrin or β3and β5 263–272, 1999. integrins,” Nature Medicine, vol. 8, no. 1, pp. 27–34, 2002. [84] F. Alves, W. Vogel, K. Mossie, B. Millauer, H. Hofler, and [69] F. D´ıaz-Gonzalez, ´ J. Forsyth, B. Steiner, and M. H. Ginsberg, A. Ullrich, “Distinct structural characteristics of discoidin “Trans-dominant inhibition of integrin function,” Molecular I subfamily receptor tyrosine kinases and complementary Biology of the Cell, vol. 7, no. 12, pp. 1939–1951, 1996. expression in human cancer,” Oncogene,vol. 10, no.3,pp. [70] K. M. Hodivala-Dilke, C. M. DiPersio, J. A. Kreidberg, and 609–618, 1995. R. O. Hynes, “Novel roles for α3β1integrinasaregulatorof [85] J. M. Auger, M. J. E. Kuijpers,Y.A.Senis,S.P.Watson, andJ. cytoskeletal assembly and as a trans-dominant inhibitor of W. M. Heemskerk, “Adhesion of human and mouse platelets integrin receptor function in mouse keratinocytes,” Journal to collagen under shear: a unifying model,” FASEB Journal, of Cell Biology, vol. 142, no. 5, pp. 1357–1369, 1998. vol. 19, no. 7, pp. 825–827, 2005. [71] D. G. Stupack, X. S. Puente, S. Boutsaboualoy, C. M. [86] L. Meyaard, “The inhibitory collagen receptor LAIR-1 Storgard, and D. A. Cheresh, “Apoptosis of adherent cells by (CD305),” Journal of Leukocyte Biology,vol. 83, no.4,pp. recruitment of caspase-8 to unligated integrins,” Journal of 799–803, 2008. Cell Biology, vol. 155, no. 4, pp. 459–470, 2001. [87] A. C. Curino, L. H. Engelholm, S. S. Yamada et al., “Intra- [72] J. Zhu, K. Motejlek, D. Wang, K. Zang, A. Schmidt, and cellular collagen degradation mediated by uPARAP/Endo180 L. F. Reichardt, “β8 Integrins are required for vascular is a major pathway of extracellular matrix turnover during morphogenesis in mouse embryos,” Development, vol. 129, malignancy,” Journal of Cell Biology, vol. 169, no. 6, pp. 977– no. 12, pp. 2891–2903, 2002. 985, 2005. [73] K. Venstrom and L. Reichardt, “Beta 8 integrins mediate [88] J. A. Kreidberg, M. J. Donovan, S. L. Goldstein et al., “Alpha interactions of chick sensory neurons with laminin-1, colla- 3 beta 1 integrin has a crucial role in kidney and lung gen IV, and fibronectin,” Molecular Biology of the Cell, vol. 6, organogenesis,” Development, vol. 122, no. 11, pp. 3537– no. 4, pp. 419–431, 1995. 3547, 1996. [74] R. Milner,J.B.Relvas, J. Fawcett, andC.Ffrench-Constant, [89] C. M. DiPersio,K.M.Hodivala-Dilke, R. Jaenisch,J.A. “Developmental regulation of αv integrins produces func- Kreidberg, andR.O.Hynes,“α3β1 integrin is required for tional changes in astrocyte behavior,” Molecular and Cellular normal development of the epidermal basement membrane,” Neuroscience, vol. 18, no. 1, pp. 108–118, 2001. Journal of Cell Biology, vol. 137, no. 3, pp. 729–742, 1997. [75] D. Mu,S.Cambier,L.Fjellbirkelandetal., “The integrin [90] U. Mayer, G. Saher, R. Fassler ¨ et al., “Absence of integrin α7 ανβ8 mediates epithelial homeostasis through MT1-MMP- causes a novel form of muscular dystrophy,” Nature Genetics, dependent activation of TGF-β1,” Journal of Cell Biology, vol. vol. 17, no. 3, pp. 318–323, 1997. 157, no. 3, pp. 493–507, 2002. [91] M. A. Stepp, S. Spurr-Michaud, A. Tisdale, J. Elwell, and I. [76] A. Kern, J. Eble, R. Golbik, and K. Kuhn, “Interaction of K. Gipson, “α6β4 integrin heterodimer is a component of type IV collagen with the isolated integrins α1β1and α2β1,” hemidesmosomes,” Proceedings of the National Academy of European Journal of Biochemistry, vol. 215, no. 1, pp. 151– Sciences of the United States of America, vol. 87, no. 22, pp. 159, 1993. 8970–8974, 1990. [77] M. Tulla, O. T. Pentikainen, T. Viitasalo et al., “Selective [92] O. Ibraghimov-Beskrovnaya, J. M. Ervasti, C. J. Leveille, C. binding of collagen subtypes by integrin α1I, α2I, and α10I A. Slaughter, S. W. Sernett, and K. P. Campbell, “Primary domains,” Journal of Biological Chemistry, vol. 276, no. 51, structure of dystrophin-associated glycoproteins linking dys- pp. 48206–48212, 2001. trophin to the extracellular matrix,” Nature, vol. 355, no. [78] M. M. Zutter and S. A. Santoro, “Widespread histologic dis- 6362, pp. 696–702, 1992. tribution of the α2β1 integrin cell-surface collagen receptor,” [93] T. Haenggi and J. M. Fritschy, “Role of dystrophin and utro- American Journal of Pathology, vol. 137, no. 1, pp. 113–120, phin for assembly and function of the dystrophin glycopro- 1990. tein complex in non-muscle tissue,” Cellular and Molecular [79] T. Bengtsson, A. Aszodi, C. Nicolae, E. B. Hunziker, E. Life Sciences, vol. 63, no. 14, pp. 1614–1631, 2006. Lundgren-Akerlund, and R. Fassler ¨ , “Loss of α10β1 integrin [94] J. Nelson, N. V. McFerran, G. Pivato et al., “The 67 kDa expression leads to moderate dysfunction of growth plate laminin receptor: structure, function and role in disease,” chondrocytes,” Journal of Cell Science, vol. 118, no. 5, pp. 929– Bioscience reports, vol. 28, no. 1, pp. 33–48, 2008. 936, 2005. [95] G. Fontanini, S. Vignati, S. Chine´ et al., “67-kilodalton [80] S. N. Popova, M. Barczyk, C. F. Tiger et al., “α11β1 integrin- laminin receptor expression correlates with worse prognostic dependent regulation of periodontal ligament function in the indicators in non-small cell lung carcinomas,” Clinical Cancer erupting mouse incisor,” Molecular and Cellular Biology, vol. Research, vol. 3, no. 2, pp. 227–231, 1997. 27, no. 12, pp. 4306–4316, 2007. [96] D. Waltregny, L. De Leval, S. Menar ´ d, J. De Leval, and V. [81] T. Riikonen, J. Westermarck, L. Koivisto, A. Broberg, V. M. Castronovo, “Independent prognostic value of the 67-kd Kahari, and J. Heino, “Integrin α2β1isapositive regulator laminin receptor in human prostate cancer,” Journal of the Journal of Oncology 17 National Cancer Institute, vol. 89, no. 16, pp. 1224–1227, [113] N. E. Vlahakis, B. A. Young, A. Atakilit et al., “Integrin α9β1 1997. directly binds to vascular endothelial growth factor (VEGF)- A and contributes to VEGF-A-induced angiogenesis,” Journal [97] E. Ardini, E. Tagliabue, A. Magnifico et al., “Co-regulation of Biological Chemistry, vol. 282, no. 20, pp. 15187–15196, and physical association of the 67-kDa monomeric laminin receptor and the α6β4 integrin,” Journal of Biological Chem- istry, vol. 272, no. 4, pp. 2342–2345, 1997. [114] N. E. Vlahakis, B. A. Young, A. Atakilit, and D. Sheppard, “The lymphangiogenic vascular endothelial growth factors [98] T. Ogawa, Y. Tsubota, J. Hashimoto, Y. Kariya, and K. VEGF-C and -D are ligands for the integrin α9β1,” Journal Miyazaki, “The short arm of laminin γ2 chain of laminin- of Biological Chemistry, vol. 280, no. 6, pp. 4544–4552, 2005. 5 (laminin-332) binds syndecan-1 and regulates cellular [115] T. R. Carlson, Y. Feng, P. C. Maisonpierre, M. Mrksich, and A. adhesion and migration by suppressing phosphorylation of O. Morla, “Direct cell adhesion to the angiopoietins mediated integrin β4 chain,” Molecular Biology of the Cell, vol. 18, no. by integrins,” Journal of Biological Chemistry, vol. 276, no. 28, 5, pp. 1621–1633, 2007. pp. 26516–26525, 2001. [99] R. G. Da Silva, B. Tavora, S. D. Robinson et al., “Endothelial [116] S. J. Leu, S. C. T. Lam, and L. F. Lau, “Pro-angiogenic α3β1-integrin represses pathological angiogenesis and sus- activities of CYR61 (CCN1) mediated through integrins tains endothelial-VEGF,” American Journal of Pathology, vol. αvβ3and α6β1 in human umbilical vein endothelial cells,” 177, no. 3, pp. 1534–1548, 2010. Journal of Biological Chemistry, vol. 277, no. 48, pp. 46248– [100] R. Van der Neut, P. Krimpenfort, J. Calafat, C. M. Niessen, 46255, 2002. and A. Sonnenberg, “Epithelial detachment due to absence of [117] S. J. Leu, Y. Liu, N. Chen, C. C. Chen, S. C. T. Lam, and L. F. hemidesmosomes in integrin β null mice,” Nature Genetics, Lau, “Identification of a novel integrin α6β1 binding site in vol. 13, no. 3, pp. 366–369, 1996. the angiogenic inducer CCN1 (CYR61),” Journal of Biological [101] S. N. Nikolopoulos, P. Blaikie, T. Yoshioka, W. Guo, and Chemistry, vol. 278, no. 36, pp. 33801–33808, 2003. F. G. Giancotti, “Integrin β4 signaling promotes tumor [118] S. Mori, C. Y. Wu, S. Yamaji et al., “Direct binding of integrin angiogenesis,” Cancer Cell, vol. 6, no. 5, pp. 471–483, 2004. αvβ3 to FGF1 plays a role in FGF1 signaling,” Journal of [102] T. S. Hiran, J. E. Mazurkiewicz, P. Kreienberg, F. L. Rice, and Biological Chemistry, vol. 283, no. 26, pp. 18066–18075, 2008. S. E. LaFlamme, “Endothelial expression of the α6β4 integrin [119] K. Suzuki, T. Okuno, M. Yamamoto et al., “Semaphorin is negatively regulated during angiogenesis,” Journal of Cell 7A initiates T-cell-mediated inflammatory responses through Science, vol. 116, no. 18, pp. 3771–3781, 2003. α1β1 integrin,” Nature, vol. 446, no. 7136, pp. 680–684, 2007. [103] S. M. Frisch and H. Francis, “Disruption of epithelial [120] J. T. Chao, L. A. Martinez-Lemus, S. J. Kaufman, G. A. cell-matrix interactions induces apoptosis,” Journal of Cell Meininger, K. S. Ramos, and E. Wilson, “Modulation of Biology, vol. 124, no. 4, pp. 619–626, 1994. α7-integrin-mediated adhesion and expression by platelet- [104] S. A. Wickstrom, ¨ K. Radovanac, and R. Fassler ¨ , “Genetic derived growth factor in vascular smooth muscle cells,” analyses of integrin signaling,” Cold Spring Harbor Perspec- American Journal of Physiology, vol. 290, no. 4, pp. C972– tives in Biology, vol. 3, no. 2, 2011. C980, 2006. [105] F. G. Giancotti and E. Ruoslahti, “Integrin signaling,” Science, [121] N. L. Flintoff-Dye, J. Welser, J. Rooney et al., “Role for vol. 285, no. 5430, pp. 1028–1032, 1999. the α7β1 integrin in vascular development and integrity,” [106] M. A. Arnaout, B. Mahalingam, and J. P. Xiong, “Integrin Developmental Dynamics, vol. 234, no. 1, pp. 11–21, 2005. structure, allostery, and bidirectional signaling,” Annual [122] Y. Taooka, J. Chen, T. Yednock, and D. Sheppard, “The Review of Cell and Developmental Biology, vol. 21, pp. 381– integrin α9β1 mediates adhesion to activated endothelial 410, 2005. cells and transendothelial neutrophil migration through [107] H. Methe, S. Hess, and E. R. Edelman, “Endothelial immu- interaction with vascular cell adhesion molecule-1,” Journal nogenicity—a matter of matrix microarchitecture,” Throm- of Cell Biology, vol. 145, no. 2, pp. 413–420, 1999. bosis and Haemostasis, vol. 98, no. 2, pp. 278–282, 2007. [123] I. Staniszewska, S. Zaveri, L. D. Valle et al., “Interaction of [108] Y. Wallez and P. Huber, “Endothelial adherens and tight α9β1 integrin with thrombospondin-1 promotes angiogen- junctions in vascular homeostasis, inflammation and angio- esis,” Circulation Research, vol. 100, no. 9, pp. 1308–1316, genesis,” Biochimica et Biophysica Acta, vol. 1778, no. 3, pp. 794–809, 2008. [124] D. Bouvard, C. Brakebusch, E. Gustafsson et al., “Functional [109] A. Orpana, V. Ranta, T. Mikkola, L. Viinikka, and O. Yliko- consequences of integrin gene mutations in mice,” Circula- rkala, “Inducible nitric oxide and prostacyclin productions tion Research, vol. 89, no. 3, pp. 211–223, 2001. are differently controlled by extracellular matrix and cell [125] R. Fassler ¨ and M. Meyer, “Consequences of lack of β1 integrin density in human vascular endothelial cells,” Journal of gene expression in mice,” Genes and Development, vol. 9, no. Cellular Biochemistry, vol. 64, no. 4, pp. 538–546, 1997. 15, pp. 1896–1908, 1995. [110] M. A. Schwartz and D. W. DeSimone, “Cell adhesion [126] L. E. Stephens, A. E. Sutherland, I. V. Klimanskaya et al., receptors in mechanotransduction,” Current Opinion in Cell “Deletion of β1 integrins in mice results in inner cell mass Biology, vol. 20, no. 5, pp. 551–556, 2008. failure and peri-implantation lethality,” Genes and Develop- [111] J. Ivaska and J. Heino, “Interplay between cell adhesion and ment, vol. 9, no. 15, pp. 1883–1895, 1995. growth factor receptors: from the plasma membrane to the [127] T. R. Carlson, H. Hu, R. Braren, Y. H. Kim, and R. A. Wang, endosomes,” Cell and Tissue Research, vol. 339, no. 1, pp. 111– “Cell-autonomous requirement for β1 integrin in endothelial 120, 2010. cell adhesion, migration and survival during angiogenesis in [112] H. Hutchings, N. Ortega, and J. Plouet, ¨ “Extracellular mice,” Development, vol. 135, no. 12, pp. 2193–2202, 2008. [128] L. Lei, D. Liu, Y. Huang et al., “Endothelial expression of matrix-bound vascular endothelial growth factor promotes endothelial cell adhesion, migration, and survival through β1 integrin is required for embryonic vascular patterning integrin ligation,” The FASEB Journal, vol. 17, no. 11, pp. and postnatal vascular remodeling,” Molecular and Cellular 1520–1522, 2003. Biology, vol. 28, no. 2, pp. 794–802, 2008. 18 Journal of Oncology [129] H. Tanjore, E. M. Zeisberg, B. Gerami-Naini, and R. Kalluri, [146] J. P. Xiong, T. Stehle, B. Diefenbach et al., “Crystal structure “β1 integrin expression on endothelial cells is required of the extracellular segment of integrin αVβ3,” Science, vol. for angiogenesis but not for vasculogenesis,” Developmental 294, no. 5541, pp. 339–345, 2001. Dynamics, vol. 237, no. 1, pp. 75–82, 2008. [147] J. P. Xiong, T. Stehle, S. L. Goodman, and M. A. Arnaout, [130] A. C. Zovein, A. Luque, K. A. Turlo et al., “β1 integrin “New insights into the structural basis of integrin activation,” establishes endothelial cell polarity and arteriolar lumen Blood, vol. 102, no. 4, pp. 1155–1159, 2003. formation via a Par3-dependent mechanism,” Developmental [148] J.-P. Xiong, T. Stehle, R. Zhang et al., “Crystal structure of Cell, vol. 18, no. 1, pp. 39–51, 2010. the extracellular segment of integrin αVβ3incomplex with [131] A. Pozzi, P. E. Moberg, L. A. Miles, S. Wagner, P. Soloway, an Arg-Gly-Asp ligand,” Science, vol. 296, no. 5565, pp. 151– and H. A. Gardner, “Elevated matrix metalloprotease and 155, 2002. angiostatin levels in integrin α1 knockout mice cause reduced [149] M. J. Humphries, E. J. H. Symonds, and A. P. Mould, “Map- tumor vascularization,” Proceedings of the National Academy ping functional residues onto integrin crystal structures,” of Sciences of the United States of America, vol. 97, no. 5, pp. Current Opinion in Structural Biology, vol. 13, no. 2, pp. 236– 2202–2207, 2000. 243, 2003. [132] Z. Zhang, N. E. Ramirez, T. E. Yankeelov et al., “α2β1 integrin [150] A. P. Mould, E. J. Koper, A. Byron, G. Zahn, and M. J. expression in the tumor microenvironment enhances tumor Humphries, “Mapping the ligand-binding pocket of integrin angiogenesis in a tumor cell-specific manner,” Blood, vol. 111, α5β1 using a gain-of-function approach,” Biochemical Jour- no. 4, pp. 1980–1988, 2008. nal, vol. 424, no. 2, pp. 179–189, 2009. [133] J. T. Yang, H. Rayburn, and R. O. Hynes, “Embryonic meso- [151] D. A. Calderwood, “Integrin activation,” Journal of Cell dermal defects in α5 integrin-deficient mice,” Development, Science, vol. 117, no. 5, pp. 657–666, 2004. vol. 119, no. 4, pp. 1093–1105, 1993. [152] S. J. Shattil, C. Kim, and M. H. Ginsberg, “The final steps of [134] S. E. Francis, K. L. Goh, K. Hodivala-Dilke et al., “Central integrin activation: the end game,” Nature Reviews Molecular roles of α5β1 integrin and fibronectin in vascular develop- Cell Biology, vol. 11, no. 4, pp. 288–300, 2010. ment in mouse embryos and embryoid bodies,” Arteriosclero- [153] A. R. Ramjaun and K. Hodivala-Dilke, “The role of cell sis, Thrombosis, and Vascular Biology, vol. 22, no. 6, pp. 927– adhesion pathways in angiogenesis,” International Journal of 933, 2002. Biochemistry and Cell Biology, vol. 41, no. 3, pp. 521–530, [135] P. Parsons-Wingerter, I. M. Kasman, S. Norberg et al., “Uni- form overexpression and rapid accessibility of α5β1 integrin [154] K. L. Wegener, A. W. Partridge, J. Han et al., “Structural Basis on blood vessels in tumors,” American Journal of Pathology, of Integrin Activation by Talin,” Cell, vol. 128, no. 1, pp. 171– vol. 167, no. 1, pp. 193–211, 2005. 182, 2007. [136] E. Georges-Labouesse, N. Messaddeq, G. Yehia, L. Cadalbert, [155] M. Moser, B. Nieswandt, S. Ussar, M. Pozgajova, and R. A. Dierich, and M. Le Meur, “Absence of integrin α6leads Fassler ¨ , “Kindlin-3 is essential for integrin activation and to epidermolysis bullosa and neonatal death in mice,” Nature platelet aggregation,” Nature Medicine, vol. 14, no. 3, pp. 325– Genetics, vol. 13, no. 3, pp. 370–373, 1996. 330, 2008. [137] J. V. Welser, N. D. Lange, N. Flintoff-Dye, H. R. Burkin, and [156] J. Zhu, C. V. Carman, M. Kim, M. Shimaoka, T. A. Springer, D. J. Burkin, “Placental Defects in α7 Integrin Null Mice,” andB.H.Luo,“Requirementof α and β subunit transmem- Placenta, vol. 28, no. 11-12, pp. 1219–1228, 2007. brane helix separation for integrin outside-in signaling,” [138] R. Fassler, E. Georges-Labouesse, and E. Hirsch, “Genetic Blood, vol. 110, no. 7, pp. 2475–2483, 2007. analyses of integrin function in mice,” Current Opinion in [157] T. Xiao, J. Takagi, B. S. Coller, J. H. Wang, and T. A. Springer, Cell Biology, vol. 8, no. 5, pp. 641–646, 1996. “Structural basis for allostery in integrins and binding to [139] X. Z. Huang, J. F. Wu, R. Ferrando et al., “Fatal bilateral fibrinogen-mimetic therapeutics,” Nature, vol. 432, no. 7013, chylothorax in mice lacking the integrin α9β1,” Molecular pp. 59–67, 2004. and Cellular Biology, vol. 20, no. 14, pp. 5208–5215, 2000. [158] N. Nishida, C. Xie, M. Shimaoka, Y. Cheng, T. Walz, and T. A. [140] K. M. Hodivala-Dilke, K. P. McHugh, D. A. Tsakiris et al., Springer, “Activation of leukocyte β2 integrins by conversion “β3-integrin-deficient mice are a model for Glanzmann from bent to extended conformations,” Immunity, vol. 25, no. thrombasthenia showing placental defects and reduced sur- 4, pp. 583–594, 2006. vival,” Journal of Clinical Investigation, vol. 103, no. 2, pp. [159] K. R. Legate, E. Montanez, ˜ O. Kudlacek, and R. Fassler ¨ , “ILK, 229–238, 1999. PINCH and parvin: the tIPP of integrin signalling,” Nature [141] Y. Takada, X. Ye, and S. Simon, “The integrins,” Genome Reviews Molecular Cell Biology, vol. 7, no. 1, pp. 20–31, 2006. Biology, vol. 8, no. 5, article 215, pp. 211–219, 2007. [160] M. Zoller ¨ , “Tetraspanins: push and pull in suppressing and [142] M. H. Ginsberg, A. Partridge, and S. J. Shattil, “Integrin promoting metastasis,” Nature Reviews Cancer, vol. 9, no. 1, regulation,” Current Opinion in Cell Biology, vol. 17, no. 5, pp. 40–55, 2009. pp. 509–516, 2005. [161] J. H. Park, J. M. Ryu, and H. J. Han, “Involvement of [143] M. A. Arnaout, S. L. Goodman, and J. P. Xiong, “Structure caveolin-1 in fibronectin-induced mouse embryonic stem and mechanics of integrin-based cell adhesion,” Current cell proliferation: role of FAK, RhoA, PI3K/Akt, and ERK 1/2 Opinion in Cell Biology, vol. 19, no. 5, pp. 495–507, 2007. pathways,” Journal of Cellular Physiology, vol. 226, no. 1, pp. [144] B. H. Luo, C. V. Carman, and T. A. Springer, “Structural 267–275, 2011. basis of integrin regulation and signaling,” Annual Review of [162] S. H. Lee, Y. J. Lee, S. W. Park, H. S. Kim, and H. J. Immunology, vol. 25, pp. 619–647, 2007. Han, “Caveolin-1 and integrin β1 regulate embryonic stem [145] J. Takagi, K. Strokovich, T. A. Springer, and T. Walz, cell proliferation via p38 MAPK and FAK in high glucose,” “Structure of integrin α5β1 in complex with fibronectin,” Journal of Cellular Physiology, vol. 226, no. 7, pp. 1850–1859, EMBO Journal, vol. 22, no. 18, pp. 4607–4615, 2003. 2011. Journal of Oncology 19 [163] A. Byron, M. R. Morgan, and M. J. Humphries, “Adhesion [178] B. P. Eliceiri, X. S. Puente, J. D. Hood et al., “Src-mediated signalling complexes,” Current Biology, vol. 20, no. 24, pp. coupling of focal adhesion kinase to integrin αvβ5invascular R1063–R1067, 2010. endothelial growth factor signaling,” Journal of Cell Biology, vol. 157, no. 1, pp. 149–160, 2002. [164] M. A. Del Pozo, N. B. Alderson, W. B. Kiosses, H. H. Chiang, R. G. W. Anderson, and M. A. Schwartz, “Integrins regulate [179] S. A. Wickstrom, ¨ K. Alitalo, and J. Keski-Oja, “Endostatin rac targeting by internalization of membrane domains,” associates with integrin α5β1 and caveolin-1, and activates Science, vol. 303, no. 5659, pp. 839–842, 2004. Src via a tyrosyl phosphatase-dependent pathway in human endothelial cells,” Cancer Research, vol. 62, no. 19, pp. 5580– [165] I. J. Salanueva, A. Cerezo, M. C. Guadamillas, and M. A. 5589, 2002. Del Pozo, “Integrin regulation of caveolin function: caveolae review series,” Journal of Cellular and Molecular Medicine, vol. [180] A. Aiyer and J. Varner, “The role of integrins in tumor 11, no. 5, pp. 969–980, 2007. angiogenesis,” in Cancer Drug Discovery Development—Anti- angiogenic Agents in Cancer Therapy,B.A.Teicher andL.M. [166] I. Bethani, S. S. Skanland, ˚ I. Dikic, and A. Acker-Palmer, “Spatial organization of transmembrane receptor signalling,” Ellis, Eds., pp. 49–73, Humana Press, Totowa, NJ, USA, 2008. EMBO Journal, vol. 29, no. 16, pp. 2677–2688, 2010. [181] C. Chandra Kumar, “Signaling by integrin receptors,” Onco- gene, vol. 17, no. 11, pp. 1365–1373, 1998. [167] R. W. Tilghman and J. T. Parsons, “Focal adhesion kinase as a regulator of cell tension in the progression of cancer,” [182] K. I. Nagashima, A. Endo, H. Ogita et al., “Adaptor protein Seminars in Cancer Biology, vol. 18, no. 1, pp. 45–52, 2008. Crk is required for ephrin-B1-induced membrane ruffling and focal complex assembly of human aortic endothelial [168] M. C. Brown, L. A. Cary, J. S. Jamieson, J. A. Cooper, cells,” Molecular Biology of the Cell, vol. 13, no. 12, pp. 4231– and C. E. Turner, “Src and FAK kinases cooperate to phos- 4242, 2002. phorylate paxillin kinase linker, stimulate its focal adhesion localization, and regulate cell spreading and protrusiveness,” [183] F. Paulhe, C. Racaud-Sultan, A. Ragab et al., “Differential Molecular Biology of the Cell, vol. 16, no. 9, pp. 4316–4328, regulation of phosphoinositide metabolism by α vβ3and 2005. αvβ5 integrins upon smooth muscle cell migration,” Journal of Biological Chemistry, vol. 276, no. 45, pp. 41832–41840, [169] E. G. Arias-Salgado, S. Lizano, S. Sarkar, J. S. Brugge, M. H. Ginsberg, and S. J. Shattil, “Src kinase activation by direct interaction with the integrin β cytoplasmic domain,” [184] V. Carloni, R. G. Romanelli, M. Pinzani, G. Laffi,and P. Proceedings of the National Academy of Sciences of the United Gentilini, “Focal adhesion kinase and phospholipase Cγ States of America, vol. 100, no. 23, pp. 13298–13302, 2003. involvement in adhesion and migration of human hepatic stellate cells,” Gastroenterology, vol. 112, no. 2, pp. 522–531, [170] S. K. Hanks, M. B. Calalb, M. C. Harper, and S. K. Patel, “Focal adhesion protein-tyrosine kinase phosphorylated in 1997. response to cell attachment to fibronectin,” Proceedings of the [185] X. Zhang, A. Chattopadhyay, Q. S. Ji et al., “Focal adhesion National Academy of Sciences of the United States of America, kinase promotes phospholipase C-γ1 activity,” Proceedings vol. 89, no. 18, pp. 8487–8491, 1992. of the National Academy of Sciences of the United States of America, vol. 96, no. 16, pp. 9021–9026, 1999. [171] A. Papapetropoulos, D. Fulton, K. Mahboubi et al., “Angi- opoietin-1 inhibits endothelial cell apoptosis via the Akt/ [186] L. Bi, I. Okabe, D. J. Bernard, A. Wynshaw-Boris, and R. survivin pathway,” Journal of Biological Chemistry, vol. 275, L. Nussbaum, “Proliferative defect and embryonic lethality no. 13, pp. 9102–9105, 2000. in mice homozygous for a deletion in the p110α subunit of phosphoinositide 3-kinase,” Journal of Biological Chemistry, [172] S. H. Kim and S. H. Kim, “Antagonistic effect of EGF vol. 274, no. 16, pp. 10963–10968, 1999. on FAK phosphorylation/dephosphorylation in a cell,” Cell Biochemistry and Function, vol. 26, no. 5, pp. 539–547, 2008. [187] E. Lelievre, P. M. Bourbon, L. J. Duan, R. L. Nussbaum, and G. H. Fong, “Deficiency in the p110α subunit of PI3K results [173] J. K. Slack-Davis, S. T. Eblen, M. Zecevic et al., “PAK1 in diminished Tie2 expression and Tie2-/–like vascular phosphorylation of MEK1 regulates fibronectin-stimulated defects in mice,” Blood, vol. 105, no. 10, pp. 3935–3938, 2005. MAPK activation,” Journal of Cell Biology, vol. 162, no. 2, pp. 281–291, 2003. [188] M. Graupera, J. Guillermet-Guibert, L. C. Foukas et al., [174] M. L. Edin and R. L. Juliano, “Raf-1 serine 338 phospho- “Angiogenesis selectively requires the p110α isoform of PI3K to control endothelial cell migration,” Nature, vol. 453, no. rylation plays a key role in adhesion-dependent activation 7195, pp. 662–666, 2008. of extracellular signal-regulated kinase by epidermal growth factor,” Molecular and Cellular Biology, vol. 25, no. 11, pp. [189] A. Sudhakar, H. Sugimoto, C. Yang, J. Lively, M. Zeisberg, and 4466–4475, 2005. R. Kalluri, “Human tumstatin and human endostatin exhibit [175] T. L. Shen, A. Y. J. Park, A. Alcaraz et al., “Conditional distinct antiangiogenic activities mediated by αvβ and α5β1 integrins,” Proceedings of the National Academy of Sciences of knockout of focal adhesion kinase in endothelial cells reveals its role in angiogenesis and vascular development in late the United States of America, vol. 100, no. 8, pp. 4766–4771, embryogenesis,” Journal of Cell Biology, vol. 169, no. 6, pp. 941–952, 2005. [190] S. Gupta, A. R. Ramjaun, P. Haiko et al., “Binding of ras to [176] D. L. Courter, L. Lomas, M. Scatena, and C. M. Giachelli, “Src phosphoinositide 3-kinase p110α is required for ras-driven tumorigenesis in mice,” Cell, vol. 129, no. 5, pp. 957–968, kinase activity is required for integrin αvβ 3-mediated acti- vation of nuclear factor-κB,” Journal of Biological Chemistry, 2007. vol. 280, no. 13, pp. 12145–12151, 2005. [191] M. S. Roberts, A. J. Woods, P. E. Shaw, and J. C. Norman, “ERK1 associates with αvβ3 integrin and regulates cell [177] J. Zaric and C. Ruegg ¨ , “Integrin-mediated adhesion and spreading on vitronectin,” Journal of Biological Chemistry, soluble ligand binding stabilize COX-2 protein levels in vol. 278, no. 3, pp. 1975–1985, 2003. endothelial cells by inducing expression and preventing degradation,” Journal of Biological Chemistry, vol. 280, no. 2, [192] S. M. Short, G. A. Talbott, and R. L. Juliano, “Integrin- pp. 1077–1085, 2005. mediated signaling events in human endothelial cells,” 20 Journal of Oncology Molecular Biology of the Cell, vol. 9, no. 8, pp. 1969–1980, [208] V. Samanna, H. Wei, D. Ego-Osuala, and M. A. Chellaiah, 1998. “Alpha-V-dependent outside-in signaling is required for the regulation of CD44 surface expression, MMP-2 secretion, [193] M. Huser ¨ , J. Luckett, A. Chiloeches et al., “MEK kinase and cell migration by osteopontin in human melanoma activity is not necessary for Raf-1 function,” EMBO Journal, cells,” Experimental Cell Research, vol. 312, no. 12, pp. 2214– vol. 20, no. 8, pp. 1940–1951, 2001. 2230, 2006. [194] S. Giroux, M. Tremblay, D. Bernard et al., “Embryonic [209] M. M. Zutter, S. A. Santoro, W. D. Staatz, and Y. L. Tsung, death of Mek1-deficient mice reveals a role for this kinase “Re-expression of the α2β1 integrin abrogates the malignant in angiogenesis in the labyrinthine region of the placenta,” phenotype of breast carcinoma cells,” Proceedings of the Current Biology, vol. 9, no. 7, pp. 369–372, 1999. National Academy of Sciences of the United States of America, [195] J. D. Hood, R. Frausto, W. B. Kiosses, M. A. Schwartz, and vol. 92, no. 16, pp. 7411–7415, 1995. D. A. Cheresh, “Differential αv integrin-mediated Ras-ERK [210] A. Kren, V. Baeriswyl, F. Lehembre et al., “Increased tumor signaling during two pathways of angiogenesis,” Journal of cell dissemination and cellular senescence in the absence of Cell Biology, vol. 162, no. 5, pp. 933–943, 2003. β1-integrin function,” EMBO Journal, vol. 26, no. 12, pp. [196] K. K. Wary, F. Mainiero, S. J. Isakoff,E.E.Marcantonio,and 2832–2842, 2007. F. G. Giancotti, “The adaptor protein Shc couples a class of [211] H. Zhao, F. P. Ross, and S. L. Teitelbaum, “Unoccupied integrins to the control of cell cycle progression,” Cell, vol. 87, αvβ3 integrin regulates osteoclast apoptosis by transmitting a no. 4, pp. 733–743, 1996. positive death signal,” Molecular Endocrinology, vol. 19, no. 3, [197] A. Pozzi, K. K. Wary, F. G. Giancotti, and H. A. Gardner, pp. 771–780, 2005. “Integrin α1β1 mediates a unique collagen-dependent pro- [212] S. M. Frisch and R. A. Screaton, “Anoikis mechanisms,” liferation pathway in vivo,” Journal of Cell Biology, vol. 142, Current Opinion in Cell Biology, vol. 13, no. 5, pp. 555–562, no. 2, pp. 587–594, 1998. [198] A. K. Fournier, L. E. Campbell, P. Castagnino et al., “Rac- [213] D. G. Stupack, T. Teitz, M. D. Potter et al., “Potentiation of dependent cyclin D1 gene expression regulated by cadherin- neuroblastoma metastasis by loss of caspase-8,” Nature, vol. and integrin-mediated adhesion,” Journal of Cell Science, vol. 439, no. 7072, pp. 95–99, 2006. 121, no. 2, pp. 226–233, 2008. [214] J. S. Desgrosellier, L. A. Barnes, D. J. Shields et al., “An [199] E. A. Klein, L. Yin, D. Kothapalli et al., “Cell-cycle control by integrin α(v)β(3)-c-Src oncogenic unit promotes anchorage- physiological matrix elasticity and in vivo tissue stiffening,” independence and tumor progression,” Nature Medicine, vol. Current Biology, vol. 19, no. 18, pp. 1511–1518, 2009. 15, no. 10, pp. 1163–1169, 2009. [200] S. Klein, A. R. De Fougerolles, P. Blaikie et al., “α5β1 integrin [215] M. L. Matter and E. Ruoslahti, “A signaling pathway from the activates an NF-κB-dependent program of gene expression α5β1and αvβ3 integrins that elevates bcl-2 transcription,” important for angiogenesis and inflammation,” Molecular Journal of Biological Chemistry, vol. 276, no. 30, pp. 27757– and Cellular Biology, vol. 22, no. 16, pp. 5912–5922, 2002. 27763, 2001. [201] M. Reidy, P. Zihlmann, J. A. Hubbell, and H. Hall, “Acti- [216] F. Aoudjit and K. Vuori, “Matrix attachment regulates Fas- vation of cell-survival transcription factor NFκBinL1Ig6- induced apoptosis in endothelial cells: a role for c-Flip and stimulated endothelial cells,” Journal of Biomedical Materials implications for anoikis,” Journal of Cell Biology, vol. 153, no. Research Part A, vol. 77, no. 3, pp. 542–550, 2006. 3, pp. 633–643, 2001. [202] M. Scatena, M. Almeida, M. L. Chaisson, N. Fausto, R. F. [217] W. Bao and S. Stromblad, ¨ “Integrin αv-mediated inactivation Nicosia, and C. M. Giachelli, “NF-κB mediates αvβ3 integrin- of p53 controls a MEK1-dependent melanoma cell survival induced endothelial cell survival,” Journal of Cell Biology, vol. pathway in three-dimensional collagen,” Journal of Cell 141, no. 4, pp. 1083–1093, 1998. Biology, vol. 167, no. 4, pp. 745–756, 2004. [203] O. Dormond, M. Bezzi, A. Mariotti, and C. Ruegg ¨ , “Pros- [218] A. Alavi, J. D. Hood, R. Frausto, D. G. Stupack, and D. A. taglandin E2 promotes integrin αvβ3-dependent endothelial Cheresh, “Role of Raf in vascular protection from distinct cell adhesion, Rac-activation, and spreading through cAMP/ apoptotic stimuli,” Science, vol. 301, no. 5629, pp. 94–96, PKA-dependent signaling,” Journal of Biological Chemistry, vol. 277, no. 48, pp. 45838–45846, 2002. [219] E. Ruoslahti, “Specialization of tumour vasculature,” Nature [204] C. S. Boosani, A. P. Mannam, D. Cosgrove et al., “Regulation Reviews Cancer, vol. 2, no. 2, pp. 83–90, 2002. of COX-2-mediated signaling by α3typeIVnoncollagenous [220] N. Alam, H. L. Goel, M. J. Zarif et al., “The integrin—growth domain in tumor angiogenesis,” Blood, vol. 110, no. 4, pp. factor receptor duet,” Journal of Cellular Physiology, vol. 213, 1168–1177, 2007. no. 3, pp. 649–653, 2007. [205] T. Kisseleva, L. Song, M. Vorontchikhina, N. Feirt, J. Kitajew- [221] G. Serini, L. Napione, M. Arese, and F. Bussolino, “Besides ski, and C. Schindler, “NF-κB regulation of endothelial cell adhesion: new perspectives of integrin functions in angio- function during LPS-induced toxemia and cancer,” Journal of genesis,” Cardiovascular Research, vol. 78, no. 2, pp. 213–222, Clinical Investigation, vol. 116, no. 11, pp. 2955–2963, 2006. [206] H. Lahlou, V. Sanguin-Gendreau, D. Zuo et al., “Mammary [222] E. S. Wijelath, S. Rahman, M. Namekata et al., “Heparin-II epithelial-specific disruption of the focal adhesion kinase domain of fibronectin is a vascular endothelial growth factor- blocks mammary tumor progression,” Proceedings of the binding domain: enhancement of VEGF biological activity National Academy of Sciences of the United States of America, by a singular growth factor/matrix protein synergism,” vol. 104, no. 51, pp. 20302–20307, 2007. Circulation Research, vol. 99, no. 8, pp. 853–860, 2006. [207] Y. Pylayeva, K. M. Gillen, W. Gerald, H. E. Beggs, L. F. [223] C. Q. Zhu, S. N. Popova, E. R. S. Brown et al., “Integrin α11 regulates IGF2 expression in fibroblasts to enhance Reichardt, and F. G. Giancotti, “Ras- and PI3K-dependent breast tumorigenesis in mice and humans requires focal tumorigenicity of human non-small-cell lung cancer cells,” Proceedings of the National Academy of Sciences of the United adhesion kinase signaling,” Journal of Clinical Investigation, vol. 119, no. 2, pp. 252–266, 2009. States of America, vol. 104, no. 28, pp. 11754–11759, 2007. Journal of Oncology 21 [224] A. Orecchia, P. M. Lacal, C. Schietroma, V. Morea, G. [241] A. C. Aplin, E. Fogel, and R. F. Nicosia, “MCP-1 promotes Zambruno, and C. M. Failla, “Vascular endothelial growth mural cell recruitment during angiogenesis in the aortic ring factor receptor-1 is deposited in the extracellular matrix by model,” Angiogenesis, vol. 13, no. 3, pp. 219–226, 2010. endothelial cells and is a ligand for the α5β1 integrin,” Journal [242] E. Iivanainen, V. M. Kah ¨ ar ¨ i, J. Heino, and K. Elenius, of Cell Science, vol. 116, no. 17, pp. 3479–3489, 2003. “Endothelial cell-matrix interactions,” Microscopy Research and Technique, vol. 60, no. 1, pp. 13–22, 2003. [225] K. Kajiya, S. Hirakawa, B. Ma, I. Drinnenberg, and M. [243] J. E. Rundhaug, “Matrix metalloproteinases and angiogene- Detmar, “Hepatocyte growth factor promotes lymphatic vessel formation and function,” EMBO Journal, vol. 24, no. sis,” Journal of Cellular and Molecular Medicine, vol. 9, no. 2, pp. 267–285, 2005. 16, pp. 2885–2895, 2005. [244] G. Murphy and H. Nagase, “Localizing matrix metallo- [226] M. Murakami, A. Elfenbein, and M. Simons, “Non-canonical proteinase activities in the pericellular environment,” FEBS fibroblast growth factor signalling in angiogenesis,” Cardio- Journal, vol. 278, no. 1, pp. 2–15, 2011. vascular Research, vol. 78, no. 2, pp. 223–231, 2008. [245] G. O. Ahn and J. M. Brown, “Matrix metalloproteinase-9 is [227] G. Camenisch, M. T. Pisabarro, D. Sherman et al., “ANGPTL3 required for tumor vasculogenesis but not for angiogenesis: stimulates endothelial cell adhesion and migration via inte- role of bone marrow-derived myelomonocytic cells,” Cancer grin αvβ3 and induces blood vessel formation in vivo,” Cell, vol. 13, no. 3, pp. 193–205, 2008. Journal of Biological Chemistry, vol. 277, no. 19, pp. 17281– [246] H. M. Eilken and R. H. Adams, “Dynamics of endothelial cell 17290, 2002. behavior in sprouting angiogenesis,” Current Opinion in Cell [228] S. M. Dallabrida, N. Ismail, J. R. Oberle, B. E. Himes, and M. Biology, vol. 22, no. 5, pp. 617–625, 2010. A. Rupnick, “Angiopoietin-1 promotes cardiac and skeletal [247] W. Risau, “Mechanisms of angiogenesis,” Nature, vol. 386, myocyte survival through integrins,” Circulation research, vol. no. 6626, pp. 671–674, 1997. 96, no. 4, pp. e8–e24, 2005. [248] J. Folkman, “Looking for a good endothelial address,” Cancer [229] E. C. Finger and A. J. Giaccia, “Hypoxia, inflammation, and Cell, vol. 1, no. 2, pp. 113–115, 2002. the tumor microenvironment in metastatic disease,” Cancer [249] T. Asahara, T. Murohara, A. Sullivan et al., “Isolation and Metastasis Reviews, vol. 29, no. 2, pp. 285–293, 2010. of putative progenitor endothelial cells for angiogenesis,” [230] A. Billioux, U. Modlich, and R. Bicknell, “Angiogenesis,” in Science, vol. 275, no. 5302, pp. 964–967, 1997. The Cancer Handbook, M. Alison, Ed., vol. 1, pp. 144–154, [250] F. Brellier, R. P. Tucker, and R. Chiquet-Ehrismann, John Wiley & Sons, 2007. “Tenascins and their implications in diseases and tissue [231] P. Carmeliet, “Angiogenesis in health and disease,” Nature mechanics,” Scandinavian Journal of Medicine and Science in Medicine, vol. 9, no. 6, pp. 653–660, 2003. Sports, vol. 19, no. 4, pp. 511–519, 2009. [232] J. Folkman, K. Watson, D. Ingber, and D. Hanahan, “Induc- [251] M. Kaspar, L. Zardi, and D. Neri, “Fibronectin as target for tion of angiogenesis during the transition from hyperplasia tumor therapy,” International Journal of Cancer, vol. 118, no. to neoplasia,” Nature, vol. 339, no. 6219, pp. 58–61, 1989. 6, pp. 1331–1339, 2006. [233] N. Weidner, J. P. Semple, W. R. Welch, and J. Folkman, [252] M. Midulla, R. Verma, M. Pignatelli, M. A. Ritter, N. S. “Tumor angiogenesis and metastasis—correlation in invasive Courtenay-Luck, and A. J. T. George, “Source of oncofetal breast carcinoma,” New England Journal of Medicine, vol. 324, ED-B-containing fibronectin: implications of production of no. 1, pp. 1–8, 1991. both tumor and endothelial cells,” Cancer Research, vol. 60, [234] J. Kandel, E. Bossy-Wetzel, F. Radvanyi, M. Klagsbrun, J. no. 1, pp. 164–169, 2000. Folkman, and D. Hanahan, “Neovascularization is associated [253] K. S. Midwood and G. Orend, “The role of tenascin-C in tis- with a switch to the export of bFGF in the multistep sue injury and tumorigenesis,” Journal of Cell Communication development of fibrosarcoma,” Cell, vol. 66, no. 6, pp. 1095– and Signaling, vol. 3, no. 3-4, pp. 287–310, 2009. 1104, 1991. [254] M. Degen, F. Brellier, S. Schenk et al., “Tenascin-W, a new [235] E. Y. Lin and J. W. Pollard, “Tumor-associated macrophages marker of cancer stroma, is elevated in sera of colon and press the angiogenic switch in breast cancer,” Cancer breast cancer patients,” International Journal of Cancer, vol. Research, vol. 67, no. 11, pp. 5064–5066, 2007. 122, no. 11, pp. 2454–2461, 2008. [236] M. C. Schmid and J. A. Varner, “Myeloid cell trafficking and [255] E. Martina, R. Chiquet-Ehrismann, and F. Brellier, tumor angiogenesis,” Cancer Letters, vol. 250, no. 1, pp. 1–8, “Tenascin-W: an extracellular matrix protein associated with osteogenesis and cancer,” International Journal of Biochem- [237] N. Ferrara, H. P. Gerber, and J. LeCouter, “The biology of istry and Cell Biology, vol. 42, no. 9, pp. 1412–1415, 2010. VEGF and its receptors,” Nature Medicine,vol. 9, no.6,pp. [256] B. Dome, ¨ M. J. C. Hendrix, S. Paku, J. Tov ´ ar ´ i, and J. 669–676, 2003. T´ımar ´ , “Alternative vascularization mechanisms in cancer: [238] E. C. Keeley, B. Mehrad, and R. M. Strieter, “Chemokines pathology and therapeutic implications,” American Journal of as mediators of tumor angiogenesis and neovascularization,” Pathology, vol. 170, no. 1, pp. 1–15, 2007. Experimental Cell Research, vol. 317, no. 5, pp. 685–690, 2011. [257] F. Hillen and A. W. Griffioen, “Tumour vascularization: [239] K. H. Hong, J. Ryu, and K. H. Han, “Monocyte chemoattrac- sprouting angiogenesis and beyond,” Cancer and Metastasis tant protein-1-induced angiogenesis is mediated by vascular Reviews, vol. 26, no. 3-4, pp. 489–502, 2007. endothelial growth factor-A,” Blood, vol. 105, no. 4, pp. 1405– [258] D. Lyden, K. Hattori, S. Dias et al., “Impaired recruitment 1407, 2005. of bone-marrow-derived endothelial and hematopoietic pre- cursor cells blocks tumor angiogenesis and growth,” Nature [240] J. Niu, A. Azfer, O. Zhelyabovska, S. Fatma, and P. E. Kolat- Medicine, vol. 7, no. 11, pp. 1194–1201, 2001. tukudy, “Monocyte chemotactic protein (MCP)-1 promotes angiogenesis via a novel transcription factor, MCP-1-induced [259] D. Ribatti, “The involvement of endothelial progenitor cells in tumor angiogenesis,” Journal of Cellular and Molecular protein (MCPIP),” Journal of Biological Chemistry, vol. 283, no. 21, pp. 14542–14551, 2008. Medicine, vol. 8, no. 3, pp. 294–300, 2004. 22 Journal of Oncology [260] M. Reyes, A. Dudek, B. Jahagirdar, L. Koodie, P. H. Marker, withdrawal,” Journal of Clinical Investigation, vol. 103, no. 2, and C. M. Verfaillie, “Origin of endothelial progenitors pp. 159–165, 1999. in human postnatal bone marrow,” Journal of Clinical [276] A. J. Maniotis, R. Folberg, A. Hess et al., “Vascular channel Investigation, vol. 109, no. 3, pp. 337–346, 2002. formation by human melanoma cells in vivo and in vitro: [261] M. L. Iruela-Arispe and G. E. Davis, “Cellular and molecular vasculogenic mimicry,” American Journal of Pathology, vol. mechanisms of vascular lumen formation,” Developmental 155, no. 3, pp. 739–752, 1999. Cell, vol. 16, no. 2, pp. 222–231, 2009. [277] R. Folberg and A. J. Maniotis, “Vasculogenic mimicry,” Acta [262] B. Strilic, ´ T. Kucer ˇ a, J. Eglinger et al., “The molecular basis Pathologica, Microbiologica. et Immunologica Scandinavica, of vascular lumen formation in the developing mouse aorta,” vol. 112, no. 7-8, pp. 508–525, 2004. Developmental Cell, vol. 17, no. 4, pp. 505–515, 2009. [278] A. J. G. Potgens, ¨ M. C. Van Altena, N. H. Lubsen, D. J. Ruiter, [263] C. M. Ghajar, S. C. George, and A. J. Putnam, “Matrix met- and R. M. W. De Waal, “Analysis of the tumor vasculature and alloproteinase control of capillary morphogenesis,” Critical metastatic behavior of xenografts of human melanoma cell Reviews in Eukaryotic Gene Expression, vol. 18, no. 3, pp. 251– lines transfected with vascular permeability factor,” American 278, 2008. Journal of Pathology, vol. 148, no. 4, pp. 1203–1217, 1996. [264] R. Hildenbrand, H. Allgayer, A. Marx, and P. Stroebel, [279] R. Clarijs, I. Otte-Holler ¨ , D. J. Ruiter, and R. M. W. De Waal, “Modulators of the urokinase-type plasminogen activation “Presence of a fluid-conducting meshwork in xenografted system for cancer,” Expert Opinion on Investigational Drugs, cutaneous and primary human uveal melanoma,” Investiga- vol. 19, no. 5, pp. 641–652, 2010. tive Ophthalmology and Visual Science, vol. 43, no. 4, pp. 912– 918, 2002. [265] F. Bougatef, C. Quemener, S. Kellouche et al., “EMMPRIN promotes angiogenesis through hypoxia-inducible factor-2α- [280] T. Kucer ˇ a and E. Lammert, “Ancestral vascular tube forma- mediated regulation of soluble VEGF isoforms and their tion and its adoption by tumors,” Biological Chemistry, vol. receptor VEGFR-2,” Blood, vol. 114, no. 27, pp. 5547–5556, 390, no. 10, pp. 985–994, 2009. 2009. [281] W. Ruf, E. A. Seftor, R. J. Petrovan et al., “Differential role [266] P. C. Brooks, S. Stromblad, ¨ L. C. Sanders et al., “Localization of tissue factor pathway inhibitors 1 and 2 in melanoma of matrix metalloproteinase MMP-2 to the surface of invasive vasculogenic mimicry,” Cancer Research, vol. 63, no. 17, pp. cells by interaction with integrin αvβ3,” Cell, vol. 85, no. 5, 5381–5389, 2003. pp. 683–693, 1996. [282] S. Anand, B. K. Majeti, L. M. Acevedo et al., “MicroRNA- [267] V. Djonov, M. Schmid, S. A. Tschanz, and P. H. Burri, 132-mediated loss of p120RasGAP activates the endothelium “Intussusceptive angiogenesis. Its role in embryonic vascular to facilitate pathological angiogenesis,” Nature Medicine, vol. network formation,” Circulation Research, vol. 86, no. 3, pp. 16, no. 8, pp. 909–914, 2010. 286–292, 2000. [283] S. Anand and D. A. Cheresh, “MicroRNA-mediated regula- [268] H. Kurz, P. H. Burri, and V. G. Djonov, “Angiogenesis tion of the angiogenic switch,” Current Opinion in Hematol- and vascular remodeling by intussusception: from form to ogy, vol. 18, no. 3, pp. 171–176, 2011. function,” News in Physiological Sciences,vol. 18, no.2,pp. [284] G. Bellon, L. Martiny, and A. Robinet, “Matrix metallopro- 65–70, 2003. teinases and matrikines in angiogenesis,” Critical Reviews in [269] A. N. Makanya, R. Hlushchuk, and V. G. Djonov, “Intussus- Oncology/Hematology, vol. 49, no. 3, pp. 203–220, 2004. ceptive angiogenesis and its role in vascular morphogenesis, [285] P. Nyberg, L. Xie, and R. Kalluri, “Endogenous inhibitors patterning, and remodeling,” Angiogenesis,vol. 12, no.2,pp. of angiogenesis,” Cancer Research, vol. 65, no. 10, pp. 3967– 113–123, 2009. 3979, 2005. [270] D. J. Brat and E. G. Van Meir, “Glomeruloid microvascular [286] M. Shimaoka and T. A. Springer, “Therapeutic antagonists proliferation orchestrated by VPF/VEGF: a new world of and conformational regulation of integrin function,” Nature angiogenesis research,” American Journal of Pathology, vol. Reviews Drug Discovery, vol. 2, no. 9, pp. 703–716, 2003. 158, no. 3, pp. 789–796, 2001. [287] J. A. Eble and J. Haier, “Integrins in cancer treatment,” [271] O. Straume, P. O. Chappuis, H. B. Salvesen et al., “Prognostic Current Cancer Drug Targets, vol. 6, no. 2, pp. 89–105, 2006. importance of glomeruloid microvascular proliferation indi- [288] S. M. Short, A. Derrien, R. P. Narsimhan, J. Lawler, D. cates an aggressive angiogenic phenotype in human cancers,” E. Ingber, and B. R. Zetter, “Inhibition of endothelial cell Cancer Research, vol. 62, no. 23, pp. 6808–6811, 2002. migration by thrombospondin-1 type-1 repeats is mediated [272] J. Holash, P. C. Maisonpierre, D. Compton et al., “Vessel by β1 integrins,” Journal of Cell Biology, vol. 168, no. 4, pp. cooption, regression, and growth in tumors mediated by 643–653, 2005. angiopoietins and VEGF,” Science, vol. 284, no. 5422, pp. [289] X. Zhang and J. Lawler, “Thrombospondin-based antiangio- 1994–1998, 1999. genic therapy,” Microvascular Research,vol. 74, no.2-3,pp. [273] B. Dome, ¨ S. Paku, B. Somlai, and J. Timar, “Vascularization 90–99, 2007. of cutaneous melanoma involves vessel co-option and has [290] P. C. Colorado, A. Torre, G. Kamphaus et al., “Anti- clinical significance,” Journal of Pathology, vol. 197, no. 3, pp. angiogenic cues from vascular basement membrane colla- 355–362, 2002. gen,” Cancer Research, vol. 60, no. 9, pp. 2520–2526, 2000. [274] M. Scharpfenecker, U. Fiedler, Y. Reiss, and H. G. Augustin, [291] P. Nyberg, L. Xie, H. Sugimoto et al., “Characterization of “The Tie-2 ligand angiopoietin-2 destabilizes quiescent the anti-angiogenic properties of arresten, an α1β1 integrin- endothelium through an internal autocrine loop mecha- dependent collagen-derived tumor suppressor,” Experimental nism,” Journal of Cell Science, vol. 118, no. 4, pp. 771–780, Cell Research, vol. 314, no. 18, pp. 3292–3305, 2008. [292] B. P. Woodall, A. Nystro ¨m,R.A.Iozzo et al., “Integrin α2β1 [275] L. E. Benjamin, D. Golijanin, A. Itin, D. Pode, and E. Keshet, is the required receptor for endorepellin angiostatic activity,” “Selective ablation of immature blood vessels in established Journal of Biological Chemistry, vol. 283, no. 4, pp. 2335– human tumors follows vascular endothelial growth factor 2343, 2008. Journal of Oncology 23 [293] M. Mongiat, S. M. Sweeney, J. D. San Antonio, J. Fu, and [308] S. A. Wickstrom, ¨ K. Alitalo, and J. Keski-Oja, “An endostatin- R. V. Iozzo, “Endorepellin, a novel inhibitor of angiogenesis derived peptide interacts with integrins and regulates actin derived from the C terminus of perlecan,” Journal of Biologi- cytoskeleton and migration of endothelial cells,” Journal of cal Chemistry, vol. 278, no. 6, pp. 4238–4249, 2003. Biological Chemistry, vol. 279, no. 19, pp. 20178–20185, 2004. [294] C. Marcinkiewicz, P. H. Weinreb, J. J. Calvete et al., “Obtu- [309] M. E. Cianfrocca, K. A. Kimmel, J. Gallo et al., “Phase 1 trial statin: a potent selective inhibitor of α1β1 integrin in vitro of the antiangiogenic peptide ATN-161 (Ac-PHSCN-NH 2), and angiogenesis in vivo,” Cancer Research, vol. 63, no. 9, pp. a beta integrin antagonist, in patients with solid tumours,” 2020–2023, 2003. British Journal of Cancer, vol. 94, no. 11, pp. 1621–1626, 2006. [295] M. C. Brown, I. Staniszewska, L. Del Valle, G. P. Tuszynski, [310] A. P. Mould, L. Burrows, and M. J. Humphries, “Iden- and C. Marcinkiewicz, “Angiostatic activity of obtustatin as tification of amino acid residues that form part of the α1β1 integrin inhibitor in experimental melanoma growth,” ligand- binding pocket of integrin α5β1,” Journal of Biological International Journal of Cancer, vol. 123, no. 9, pp. 2195– Chemistry, vol. 273, no. 40, pp. 25664–25672, 1998. 2203, 2008. [311] L. Marinelli, A. Meyer, D. Heckmann, A. Lavecchia, E. Nov- [296] J. A. Eble, B. Beermann, H.-J. Hinz, and A. Schmidt- ellino, and H. Kessler, “Ligand binding analysis for human Hederich, “α2β1 integrin is not recognized by rhodocytin but α5β1 integrin: strategies for designing new α5β1 integrin is the specific, high affinity target of rhodocetin, an RGD- antagonists,” Journal of Medicinal Chemistry, vol. 48, no. 13, independent disintegrin and potent inhibitor of cell adhesion pp. 4204–4207, 2005. to collagen,” Journal of Biological Chemistry, vol. 276, no. 15, [312] N. Umeda, S. Kachi, H. Akiyama et al., “Suppression pp. 12274–12284, 2001. and regression of choroidal neovascularization by systemic [297] J. A. Eble, S. Niland, A. Dennes, A. Schmidt-Hederich, P. administration of an α5β1 integrin antagonist,” Molecular Bruckner, and G. Brunner, “Rhodocetin antagonizes stromal Pharmacology, vol. 69, no. 6, pp. 1820–1828, 2006. tumorinvasioninvitro andother α2β1 integrin-mediated [313] S. K. Kuwada, “Volociximab, an angiogenesis-inhibiting cell functions,” Matrix Biology, vol. 21, no. 7, pp. 547–558, chimeric monoclonal antibody,” Current Opinion in Molec- ular Therapeutics, vol. 9, no. 1, pp. 92–98, 2007. [298] J. Zhou, V. L. Rothman, I. Sargiannidou et al., “Cloning and [314] M. L. Wahl, T. L. Moser, and S. V. Pizzo, “Angiostatin and characterization of angiocidin, a tumor cell binding protein anti-angiogenic therapy in human disease,” Recent Progress for thrombospondin-1,” Journal of Cellular Biochemistry, vol. in Hormone Research, vol. 59, pp. 73–104, 2004. 92, no. 1, pp. 125–146, 2004. [315] D. Zhang, P. L. Kaufman, G. Gao, R. A. Saunders, and J. [299] Y. Sabherwal, V. L. Rothman, S. Dimitrov et al., “Integrin X. Ma, “Intravitreal injection of plasminogen kringle 5, an α2β1 mediates the anti-angiogenic and anti-tumor activities endogenous angiogenic inhibitor, arrests retinal neovascu- of angiocidin, a novel tumor-associated protein,” Experimen- larization in rats,” Diabetologia, vol. 44, no. 6, pp. 757–765, tal Cell Research, vol. 312, no. 13, pp. 2443–2453, 2006. [300] R. C. Pandey, M. W. Toussaint, J. C. McGuire, and [316] W.R. Ji, F. J. Castellino, Y. Chang et al., “Characterization of M. C. Thomas, “Maggiemycin and anhydromaggiemycin: kringle domains of angiostatin as antagonists of endothelial two novel anthracyclinone antitumor antibiotics—isolation, cell migration, an important process in angiogenesis,” FASEB structures, partial synthesis and biological properties,” Jour- Journal, vol. 12, no. 15, pp. 1731–1738, 1998. nal of Antibiotics, vol. 42, no. 11, pp. 1567–1577, 1989. [317] Y. Hamano and R. Kalluri, “Tumstatin, the NC1 domain of [301] J. Kap ¨ yla, ¨ O. T. Pentikainen, ¨ T. Nyronen ¨ et al., “Small α3 chain of type IV collagen, is an endogenous inhibitor molecule designed to target metal binding site in the α2I of pathological angiogenesis and suppresses tumor growth,” domain inhibits integrin function,” Journal of Medicinal Biochemical and Biophysical Research Communications, vol. Chemistry, vol. 50, no. 11, pp. 2742–2746, 2007. 333, no. 2, pp. 292–298, 2005. [302] L. Nissinen, O. T. Pentikainen, ¨ A. Jouppila et al., “A small- [318] Y. Maeshima, P. C. Colorado, and R. Kalluri, “Two RGD- molecule inhibitor of integrin α2β1 introduces a new strategy independent α(v)β3 integrin binding sites on tumstatin for antithrombotic therapy,” Thrombosis and Haemostasis, regulate distinct anti-tumor properties,” Journal of Biological vol. 103, no. 2, pp. 387–397, 2010. Chemistry, vol. 275, no. 31, pp. 23745–23750, 2000. [303] Y. Funahashi, N. H. Sugi, T. Semba et al., “Sulfonamide [319] N. Floquet, S. Pasco, L. Ramont et al., “The antitumor derivative, E7820, is a unique angiogenesis inhibitor sup- properties of the α3(IV)-(185–203) peptide from the NC1 pressing an expression of integrin α2 subunit on endothe- domain of type IV collagen (tumstatin) are conformation- lium,” Cancer Research, vol. 62, no. 21, pp. 6116–6123, 2002. dependent,” Journal of Biological Chemistry, vol. 279, no. 3, [304] M. S. O’Reilly, T. Boehm, Y. Shing et al., “Endostatin: an pp. 2091–2100, 2004. endogenous inhibitor of angiogenesis and tumor growth,” [320] C. Magnon, A. Galaup, B. Mullan et al., “Canstatin acts Cell, vol. 88, no. 2, pp. 277–285, 1997. on endothelial and tumor cells via mitochondrial damage [305] R. S. Herbst, K. R. Hess, H. T. Tran et al., “Phase I study initiated through interaction with αvβ3and αvβ5 integrins,” of recombinant human endostatin in patients with advanced Cancer Research, vol. 65, no. 10, pp. 4353–4361, 2005. solid tumors,” Journal of Clinical Oncology, vol. 20, no. 18, pp. [321] E. Petitclerc, A. Boutaud, A. Prestayko et al., “New functions 3792–3803, 2002. for non-collagenous domains of human collagen type IV. [306] J. Dixelius, H. Larsson, T. Sasaki et al., “Endostatin-induced Novel integrin ligands inhibiting angiogenesis and tumor tyrosine kinase signaling through the Shb adaptor protein growth in vivo,” Journal of Biological Chemistry, vol. 275, no. regulates endothelial cell apoptosis,” Blood, vol. 95, no. 11, 11, pp. 8051–8061, 2000. pp. 3403–3411, 2000. [322] P. C. Brooks, S. Silletti, T. L. Von Schalscha, M. Friedlander, [307] S. A. Karumanchi, V. Jha, R. Ramchandran et al., “Cell and D. A. Cheresh, “Disruption of angiogenesis by PEX, surface glypicans are low-affinity endostatin receptors,” a noncatalytic metalloproteinase fragment with integrin Molecular Cell, vol. 7, no. 4, pp. 811–822, 2001. binding activity,” Cell, vol. 92, no. 3, pp. 391–400, 1998. 24 Journal of Oncology [323] L. Bello, V. Lucini, G. Carrabba et al., “Simultaneous inhibi- integrin alpha(v)beta(3), + or - dacarbazine in patients with tion of glioma angiogenesis, cell proliferation, and invasion stage IV metastatic melanoma,” Cancer, vol. 116, no. 6, pp. by a naturally occurring fragment of human metallopro- 1526–1534, 2010. teinase-2,” Cancer Research, vol. 61, no. 24, pp. 8730–8736, [338] S. J. O’Day, A. C. Pavlick, M. R. Albertini et al., “Clinical and pharmacologic evaluation of two dose levels of intetumumab (CNTO 95) in patients with melanoma or angiosarcoma,” [324] J.-O. Nam, H.-W. Jeong, B.-H. Lee, R.-W. Park, and I.-S. Kim, “Regulation of tumor angiogenesis by fastatin, the fourth Investigational New Drugs. In press. FAS1 domain of βig-h3, via αvβ3 integrin,” Cancer Research, [339] J. A. Varner, M. T. Nakada, R. E. Jordan, and B. S. Coller, vol. 65, no. 10, pp. 4153–4161, 2005. “Inhibition of angiogenesis and tumor growth by murine [325] C. Mas-Moruno, F. Rechenmacher, and H. Kessler, “Cilengi- 7E3, the parent antibody of c7E3 Fab (abciximab; ReoPro),” Angiogenesis, vol. 3, no. 1, pp. 53–60, 1999. tide: the first anti-angiogenic small molecule drug candidate. Design, synthesis and clinical evaluation,” Anti-Cancer Agents [340] M. T. Nakada, G. Cao, P. M. Sassoli, and H. M. DeLisser, in Medicinal Chemistry, vol. 10, no. 10, pp. 753–768, 2010. “c7E3 Fab inhibits human tumor angiogenesis in a SCID mouse human skin xenograft model,” Angiogenesis, vol. 9, no. [326] K. E. Shannon, J. L. Keene, S. L. Settle et al., “Anti-metastatic properties of RGD-peptidomimetic agents S137 and S247,” 4, pp. 171–176, 2006. Clinical and Experimental Metastasis, vol. 21, no. 2, pp. 129– [341] F. Mitjans, T. Meyer, C. Fittschen et al., “In vivo therapy 138, 2004. of malignant melanoma by means of antagonists of αv [327] A. Abdollahi, D. W. Griggs, H. Zieher et al., “Inhibition of integrins,” International Journal of Cancer, vol. 87, no. 5, pp. 716–723, 2000. αvβ3 integrin survival signaling enhances antiangiogenic and antitumor effects of radiotherapy,” Clinical Cancer Research, [342] A. R. Reynolds, I. R. Hart, A. R. Watson et al., “Stimulation vol. 11, no. 17, pp. 6270–6279, 2005. of tumor growth and angiogenesis by low concentrations of RGD-mimetic integrin inhibitors,” Nature Medicine, vol. 15, [328] N. E. Tsopanoglou, M. E. Papaconstantinou, C. S. Flordellis, no. 4, pp. 392–400, 2009. and M. E. Maragoudakis, “On the mode of action of thrombin-induced angiogenesis: thrombin peptide, TP508, [343] D. Hanahan, “A flanking attack on cancer,” Nature Medicine, mediates effects in endothelial cells via α vβ3 integrin,” vol. 4, no. 1, pp. 13–14, 1998. Thrombosis and Haemostasis, vol. 92, no. 4, pp. 846–857, [344] R. K. Jain, “Normalization of tumor vasculature: an emerging concept in antiangiogenic therapy,” Science, vol. 307, no. 5706, pp. 58–62, 2005. [329] K. Meerovitch, F. Bergeron, L. Leblond et al., “A novel RGD antagonist that targets both αvβ3and α5β1 induces apoptosis [345] G. Huang and L. Chen, “Tumor vasculature and microenvi- of angiogenic endothelial cells on type I collagen,” Vascular ronment normalization: a possible mechanism of antiangio- Pharmacology, vol. 40, no. 2, pp. 77–89, 2003. genesis therapy,” Cancer Biotherapy and Radiopharmaceuti- [330] C. C. Kumar, M. Malkowski, Z. Yin et al., “Inhibition of cals, vol. 23, no. 5, pp. 661–667, 2008. angiogenesis and tumor growth by SCH 221153, a dual αvβ3 [346] G. Bergers and D. Hanahan, “Modes of resistance to anti- and αvβ5 integrin receptor antagonist,” Cancer Research, vol. angiogenic therapy,” Nature Reviews Cancer, vol. 8, no. 8, pp. 61, no. 5, pp. 2232–2238, 2001. 592–603, 2008. [331] L. Belvisi, T. Riccioni, M. Marcellini et al., “Biological [347] P. Fraisl, M. Mazzone, T. Schmidt, and P. Carmeliet, and molecular properties of a new αvβ3/αvβ5 integrin “Regulation of angiogenesis by oxygen and metabolism,” antagonist,” Molecular Cancer Therapeutics, vol. 4, no. 11, pp. Developmental Cell, vol. 16, no. 2, pp. 167–179, 2009. 1670–1680, 2005. [348] A. Rapisarda and G. Melillo, “Role of the hypoxic tumor [332] K. Minamiguchi, H. Kumagai, T. Masuda, M. Kawada, M. microenvironment in the resistance to anti-angiogenic thera- Ishizuka, and T. Takeuchi, “Thiolutin, an inhibitor of huvec pies,” Drug Resistance Updates, vol. 12, no. 3, pp. 74–80, 2009. adhesion to vitronectin, reduces paxillin in huvecs and [349] S. J. Lunt, N. Chaudary, and R. P. Hill, “The tumor microen- suppresses tumor cell-induced angiogenesis,” International vironment and metastatic disease,” Clinical and Experimental Journal of Cancer, vol. 93, no. 3, pp. 307–316, 2001. Metastasis, vol. 26, no. 1, pp. 19–34, 2009. [333] R. Soldi, S. Mitola, M. Strasly, P. Defilippi, G. Tarone, and [350] K. De Bock, S. Cauwenberghs, and P. Carmeliet, “Vessel F. Bussolino, “Role of α(v)β3 integrin in the activation abnormalization: another hallmark of cancer? Molecular of vascular endothelial growth factor receptor-2,” EMBO mechanisms and therapeutic implications,” Current Opinion Journal, vol. 18, no. 4, pp. 882–892, 1999. in Genetics and Development, vol. 21, no. 1, pp. 73–79, 2010. [334] J. C. Gutheil, T. N. Campbell, P. R. Pierce et al., “Targeted [351] A. R. Reynolds, “Potential relevance of bell-shaped and antiangiogenic therapy for cancer using vitaxin: a humanized u-shaped dose-responses for the therapeutic targeting of monoclonal antibody to the integrin α(v)β3,” Clinical Cancer angiogenesis in cancer,” Dose-Response, vol. 8, no. 3, pp. 253– Research, vol. 6, no. 8, pp. 3056–3061, 2000. 284, 2010. [335] D. G. McNeel, J. Eickhoff,F.T.Lee et al., “Phase Itrial of [352] S. De, O. Razorenova, N. P. McCabe, T. O’Toole, J. Qin, a monoclonal antibody specific for α vβ3 integrin (MEDI- and T. V. Byzova, “VEGF—integrin interplay controls tumor 522) in patients with advanced malignancies, including an growth and vascularization,” Proceedings of the National assessment of effect on tumor perfusion,” Clinical Cancer Academy of Sciences of the United States of America, vol. 102, Research, vol. 11, no. 21, pp. 7851–7860, 2005. no. 21, pp. 7589–7594, 2005. [336] D. Zhang, T. Pier, D. G. McNeel, G. Wilding, and A. Friedl, [353] G. H. Mahabeleshwar, J. Chen, W. Feng, P. R. Somanath, O. V. “Effects of a monoclonal anti-αvβ3 integrin antibody on Razorenova, and T. V. Byzova, “Integrin affinity modulation blood vessels—a pharmacodynamic study,” Investigational in angiogenesis,” Cell Cycle, vol. 7, no. 3, pp. 335–347, 2008. New Drugs, vol. 25, no. 1, pp. 49–55, 2007. [354] P. R. Somanath, A. Ciocea, and T. V. Byzova, “Integrin [337] P. Hersey, J. Sosman, S. O’Day et al., “A randomized phase and growth factor receptor alliance in angiogenesis,” Cell 2 study of etaracizumab, a monoclonal antibody against Biochemistry and Biophysics, vol. 53, no. 2, pp. 53–64, 2009. Journal of Oncology 25 [355] P. R. Somanath, N. L. Malinin, and T. V. Byzova, “Cooper- ation between integrin ανβ3 and VEGFR2 in angiogenesis,” Angiogenesis, vol. 12, no. 2, pp. 177–185, 2009. [356] A. Cretu, J. M. Roth, M. Caunt et al., “Disruption of endothe- lial cell interactions with the novel Hu177 cryptic collagen epitope inhibits angiogenesis,” Clinical Cancer Research, vol. 13, no. 10, pp. 3068–3078, 2007. [357] K. Chen and X. Chen, “Integrin targeted delivery of chemotherapeutics,” Theranostics, vol. 1, pp. 189–200, 2011. [358] Z. Wang, W. K. Chui, and P. C. Ho, “Integrin targeted drug and gene delivery,” Expert Opinion on Drug Delivery, vol. 7, no. 2, pp. 159–171, 2010. [359] K. N. Sugahara, T. Teesalu, P. Prakash Karmali et al., “Coad- ministration of a tumor-penetrating peptide enhances the efficacy of cancer drugs,” Science, vol. 328, no. 5981, pp. 1031–1035, 2010. [360] G. J. Strijkers, E. Kluza, G. A. F. Van Tilborg et al., “Param- agnetic and fluorescent liposomes for target-specific imaging and therapy of tumor angiogenesis,” Angiogenesis, vol. 13, no. 2, pp. 161–173, 2010. [361] A. J. Beer, H. Kessler, H. J. Wester, and M. Schwaiger, “PET Imaging of Integrin alphaVbeta3 expression,” Theranostics, vol. 1, pp. 48–57, 2011. [362] F. Kiessling, J. Gaetjens, and M. Palmowski, “Application of molecular ultrasound for imaging integrin expression,” Theranostics, vol. 1, pp. 127–134, 2011. [363] E. Mery, E. Jouve, S. Guillermet et al., “Intraoperative flu- orescence imaging of peritoneal dissemination of ovarian carcinomas. A preclinical study,” Gynecologic Oncology, vol. 122, no. 1, pp. 155–162, 2011. 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Integrin-Mediated Cell-Matrix Interaction in Physiological and Pathological Blood Vessel Formation

Journal of Oncology , Volume 2012 – Sep 18, 2011

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Hindawi Publishing Corporation
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Copyright © 2012 Stephan Niland and Johannes A. Eble. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
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1687-8469
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10.1155/2012/125278
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Abstract

Hindawi Publishing Corporation Journal of Oncology Volume 2012, Article ID 125278, 25 pages doi:10.1155/2012/125278 Review Article Integrin-Mediated Cell-Matrix Interaction in Physiological and Pathological Blood Vessel Formation Stephan Niland and Johannes A. Eble Center for Molecular Medicine, Department of Vascular Matrix Biology, Excellence Cluster Cardio-Pulmonary System, J. W. Goethe University Hospital, Theodor-Stern-Kai 7, Building 9 b, 60590 Frankfurt, Germany Correspondence should be addressed to Johannes A. Eble, eble@med.uni-frankfurt.de Received 25 May 2011; Accepted 15 July 2011 Academic Editor: Debabrata Mukhopadhyay Copyright © 2012 S. Niland and J. A. Eble. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Physiological as well as pathological blood vessel formation are fundamentally dependent on cell-matrix interaction. Integrins, a family of major cell adhesion receptors, play a pivotal role in development, maintenance, and remodeling of the vasculature. Cell migration, invasion, and remodeling of the extracellular matrix (ECM) are integrin-regulated processes, and the expression of certain integrins also correlates with tumor progression. Recent advances in the understanding of how integrins are involved in the regulation of blood vessel formation and remodeling during tumor progression are highlighted. The increasing knowledge of integrin function at the molecular level, together with the growing repertoire of integrin inhibitors which allow their selective pharmacological manipulation, makes integrins suited as potential diagnostic markers and therapeutic targets. 1. Introduction growth, the tumor needs to hook up to the vascular system by forming neovessels. Invasive cancer is among the leading causes of death world- During tumor progression, an angiogenic switch is acti- wide, and rates are still increasing, due to ageing and vated causing a continuous neovessel formation emanating changes in lifestyle [1]. Cancer is a collective term for many from the normally quiescent vasculature, which sustains diseases, rather than a single disease, with the common tumor growth [6]. This process called tumor angiogenesis characteristic that tissue growth goes haywire [2]. Patients is a collective term that is generally used for all types of who have undergone cancer treatment show an increased risk tumor neovascularization. In addition to vessel co-option of developing a second tumor, mainly due to the same risk and to endothelial cell (EC) sprouting, tumor vessels can also factors that were responsible for the first tumor but also in develop by intussusceptive or glomerular angiogenesis, or, part due to the treatment of the first tumor with mutagenic in a way of vascular mimicry, even tumor cells themselves chemotherapeutics or radiation [3]. Therefore, new strate- can form vessel-like hollow structures. These types of vessel gies for cancer treatment with as little as possible adverse formation can occur in parallel, and also gradual transitions side effects are needed that effectively eradicate the primary are possible. Vessel formation by the latter types requires less tumor and also do not increase the risk of recurrence. energy than sprouting angiogenesis, is thus carried out faster, A tumor initially grows without any connection to the and usually can be observed in, for example, gliosarcoma vasculature until it reaches a critical size of about two mm multiforme, melanoma, and breast and colon cancer [7]. in diameter. Then it remains in a dormant state, in which For neovessel formation, ECs need to migrate into a pre- proliferation and apoptosis due to lack of oxygen, are in a viously avascular region and to extensively remodel the dynamic equilibrium unless it develops in a well-vascularized extracellular matrix (ECM). In this process, integrins, which region or is able to recruit its own vasculature. Hanahan are cell adhesion receptors for various ECM proteins and Weinberg have proposed six hallmarks of cancer, one of and immunoglobulin superfamily molecules, are the most them being the induction of angiogenesis [4, 5]. For further important matrix receptors [8, 9]. Therefore, integrins are 2 Journal of Oncology appealing targets for cancer therapy using a variety of inte- In different vessel types, that is, arteries, arterioles, cap- grin-specific antagonists, ranging from endogenous antago- illaries, venules, and veins, this general blueprint is mod- nists over humanized or chimeric antibodies to peptides and ified corresponding to the respective functional require- small nonpeptidic compounds [10–12]. ments. For example, endothelia, which are continuous in most instances, can become fenestrated, as in exocrine or In this paper, based on the general assembly of blood endocrine gland tissues, or even discontinuous, as in liver, vessels, the specific organization of tumor vasculature will be spleen, or bone marrow, in order to facilitate the exchange described, as well as the dynamic sequence of events by which of hormones or metabolites. Elastic and muscular arteries a tumor gains access to the body’s vasculature. In this context, illustrate other examples for a modification of this general the role of integrins and possibilities of their pharmacological blueprint. In order to even the pulsatile blood flow coming manipulation are explored. from the heart, the proteins elastin and fibrillin are abundant in the tunica media ECM of elastic arteries, which is the 2. The Static Picture: The Extracellular Matrix direct cause for the vessel wall’s elastic properties. Muscular of Blood Vessels arteries possess numerous concentric sheaths of smooth muscle cells. By means of vasoconstriction and vasodilation, The tissue’s ECM is a structure-shaping molecular scaffold they can distribute and direct the blood to different organs. and also a repository for cytokines and other growth factors [13]. Cells embedded in this matrix need to be supplied 2.2. Extracellular Matrix in the Vessel Wall. The ECM of with oxygen and nutrients, signaling molecules need to be blood vessels together with their resident cells contributes to received and emitted, and metabolic waste products need essentially all physiological functions of blood vessels and has to be disposed of. These tasks are optimally fulfilled by the been reviewed recently [15]. cardiovascular system with its intricate and dynamic network The subendothelial basement membrane (BM) com- of blood vessels. Depending on their functions, different partmentalizes the vessel’s single-layered endothelium from types of blood vessels show special histological and molecular the vascular connective tissue. The molecular architecture adaptations. The heart, as a double-acting pump, drives of BMs has recently been reviewed [16–18]. Fibronectin, the blood circulation within the vasculature via the aorta incorporated between endothelial and perivascular cells, is through arteries and arterioles into capillaries, from where essential for blood vessel morphogenesis [19]. The presence the blood flows back through venules and veins. Due to of von Willebrand factor (vWF) is characteristic for the the prevailing pressure conditions, the body fluid is forced subendothelial BM, where also other BM proteins, such as through the vessel wall to form the lymph, which then is the network-forming collagens IV and XVIII can be found, drained by lymph vessels back to the blood circulation. together with laminins, nidogens, and perlecan. Thirteen Additionally, the vasculature serves as “highway” system different collagens are present in the vascular wall [20, 21]. for leukocytes to patrol the body during immunological The network-forming collagen IV [22] plays a key role for surveillance and to quickly reach sites of inflammation. The the mechanical stability of the BM [23], which, especially in vascular wall is capable of self-sealing upon smaller injuries, arterial regions of the circulatory system, has to withstand a and leukocytes are able to penetrate the blood vessel wall considerable blood pressure. in a complex interplay without any obvious vessel leakage. In the tunica media of elastic and muscular arteries, Pathologically, tumor cells capitalize the blood vessel system covalently crosslinked supramolecular aggregates of elastin to disseminate from a primary tumor and to colonize distant form concentric lamellae and fibers in a proportion of up organs where they develop metastases. to 50% of the vessel’s dry weight and confer resilience to pulsatile blood flow [24–26]. Regions of the ECM that 2.1. General Organization of the Vessel Wall. Histologically, consist mostly of elastin are confined by EMILINs, that is, the walls of blood vessels comprise three concentric layers, homotrimeric elastin microfibril interphase-located proteins that is, tunica intima, tunica media, and tunica adventitia [27]. Anchored to microfibrillar bridges of fibrillin-1 and [14], which are separated by two sheet-like structures of fibulin-5 between these concentric elastin lamellae, vascular ECM proteins. The membrana limitans interna and externa smooth muscle cells (VSMCs) are sandwiched in a fishbone- establish a border between tunica media and tunica interna like pattern and thus can effectively regulate the vessel’s and adventitia, respectively. These ECM sheaths tightly caliber [25, 28–31]. Dependent on the vessel type, distinct connect the cell layers of the vessel wall to form a functional fibulins are involved in the assembly of the ECM. While unit, which becomes evident when too weak cell-matrix fibulin-1 is widespread and occurs in the BMs of all blood interactions lead to life-threatening aneurysms. vessels, heart valves and septa, fibulin-3, and fibulin-4 occur The tunica intima comprises a single layer of squamous in the walls of capillaries and larger blood vessels [32]. The ECs and lines the inner surface of all blood vessels. The innermost and outermost elastic lamellae are referred to as tunica media, which is usually the thickest layer in arteries, membrana limitans interna and membrana limitans externa, is composed of mural cells, which are smooth muscle cells respectively. Between the elastic lamellae, type I and III in larger blood vessels and pericytes in capillaries. The collagens are deposited that bear tensile forces exerted on tunica adventitia finally interconnects the blood vessel with the vessels and limit their elastic dilatability. In contrast, in the surrounding connective tissue, and it is usually most the interstitial connective tissue between the subendothelial prominent in veins. membrane and the membrana limitans interna, type VI and Journal of Oncology 3 type VIII collagens are found [21, 33]. The connection of the osteopontin receptor α8β1, and integrin αvβ3 is also the membrana limitans interna to the subendothelial BM by expressed on glial cells [49]. type XVIII collagen is assumed [34]. Also type XVI collagen, As EC-derived tumors, angiosarcomas express the inte- which is produced by VSMCs and found close to both grins α1β1, α2β1, α3β1, α5β1, and α6β1, and in benign and elastic microfibrils and fibrillar type I and type III collagens, malignant mesenchymal tumors as well as in the desmoplas- may contribute to the connection between the elastic and tic stroma of carcinomas, integrins α1β1and α5β1are widely collagenous phases of the ECM [35, 36], especially, as type distributed [50]. Integrins α1β1and α2β1 bind to the same XVI collagen contains a binding site for the major collagen ligand in the ECM and are VEGF-dependently upregulated receptor on VSMCs, integrin α1β1[37, 38]. on migrating ECs, and antagonists against both integrins The ECM of the tunica media is synthesized by VSMCs, inhibit VEGF-mediated angiogenesis without affecting the which are all encapsulated by an (incomplete) BM containing existing vasculature [51, 52]. Therefore, and against the the usual BM proteins, type IV collagen and laminins background of gene ablation studies, they are believed to dif- [33, 39]. Depending on microenvironmental cues, VSMCs ferentially regulate angiogenesis [49]. Important coreceptors can reversibly acquire distinct phenotypes, which can be for integrin α2β1 are the syndecans-1 and -4, which weaken characterized as either (i) contractile and differentiated or the invasiveness of tumor cells into a collagenous matrix [53]. (ii) secretory, migratory, and less differentiated [37, 39]. Cells bind to fibronectin and vitronectin preferentially Under physiological conditions, the contractile phenotype via the RGD-dependent integrins αvβ3and α5β1[54]. prevails, at which the VSMCs transduce forces on the Fibronectin can also be bound by the leukocyte-specific pericellular matrix especially by the collagen-binding inte- integrins α4β1and α4β7[55]. Cell-fibronectin interactions grin α1β1, by the laminin-binding integrin α7β1and by are modulated by proteoglycans, glycoproteins of the ECM, dystroglycan [37]. In contrast, in the secretory, proliferatory, and the coreceptors syndecans [56]. and migratory phenotype, the integrin equipment of the Integrin αvβ3 was identified as a marker for angiogenic VSMCs predominantly consists of the fibronectin receptor, vascular tissue [57]. In contrast to quiescent ECs, integrin α5β1, and the integrins α4β1and α9β1. Consistently, in the αvβ3 is highly expressed on activated ECs during tumor proximity of secretory VSMCs, the fibronectin splice variants angiogenesis,aswellasonsometumor cells[58, 59]. In the V (IIICS) and EIIIA with binding sites for the integrins α4β1, tumor microenvironment, angiogenic ECs can interact due α5β1, and α9β1 are abundant [39]. In capillaries, scattered to their increased levels of the integrins αvβ3and αvβ5 pericytes, each encapsulated by an own BM, stabilize the with provisional matrix proteins, such as vitronectin, fib- endothelium and its subendothelial BM [40–42]. rinogen, vWF, osteopontin, and fibronectin. Also, partially The fibroelastic connective tissue of the tunica adventitia proteolyzed collagen in the tumor exposes RGD sites and is connects the blood vessel with the perivascular connective a further ligand for integrin αvβ3[60]. Thus, the ECM of tissue. It is rich in versican, a glycoprotein, which can interact the tumor microenvironment both provides survival signals with fibrillin-1 [43], fibulin-1 [44], and fibulin-2 [45], as well and facilitates invasion. Integrin-αvβ3-mediated adhesion as with other ECM molecules. to platelets protects malignant cells from clearance through the immune system, and moreover, αvβ3 integrin also helps tumor cells to adhere to the vessel endothelium and to spread 2.3. Receptors for ECM Molecules. To interact with their microenvironment and to spatiotemporally regulate their into adjacent tissues [61]. differentiation state, morphology, metabolism, and survival, The pharmacological inhibition of integrin-αvβ3- mediated cell-matrix interaction impedes tumor angiog- cells are equipped with a variety of receptors for all the ECM molecules [13]. Integrins are the largest family of these enesis and growth [62], as does a replacement of the β3 receptors, and they mediate adhesion to collagens, laminins, subunit with a mutated nonphosphorylatable subunit in a and fibronectin. In addition, there are other receptors and murine model [63], which provides evidence for a proan- coreceptors, such as the syndecans [46]. giogenic role of integrin αvβ3, in contrast to integrin αvβ5, Binding to a wide variety of different ECM molecules which does not seem to play an essential role in angiogenesis and transmitting signals bi-directionally in an outside-in [64]. Interestingly, the analysis of αv-knock-out mice and inside-out manner, integrins constitute functional hubs, revealed that, despite being embryonic or perinatally lethal, the vascular endothelium was not impaired in the absence which, according to an interesting concept in network theory and systems biology, integrate networks of angiogenic sig- of the αv subunit, whereas the primary cause of death was naling cues that orchestrate the behavior of ECs and VSMCs brain hemorrhage [65–67]. Also endothelial Tie-2-specific knockout of the αv subunit did not result in any vascular during angiogenesis [47, 48]. Thus, therapeutically targeting integrins as the operationally important circuit-integrating or angiogenesis defect [67]. Moreover, in an integrin hubs rather than single pathways of the complex system may subunit β3- and also β5-deficient mouse model, pathologic result in a more pronounced inhibition of angiogenesis [47]. angiogenesis and tumor growth are increased [68]. A ECs express the vitronectin receptors αvβ3and αvβ5; possible cause for these seemingly contradictory phenomena could be a relief of a transdominant inhibition by αvβ3on moreover, on ECs and pericytes the following integrins are expressed: the collagen receptors α1β1and α2β1, the laminin other integrins or other molecules, which would enhance receptors α3β1, α3β6, and α6β4, the osteopontin receptor their proangiogenic function [69, 70]. Likewise, there could be a compensatory role of other integrins with overlapping α9β1, and the fibronectin receptors α4β1and α5β1[49]. Pericytes additionally express the laminin receptor α7β1, and function [49]. Moreover, inhibition could also stabilize the 4 Journal of Oncology integrin αvβ3 in its unligated conformation and thus induce interaction with a multitude of proteins, such as MMPs, apoptosis by triggering an integrin-mediated death program uPA/uPAR, tissue inhibitor of matrixmetalloproteinase-2 [71]. (TIMP-2), vWF, TSP-1, osteopontin, syndecan-1, insulin- Integrin αvβ8isimportant forvasculardevelopment in receptor substrate-1 (IRS-1), cytohesin-1, integrin cytoplas- the embryonic brain and in the yolk sac [72]. It is expressed mic domain-associated protein-1 (ICAP-1), integrin-linked on astrocytes but not on ECs or pericytes, nevertheless plays kinase (ILK), calcium- and integrin-binding protein (CIB), an important role in angiogenesis, as it binds in addition to β3-endotoxin, talin, actinin, tensin, nischarin, and the Ras- several ECM proteins also to the latency-associated peptide related protein Rab 25 [9]. (LAP) of TGFβ1, which in cooperation with the membrane- The subendothelial BM of the tunica intima serves as type metalloproteinase MT1-MMP/MMP14 results in acti- a mechanical support to which ECs are anchored by vari- vation of TGFβ and triggering of its downstream signal ous adhesion molecules, especially integrins [46, 105–108]. cascades [73–75]. Additionally, the subendothelial BM provides microenviron- Collagen IV, an essential component of BMs, is bound mental information that regulate the metabolic activity of by integrin α1β1, whichisexpressedonmesenchymal cells attached ECs, such as their production of leukocyte adhesion and can also bind to other collagens [76, 77]. Further molecules [107] or antithrombotic prostacyclins [109], as collagen-binding integrins are α2β1, the main receptor for well as other properties, for example, the tightness of inter- fibrillar collagens, which is expressed on epithelial and some cellular contacts [108]. Therefore, angiogenesis is regulated mesenchymal cells as well as on thrombocytes [78], α10β1in not least by integrins which are adhesion receptors for cartilage [79], and α11β1, a key receptor for fibrillar collagen matricellular proteins, ECM proteins, and immunoglobulin on fibroblasts [80]. The integrins α1β1and α2β1are involved superfamily molecules, on nearly all cells including ECs in the regulation of collagen and MMP synthesis and thus [8, 58]. of special importance for ECM turnover [81–83]. Discoidin In addition to their mechanical function [110], integrins domain receptors DDR1 on epithelial cells and DDR2 on also assist growth factor receptors and play important roles mesenchymal cells are further collagen receptors with tyro- in signaling processes, in particular as soluble growth factors, sine kinase function and are relevant for cancer [84]. and other signaling molecules are bound by integrins as well Other collagen receptors are glycoprotein GPIV on platelets [111]. For example, the proangiogenic VEGF-A is bound [85], the leukocyte-associated immunoglobulin-like receptor by integrins αvβ3and a3β1[112] and also by the tenascin- LAIR-1/CD305 [86], and the urokinase-type plasmino- C- and osteopontin-receptor integrin α9β1[113]. The latter gen activator receptor-associated protein uPARAP/Endo180, integrin, furthermore, binds the lymphangiogenic growth which is involved in matrix turnover during malignancy [87]. factors VEGF-C and VEGF-D [114]. Angiopoietins-1 and Laminin, as a further integral component of BMs, is -2 are bound by integrin α5β1[115]. Integrin α6β1isa bound by the integrins α3β1, α6β1, α6β4, and α7β1[88– receptor for the proangiogenic CCN-family member CYR61, 91] and also by α-dystroglycan [92, 93] and by the 67 kDa and is involved in in vivo in tube formation [116, 117]. The laminin receptor 67LR [94]. 67LR is increased in various fibronectin receptor integrin αvβ3, which is the best-studied tumors and correlates with their metastatic potential [95, integrin in relation to angiogenesis and is upregulated during 96]. The different laminin receptors may also act coopera- wound healing and retinal vascularization and especially on tively in laminin binding, for example, laminin-binding β1 tumor blood vessels, also binds to fibroblast growth factor integrins and 67LR [97] or integrin α6β4 and syndecan 1 FGF-1 [118]. Semaphorin 7A binding is also reported for the [98]. collagen receptor integrin α1β1[119]. Integrin α3β1, which in the vascular wall binds to lam- Stimulated by PDGF, vascular smooth muscle cells inins-411 (laminin 8) and-511 (laminin 10), thrombospon- express the laminin receptor integrin α7β1, which plays an din (TSP), TIMP2, tetraspanin CD151, and to the C-termi- important role in recruitment and differentiation of VSMCs nal domain of the collagen IV α3 chain, is controversially [120, 121]. ascribed either a positive or a negative role in angiogenesis Integrin α9β1 is not only involved in lymphangiogenesis (cf. [99]). [114] but also plays a role in EC adhesion [122]. While There is controversy whether the hemidesmosomal inte- binding of TSP-1 to integrin α9β1 promotes angiogenesis grin α6β4, whichisexpressedonasubset of ECs[100]and on [123], VEGF-A is another ligand of integrin α9β1[113]. tumor ECs [101], aggravates pathological angiogenesis [101] or whether it is a negative regulator of angiogenesis that is downregulated at its onset [102]. 2.4. Vascular-Relevant Integrin-Deficient Mouse Models. The Thus, many molecules of the ECM scaffold, for example, crucial involvement of integrins in EC biology has been laminins, collagens, fibronectin, and vitronectin, are ligands elucidated substantially by the examination of genetic knock- for integrins that link the cell’s cytoskeleton to the ECM. Loss out studies [124]. By ablation of the respective genes, the of this matrix-integrin contact triggers apoptotic cell death EC integrins α1β1, α2β1, α4β1, α5β1, α6β1, α6β4, α9β1, [103]. Picking up signals from the cell’s microenvironment, αvβ3, and αvβ5 and also the VSMC integrin α7β1 and the integrins functionally sense, interpret, and distribute infor- glial cell integrin αvβ8havebeenimplicatedinregulation mation, which allows the cell to modulate its proliferation, of cell growth, survival, and migration during angiogenesis differentiation, migration, and shape [104]. The modulatory (for recent reviews of the findings from knock-out mice cf. and regulating function of integrins is emphasized by direct [8, 10]). However, due to redundancy and compensatory Journal of Oncology 5 mechanisms, the interpretation of knock-out results is often angiogenesis [101], but die of severe skin defects [100]. In difficult. neovascularization, the endothelial expression of integrin Itgb1−/− mice die at E5.5 before they start to develop α6β1 is downregulated [102]. While it is not required for EC their vasculature [125, 126]. Mice with a conditional knock- proliferation and survival, it promotes tumor angiogenesis out in Tie-2-positive ECs survive until E9.5–E10.5, and [101]. In contrast, genetic ablation of α7β1, which is they are capable of vasculogenesis, but their angiogenesis is expressed on VSMCs but not on ECs, leads to incomplete disturbed showing defects in sprouting and branching [127– cerebral vascularization and hemorrhage and also to pla- 129]. Another endothelial-specific knockout of the integrin cental vascular defects, which results in partial embryonic β1 subunit is mediated via VE-cadherin-Cre recombinase lethality and demonstrates that integrin α7β1isimportant and becomes manifest later in embryogenesis resulting in for recruitment and survival of VSMCs [121, 137]. lethality between E13.5 and E17.5 [130]. In this mouse Deletion of Itga8 resulting in lack of integrin α8β1, model, loss of β1 integrin leads to a decreased expression a receptor for fibronectin and tenascin, results in partial of the cell polarity gene PAR3 and thus to disruption of EC embryonic lethality, but no defects in vascular development polarity and lumen formation [130]. (Mul ¨ ler and Reichardt, cited in [138]). Itga1−/− mice, deficient for the collagen-binding inte- Itga9−/− mice lacking integrin α9β1, which is the grin α1β1, show a normal vascular development and a receptor for tenascin-C, osteopontin, VCAM-1, and also for reduced tumor angiogenesis in adulthood, which has been VEGF-A, -C, and -D [113, 114], have defects in large lym- attributed to increased MMP activity [131], while α2β1- phatic vessels and die postnatally at P8-12 from a bilateral deficient Itga2−/− mice show an enhanced tumor angio- chylothorax [139]. genesis in adulthood, but an otherwise normal vascular Ablation of Itgav, resulting in simultaneous loss of the development [131, 132], and integrin α2β1isinvolvedin two integrins αvβ5, a receptor for vitronectin, osteopontin, the PlGF-dependent regulation of VEGFR-1 [132]. Although and Del-1 (developmental locus 1), and αvβ3, a recep- integrin α1β1and α2β1 bind to the same ligand in the tor for a variety of ECM proteins, such as fibronectin, ECM, their differential knockout results in opposing effects vitronectin, laminins, fibrinogen, fibrin, TSP, tenascin-C, on angiogenesis, suggesting a regulatory role for this pair of vWF, denatured collagen, osteopontin, MMP-2, Del-1, bone integrins. sialoprotein, FGF-2, thrombin, and CCN1 (cystein-rich Da Silva and coworkers generated EC-specific condi- protein 61), leads to 80% embryonic lethality at E9.5, and tional α3 integrin knock-out mice and showed that these the other 20% die at P0 with brain hemorrhage [65]. On mice, in contrast to a global ablation, are viable and fertile the other hand, Itgb3−/− mice, which are just integrin-αvβ3 but display enhanced tumor growth, elevated hypoxia- deficient, show 50% embryonic and early postnatal lethality induced retinal angiogenesis and tumor angiogenesis, and and an enhanced angiogenesis in surviving adult animals, increased VEGF-mediated neovascularization [99]. The indicating that this integrin is not strictly required for vascu- authors also could show that α3β1 is a positive regulator of lar development [140]. Surprisingly, animals with an intact EC-derived VEGF, which again represses VEGFR2 expres- but nonfunctional β3 integrin subunit develop normally but sion. Their data demonstrated that endothelial α3β1nega- show defects in angiogenesis in adulthood [63]. In contrast, tively regulates pathological angiogenesis and implicated an Itgb5−/− animals lacking integrin αvβ5develop normally unexpected role for low levels of EC-derived VEGF as an and angiogenesis is not significantly affected, indicating that activator of neovascularization. this integrin is not mandatory for vascular development [64]. Itga4−/− mice, deficient for fibronectin- and VCAM1- Integrins β3and β5 doubly deficient mice show enhanced binding integrin α4β1, are embryonic lethal with 50% dying tumor growth and angiogenesis. This strongly suggests that at E9.5–10.5 due to failure of chorion-allantois fusion and these integrins are not required for vascular development or 50% dying at E11.5 due to cardiovascular defects [55]. for pathological angiogenesis, pointing out that the mode of Mice, which by ablation of Itga5 are deficient for the action of αvβ3 antagonists and antiangiogenic therapeutics is fibronectin receptor integrin α5β1, show normal vasculo- still insufficiently understood [68]. Ablation of Itgb8 leads to genesis but no angiogenesis, which results in embryonic the loss of integrin αvβ8 on glial cells and thus to disrupted lethality at E10-11 due to defects in posterior somites, blood vessel formation in the brain, thereby demonstrating yolk sac, and embryonic vessels [133, 134]. This demon- that this integrin is mandatory for brain’s blood vessel strates the requirement of the integrin α5 subunit during development [72]. Moreover, the phenotype of β8-deficient embryonic development of early blood vessels and other mice resembles that of αv-deficient mice, which provides tissues. Accordingly, integrin α5β1, which is poorly expressed evidence that most defects in αv-deficient mice are due to on normal quiescent ECs, is markedly upregulated during the loss of integrin αvβ8[72]. tumor angiogenesis [135]. Among the laminin-binding integrins, integrin α6isnot 2.5. Integrin Structure. The family of integrins contains 24 essentially required for vascular development, although α6- structurally related N-glycosylated heterodimeric proteins deficiency is lethal with skin blistering defects resembling assembled noncovalently from 18 α-subunits and eight epidermolysis bullosa [136]. In line with the α6 knock- β-subunits. Each subunit comprises a large extracellular out mice, Itgb4−/− mice, lacking a functional laminin- domain, a single transmembrane domain, and with the binding integrin α6β4 by deletion of its signaling domain, exception of the β4 integrin subunit, a short noncatalytic show normal vascular development, although with reduced cytoplasmic tail [141]. Integrins are of special importance as α β 6 Journal of Oncology Collagen α β α β α ββ α α β 13 2 Integrin signaling Figure 1: Integrin activation. Integrins are a family of heterodimeric transmembrane adhesion receptors that bidirectionally relay signals with the extracellular matrix (ECM) and also with other cells. When activated, a conformational change increases the affinity, and clustering increases the avidity towards the ligand. (1) By inside-out signaling, integrins can reversibly undergo a conformational change from a bent inactive to an upright activated conformation with intermediate ligand affinity, at which the cytoplasmic domains are still close together. (2) Upon ligand binding, the integrin adopts a high-affinity conformation with a concomitant parting of the legs and a separation of the cytosolic α-and β-tails that unlocks docking sites for cytosolic molecules. (3) Clustering of ligand-occupied and activated integrins establishes a mechanical link between ECM and cytoskeleton and leads to the recruitment of scaffolding molecules and kinases. (4) The assembly of focal adhesions triggers intracellular signaling cascades. Details can be found in the text. they mediate cell matrix crosstalk via both outside-in and adopts an activated upright conformation [106, 151]. This inside-out signaling [54, 142]. Moreover, the 24 different conformational change is conveyed through the transmem- integrins possess promiscuous and redundant ligand speci- brane domains towards the cytoplasmic tails [54, 105, 152], ficities, which is of importance when distinct signals are to be where cytoskeletal proteins and signaling molecules relay the transduced or when in a particular context a defined cellular incoming signal intracellularly [153]. In inside-out signaling, response is elicited, as is discussed by Ruegg ¨ and Alghisi [11]. the binding of intracellular molecules, such as talin or Integrin structure and function have been studied in kindlins [154, 155], to the cytoplasmic integrin tails leads via detail at the molecular level [143, 144]. The extracellular a separation of the transmembrane domains [156]toaswitch headpiece is formed by a disk-like propeller domain of the blade-like erection of the extracellular domains [147, 157, α subunit and globular domains of the β subunit [145, 146]. 158]. Likewise, in outside-in signaling, ECM ligand binding The joint globular head harbors the ligand-binding site [146, to the integrin headpiece also induces a conformational 147]. The crystal structure of the integrin-αvβ3-binding site change in the hybrid domain and thereby a separation of with an inserted RGD ligand [148] helped to map functional the integrin subunits’ legs [144]. This parting of the legs amino acid residues on other integrins [149]. Recently, separates the cytosolic tails and allows binding of cytosolic the binding pocket of integrin α5β1has been mapped by proteins and thus clustering of integrins and formation of swapping regions of zebrafish and human α5 subunit in a focal adhesion sites (Figure 1). gain-of-function approach [150]. By clustering into focal adhesions, integrins recruit talin, paxillin, α-actinin, tensin, and vinculin and thereby mechan- 2.6. Integrin Signaling. Depending on their activity, integrins ically couple the ECM scaffold to the actin cytoskeleton. adopt distinct conformations (Figure 1). In the inactive rest- Additionally, integrins bind scaffolding molecules, such as ing conformation, the headpiece of the heterodimer bends p130 CRK/BCAR1, and recruit and activate kinases, such as towards the plasma membrane, and the transmembrane focal adhesion kinases (FAKs), Src family kinases (SFKs), and domains of the α and β subunits are associated [146]. Upon integrin-linked kinase (ILK), the latter forming a complex ligand binding, the previously bent integrin ectodomain with the adapter molecules parvin and PINCH/LIMS1 [159]. Inactive integrin Activatedintegrin Ligand-occupied integrin Clustered integrin Journal of Oncology 7 In addition, tetraspanins can recruit integrins to mem- the activation of PI3K by Ras is important for lymphangio- brane microdomains, thus regulating integrin function genesis [190]. [160]. Thereby, the rather unstable nascent adhesions are In addition to a direct activation of ERK, integrins can transformed into focal complexes, focal adhesions, fibrillar also activate a Raf/MEK/ERK signaling cascade in ECs [189, adhesions, or podosomes. This clustering of integrins leads 191, 192]. Raf-deficient and MEK-deficient mice have severe to a reorganization of the plasma membrane around the focal vascular defects [193, 194]. Growth-factor-mediated ERK adhesion into caveolin-containing lipid rafts, to which also signaling is linked with integrin-mediated signaling via FAK growth factor receptors often localize, and to the assembly [195]. Integrin-mediated ERK signaling is important for cell of adhesion signaling complexes [161–163]. This allows a proliferation and migration of ECs [191, 196]. Integrin α1β1 regulation of growth factor signals by integrin-mediated is unique among the collagen-binding integrins because it caveolae trafficking [164, 165]. In the assembly of such promotes cell proliferation by activating the Ras-Shc-MAPK integrin adhesions, up to 156 distinct molecules, amongst pathway, and cell cycle progression is regulated via FAK, other adaptor proteins, kinases, and phosphatases, are Rac, and cyclin D by integrin-mediated adhesion and matrix involved [48, 163]. Membrane lipid-protein interactions that stiffness [197–199]. modulate the homo- or heterotypic association of receptor Integrins can also activate the NF-κB pathway in ECs and molecules in the cell surface, or between adjacent cells, protect them from apoptosis [200–202]. Additionally, NF-κB have been reviewed recently [166]. From the focal adhesion signaling regulates the expression of cyclooxgenase-2 (COX- sites signal pathways diverge that regulate diverse cellular 2), which again is involved in EC spreading and migration programs, such as adhesion, migration, proliferation, and and in the induction of VEGF and FGF-2 [177, 203, survival. To provide an overview, integrins generally relay 204]. However, inhibition of the NF-κB pathway increases their signals via the FAK, ERK, and NF-κBpathways[153]. angiogenesis pathologically [205]. In most cases, in mechanosensory signaling FAK, Src, Integrins alone are not oncogenic, but some oncogenes and SH2, domains containing protein tyrosine phosphatase may depend on integrin signaling for tumor growth and 2(SHP2)are involved [167]. Upon integrin binding, FAK invasion. For example, integrin-triggered FAK signaling is autophosphorylates and binds to Src, which further phos- essential for Ras- and PI3K-mediated oncogenesis [206, phorylates FAK and several downstream binding partners, 207]. Also the expression of the cancer stem cell marker amongst others, JNK and Rho [168–170]. CD44 is integrin-regulated, and it can be speculated that Activated FAK also recruits PI3K, which mediates the integrin-relayed signals are needed to maintain a cancer activation of AKT and procures integrin-mediated cell sur- stem cell population [12, 208]. On the other hand, there vival, and likewise the antiapoptotic AKT can be activated via is evidence that the collagen receptor integrin α2β1has a Ang-1 [171]. Moreover, signals relayed via integrins and Src tumor-suppressing function [209, 210]. can be integrated by FAK with growth factor receptor-relayed Ligated integrins promote survival, whereas unligated signals via Ras, MEK, and MAPK [172]. Growth factors integrins recruit caspase-8 to the plasma membrane and can activate Ras signaling independently from integrin- promote apoptosis in a process termed integrin-mediated relayed adhesion signals. Nevertheless, MEK1 and Raf1 are death [71, 211], which differs from anoikis induced by loss important interfaces between integrin-relayed and growth- of cell adhesion to the ECM [103, 212]. Loss of caspase-8 factor-relayed signaling, because both MEK1 and Raf1 need confers resistance to integrin-mediated death of tumor cells, to be activated via adhesion-mediated activation of Src and and unligated integrin αvβ3 promotes the malignancy of FAK in order to activate MAPK [173, 174]. such tumors [213, 214]. Cell survival is promoted by integrin An endothelial-specific ablation of FAK results in im- ligation-dependent upregulation of BCL2 and FLIP/CFLAR, paired blood vessel development and embryonic lethality activation of the PI3K-AKT pathway, NF-κB signaling, [175] Downstream of FAK, Src couples integrin-mediated and p53 inactivation [176, 202, 215–217]. Survival is also and VEGF-receptor-mediated proangiogenic signaling in promoted by crosstalk between integrins and growth factor ECs [176–178]. However, endostatin can also activate Src receptors, for example, αvβ3 and FGFR or αvβ5 and VEGFR2 via integrin α5β1 and thereby disassemble actin stress fibers [195, 218]. and focal adhesions and thus inhibit cell migration, which is In various steps of angiogenesis and tumor progression, regulated by integrins via the Ras/ERK pathway [179–181]. crosstalk between integrins and growth factor receptors Important for adhesion and migration of endothelial and on tumor cells and also on host cells is important. This VSMCs are also p130Cas and PLC-γ, which can interact with crosstalk can consist in either an activation of a latent growth FAK [182–185]. factor, a regulation of common pathways for signaling or PI3K is of pivotal importance for angiogenesis, because internalization and recycling, a collaborative or a direct its deletion results in embryonic lethality E9.5 to E10.5, activation, or also a negative regulation [111]. The outcome when angiogenesis is important for vascular development. of a growth factor signal in a particular context is often PI3K deletion also causes decreased Tie-2 expression and determined by a synergistic and reciprocal interaction of thus creates a phenotype resembling Tie-2 deficiency [186, integrins with growth factor receptors, such as tyrosine 187]. Moreover, EC-specific deletion of the PI3K isoform kinase receptors like VEGFRs and Tie-2, Met, and FGFR, and p110α impairs angiogenesis [188]. In ECs, adhesion via semaphorins regulate integrin function as well [111, 219– integrins elicits a survival signal via FAK/PI3K/mTOR/4E- 221]. A complex of VEGF with the fibronectin heparin II BP1 and Cap-dependent translation [189]. Furthermore, domain increases, upon cell binding via integrin α5β1and 8 Journal of Oncology the signaling via VEGFR2 synergistically [222]. Expression of intensively studied one [237]. Mediated by HIF-1, VEGF-A integrin α11β1 on tumor-associated fibroblasts has a tumor- synergizes with FGF-2. VEGF is upregulated under hypoxic promoting effect, because it upregulates the expression and hypoglycemic conditions prevailing within tumor tissue of insulin-like growth factor 2 (IGF2), which is another [230]. example of integrin-regulated growth factor signaling [223]. The role of chemokines in tumor angiogenesis and neo- Beside binding ECM proteins and thus regulating adhe- vascularization has been reviewed recently [238]. Tumor sion and migration, integrins can also directly interact with cells express CCL2/MCP-1 (C-C-motif ligand 2/mono- pro- and antiangiogenic factors [221]. Integrin α5β1can cyte chemotactic protein-1), and thus, tumor-associated bind to matrix-bound VEGFR-1 [224]. In addition, integrin macrophages (TAMs) are recruited, resulting in an inflam- α9β1 can directly interact with VEGF-A, -C, and -D and matory response. These TAMs are again a source for also with hepatocyte growth factor (HGF) [113, 114, 225]. angiogenic growth factors, such as, VEGF and FGF-2 [239, Moreover, integrin α3β1and αvβ3 bind VEGF-A and 240]. MCP-1 also mediates the recruitment of mural cells in VEGF-A [112]. FGF is directly bound by integrin αvβ3 an Ang-1-dependent manner in an ex vivo model [241]. [226]. Angiopoietins also can directly interact with many Multiple sequential steps are required for angiogenesis integrins [115, 221, 227, 228]. to be successful and in all steps of this angiogenic cascade In the context of a hypoxic tumor microenvironment, it integrins, which mediate interactions of cells with surround- is especially interesting that the expression of integrins α1β1 ing insoluble ECM proteins, in addition to soluble growth and α2β1 is upregulated by VEGF [51]. factors, play an important role [15]. In a first step, the BM of an existing vessel is degraded by MMPs that are expressed by ECs, such as MMP-1, MMP-2, MMP-9, and 3. The Dynamic Process: Connection of MT1-MMP/MMP14 [242–244], at which MMP-9 is required a Tumor to the Host Vasculature for tumor vasculogenesis rather than angiogenesis [245]. Angiogenesis is an important step in the metastatic cascade, Subsequently, cell-matrix contact influences the outgrowth of tip cells and the proliferation of stalk cells that thereupon which not only provides the tumor with nutrients but also form endothelial tubes [246]. A new BM is assembled by is a route for dissemination. An important trigger for this is hypoxia [229]. newly synthesized BM proteins. Finally, the newly generated capillaries undergo maturation, pruning, and expansion. 3.1. An Angiogenic Switch Triggers the Angiogenic Cascade. In avascular tissue regions, an oxygen diffusion limit of about 3.2. Tumor Vessels Can Arise by Different Types of Vessel 150 μm restricts tumor growth to just a few millimeters in Formation. During embryonic morphogenesis, endothelial diameter. Thus, in this prevascular phase of tumor dor- precursor cells called angioblasts initiate the body’s vascu- mancy, there is a dynamic equilibrium between proliferation lature by forming tubes in a process called vasculogenesis. and hypoxia-induced apoptosis [230]. The dormant phase This is subsequently accompanied by sprouting (angio- ceases when a tumor recruits its own vasculature by the genesis) of new vessels from already existing ones. Once secretion of angiogenic factors into its environment [231], morphogenesis is completed, the adult vasculature is largely a process denoted as angiogenic switch [2, 6]. After this quiescent, except for transient events, such as wound healing angiogenic switch is thrown, the tumor hooks up to the or menorrhea [247]. However, angiogenesis takes place body’s vascular system and thus resumes its growth. under many pathological conditions, such as atherosclerosis, In tumor development, the establishment of an angio- endometriosis, osteomyelitis, diabetic retinopathy, rheuma- genic phenotype is a crucial and general step [232–234]. toid arthritis, psoriasis, and tumor growth [230]. During Depending on tumor type and environment, this induction tumor progression, the quiescent vasculature becomes per- of new vessel sprouting can occur at different stages of manently activated to sprout new vessels that enable blood the tumor progression pathway, and it leads to exponential supply and thus help sustain tumor growth [5, 6]. Due to macroscopic tumor growth [2, 4, 6]. In addition, recent data its increased metabolic rate, tumor tissue requires blood indicate that angiogenesis also contributes to the microscopic supply for expansive growth, which is circumstantiated by premalignant phase of neoplastic progression [5]. the observation that tumor cells, which are p53 deficient Infiltration of bone-marrow-derived monocytes that and thus show a reduced apoptosis rate, die beyond an differentiate into macrophages can trigger this angiogenic oxygen diffusion limit in the range of 150 μm[248]. Tumor switch in spontaneous tumors by releasing both numerous cells proliferate around the continuously formed neovessels proangiogenic cytokines, for example, VEGF, TNFα,IL-8, which markedly differ from normal vessels in morphology and bFGF [235, 236] and MMPs (e.g., MMPs-2, -7, and and molecular composition [219, 249]. Tumor vasculature -9) together with elastase and uPA [236]. These matrix- generally appears highly tortuous, chaotic, and disorganized. degrading enzymes loosen the avascular ECM for the angio- The vessels themselves are leaky due to a discontinuous genic ingrowth of neovessels. endothelium, a poorly formed BM, and a lack of mural From the multitude of proangiogenic molecules, such as cells. In addition, tumor cells sometimes mimic ECs. This FGF-1 and -2, G-CSF, HGF, IL-8, PD-ECGF, PGE-1 and -2, poor quality of tumor-associated blood vessels compromises PlGF-1, and -2, TGF-α and -β,TNF-α, and VEGF-A through blood flow, impairs drug delivery, and facilitates tumor E, only the VEGFs and PlGFs are specific for ECs [230]. cell intravasation leading to hematogenous or lymphatic VEGF-A, which exists in five splice variants, is the most metastasis. In addition to histological vessel malformations, Journal of Oncology 9 tumor vessels show an anomalous composition of their ECM, for example, tenascin-C and –W, and the oncofetal fibronectin ED-B splice variants are associated with tumor vessels [250, 251]. ED-B fibronectin is synthesized by neo- plastic cells [252]. Melanoma and glioblastoma cells secrete tenascin-C as do cancer-associated fibroblasts (CAF) of most carcinomas [253]. Tenascin-C stimulates angiogenesis in ECs, mediates survival of tumor stem cells, enhances proliferation, invasiveness, and metastasis in tumor cells, and blocks immunosurveillance [250, 253]. Tenascin-W is more strictly associated with tumorigenesis and can be used as a tumor biomarker for breast and colon cancer, because it is undetectable in healthy stroma but overexpressed in the tumor stroma [254, 255]. Vascularization mechanisms in cancer have been re- viewed recently [256, 257]. New tumor blood vessels can either arise by vessel co-option or be formed by tumor angiogenesis, but there is also evidence for vasculogenesis or recruitment of circulating bone-marrow-derived endothelial progenitor cells that differentiate into ECs [230, 258–260] (Figure 2(A)). Depending on the tumor type, tumor blood F vessels build different and characteristic vascular beds, and, according to the function of the vascular bed and Endothelial cell (EC) Tip cell the osmotic pressure of the surrounding tissue, endothelia Endothelial progenitor cell (EPC) Tumor cell represent highly heterogeneous “vascular addresses” [230]. Tumor vessels constantly change their shape due to persis- EPC-derived cell tent growth, and about 30% of the vasculature comprise Figure 2: Diverse types of vessel formation. Tumor neovasculariza- arteriovenous shunts bypassing capillaries. The concomitant tion can take place by distinct types of vessel formation, which can poor perfusion leads to hypoxia of ECs, which consequently proceed simultaneously and also merge seamlessly. (A) Neovessel synthesize more proangiogenic molecules and thus crank formation by recruitment of bone-marrow-derived endothelial tumor angiogenesis [230]. progenitor cells. (B) Sprouting angiogenesis is initiated by the differentiation of an EC into a migratory but nonproliferating tip 3.2.1. Endothelial Sprouting. Endothelial sprouting can be cell. (C) Intussusceptive angiogenesis starts with the insertion of a triggered by hypoxia, hypoglycemia, and inflammatory or connective tissue pillar into a preexisting vessel, and the vessel is dis- mechanical stimuli, such as blood pressure, and is regulated placed as the pillar extends in size. (D) In glomeruloid angiogenesis, by many angiogenic growth factors, such as VEGF, and complex vascular aggregates of several closely associated vessels are matrix proteases. When neovessels sprout from capillaries, formed. (E) Vessel co-option is the acquisition of host capillaries by the tumor. (F) In vascular mimicry, tumor cells can partly assume pericytes are selectively lost, and upon receiving an angio- EC function and form vessel-like hollow structures. Arrows denote genic stimulus, select ECs differentiate into tip cells that consecutive stages of vessel formation. Tumor tissue is depicted dark invade the avascular ECM (Figure 2(B)). These tip cells gray. See text for details. migrate into the ECM following the stimulatory gradient. Behind the tip cells, other ECs begin to proliferate and, as stalk cells, form cord-like structures. These develop cell invasion [266]. In addition to VEGF, FGF, PDGF, and into endothelial tubes [130, 261, 262] that subsequently PlDGF are involved, and Ang-2/Tie-2 signaling regulates the anastomose and thus allow blood flow. Finally, pericytes and detachment of pericytes. Later, PDGF-BB recruits pericytes smooth muscle cells are recruited, a new BM is synthesized, andsmoothmusclecells to the newlyformedECtube, and the ECs become quiescent again. and TGF-β1 and Ang-1/Tie-2 stabilize the EC-mural cell The molecular background of capillary sprouting and interaction [231]. the key role of VEGF have been reviewed by Carmeliet [231]. Upon a hypoxic stimulus, VEGF is produced, and as a consequence the endothelium’s permeability is increased 3.2.2. Intussusceptive Angiogenesis. Another way of tumor and the BM loosened by the activity of MMPs [243, 263] neovascularization is intussusceptive angiogenesis, which and the urokinase plasminogen activator system [264]. The represents a nonproliferative and noninvasive mechanism MMP inducer EMMPRIN/CD147 also upregulates soluble for the enlargement of a capillary plexus by intussusceptive VEGF isoforms 121 and 165 and VEGFR-2 on ECs and growth, arborization, and remodeling [267](Figure 2(C)). thus promotes sprouting angiogenesis [265]. Integrin αvβ3 As this mode of vascularization is mostly independent from mediates migration into the fibrin-rich cancer stroma and EC proliferation and migration, as well as BM degradation, furthermore can associate with MMP-2, thus enabling ECs this process is more economical and, occurring within to maintain the BM in the sol state and to promote tumor hours or even minutes, is noticeably faster than sprouting 10 Journal of Oncology angiogenesis [268]. It begins with the formation of translu- of angiogenesis, and they can arise by two types of vascu- minal pillars from the EC walls. Their subsequent expansion logenic mimicry, designated the tubular and the patterned splits the preexisting vessel into two, thereby enhancing the matrix type [277]. These tubular vessel-like networks resem- vascular surface. In a subsequent process of arborization, ble the pattern of embryonic vascular networks, and, in the disorganized capillary network is remodeled into a their gene expression pattern, aggressive tumors that form functional tree-like structure by serial pillar formation. In such channels resemble endothelial, pericytes, and other a final remodeling step, the branching angles are modified, precursor stem cells, suggesting that tumor cells might and the capillary network is pruned. The formation of new disguise as embryonic stem-cell-like or other cell types capillaries is initiated by sprouting angiogenesis that is later [256]. Vasculogenic mimicry of the patterned matrix type accompanied or followed by intussusceptive angiogenesis, looks completely different and is characterized by a fluid- which increases the EC surface [269]. Intussusceptive angio- conducting meshwork of extravascular patterned depositions genesis is synergistically regulated by VEGF and Ang-1, of matrix proteins such as laminins, collagens IV and VI, and and it seems to be induced by laminar shear stress on the heparin sulfate proteoglycans that anastomose with blood vessel walls, whereas oscillating shear stress favors sprouting vessels [277–279]. Although it is not yet elucidated how such angiogenesis [269]. channels are connected to the vasculature, the latter type of vascular mimicry has been reported for many cancers, such as breast, ovarian, and prostate carcinoma, melanoma, 3.2.3. Glomeruloid Angiogenesis. In many aggressive tumors, soft tissue sarcomas, osteosarcoma, and phaeochromocy- glomeruloid angiogenesis gives rise to complex vascular toma [277, 280]. In aggressive melanoma, the expression structures termed glomeruloid bodies, in which several of tissue factor pathway-associated genes, such as tissue microvessels together are ensheathed by a BM of varying factor (TF), TF pathway inhibitor-1 (TFPI-1), and TFPI- thickness containing sparse pericytes [270](Figure 2(D)). 2, is upregulated, suggesting an anticoagulation mechanism The frequency of occurrence of such glomeruloid bodies in the channel-forming tumor cells [281]. Fluid propelled is an indication for the tumor’s aggressiveness and the through these channels by a pressure gradient might facilitate patient’s survival [271]. The formation of such glomeruloid the supply with nutrients and oxygen, and, additionally, this bodies is rather a remodeling than true angiogenesis, because fluid-conducting network could substitute for a lymphatic proliferating and migrating tumor cells can actively pull vascular system and drain extravasated interstitial fluid capillaries of the surrounding host vasculature and adjacent in tumors that lack lymphatic vessels, for example, uveal capillary branching points into the tumor node. Thereby, melanoma [279, 280]. formed coiled vascular structures develop subsequently into glomeruloid bodies that are connected to the surrounding vasculature via numerous narrowed capillaries [256]. 4. Manipulation of Cell Matrix Interaction in Tumor Angiogenesis 3.2.4. Vessel Co-Option. Malignant cells can initially grow Cell-matrix interactions regulate signaling pathways that are in the vicinity and along pre-existing microvessels and thus intricately interconnected with cytokine-regulated pathways, use the host vasculature for their own benefit (Figure 2(E)). which complicates the analysis of their contribution to a This co-option of the host vasculature was originally believed particular step in angiogenesis [153]. ECM receptors can to be limited to the initial phase of tumorigenesis [272]. be manipulated with a wide variety of different compounds Meanwhile, however, there is evidence that vessel co-option ranging from endogenous compounds, such as matrikines, might persist during all stages of primary and metastatic over their synthetic analogues and peptides mimicking only growth of various tumors [256], for example, cutaneous integrin-binding sites to function-blocking antibodies and melanoma, which appears to grow by co-opting the vascular small molecules with integrin inhibitory function. Other plexus in its surrounding connective tissue, while there is no starting points for an antiangiogenic therapy are the inhibi- sign of directed vessel ingrowth [273]. tion of signaling cascades downstream of the ECM receptors Vessel co-option is regulated dependent on the tumor or cytokine receptors and as a new avenue the blocking type and the host environment, but the key regulators are of microRNAs with antisense RNAs in ECs [282, 283]. An again VEGF and angiopoietins [272, 274]. Ang-1 binds to efficient antivascular cancer therapy can target either the Tie-2 and thus triggers signaling cascades, assuring survival angiogenic signaling pathways or the vascularization mech- and quiescence of ECs, and thus causing tumor vessel anism [256]. A combination of conventional chemotherapy maintenance, whereas the nonsignaling Tie-2 ligand Ang-2 with angiosuppressive or vascular disrupting therapy is often acts as a negative regulator and destabilizes the capillary walls problematic and needs careful design [256]. by detachment of pericytes [272, 274]. Subsequently, VEGF via its receptor VEGFR-2 promotes both survival of ECs and growth of new vessels [237, 275]. 4.1. Pharmacological Intervention of Integrin-ECM Interac- tion. In addition to soluble growth factors, such as VEGF, 3.2.5. Vascular Mimicry. Aggressive melanomas can form there are several endogenous angiogenesis inhibitors, for fluid-filled vessel-like channels without any EC lining in example, endostatin, endorepellin, and tumstatin, which a nonangiogenic process termed vascular mimicry [276] share the common feature that they all are proteolytic frag- (Figure 2(F)). These channels allow perfusion independent ments of ECM molecules [284, 285]. In tumor angiogenesis Journal of Oncology 11 within a primary tumor, such ECM fragments are generated In a phase I clinical trial, endostatin, the C-terminal by the release of MMPs, in order to degrade the BM. This fragment of collagen XVIII, blocks the function of integrin results not only in labile and leaky tumor vessels but at α5β1[179, 304, 305] and also binds to heparin and with the same time keeps metastases from growing, as these lower affinities to other heparan sulfate proteoglycans that endogenous angiogenesis inhibitors are distributed via the are involved in growth factor signaling [306, 307]. Endo- blood stream [230]. Therefore, they are of pharmacological statin’s antiangiogenic activity can also be mimicked with interest with regard to their use as angiogenesis inhibitors. derived short non-RGD but arginine-rich peptides [308]. Intensive efforts have been directed towards the development Integrin α5β1 can also be blocked by the synthetic non- of integrin antagonists for the treatment of cancer and many RGD peptides PHSCN, named ATN-161, [309] and cyclic other diseases, ranging from autoimmune diseases over CRRETAWAC [310], as well as by the peptide mimetics SJ749 inflammatory to thrombotic diseases, and their applications [311] and JSM6427 [312], and it can be inhibited by the seem promising [11, 286]. Integrin-mediated interactions affinity-matured humanized chimeric monoclonal antibody of cells with their surrounding ECM can be manipulated M200/volociximab [313]. by antibodies, peptides, small nonpeptidic compounds, and Angiostatin is a proteolytic fragment of plasminogen that endogenous inhibitors (Figure 3). Integrin antagonists with effectively inhibits integrin αvβ3[314], and its antiangio- antiangiogenic activities have been reviewed recently with genic effect can also be achieved by its isolated kringle-5 special emphasis on drugs that are in clinical trials [11]. domain [315]. Kringle-1 to 3 show the same antiproliferative Spurred by the success in pharmacologically targeting effect as the whole angiostatin, but hardly inhibit migration, RGD-dependent integrins, there are also attempts to phar- whereas kringle-4 inhibits EC migration but shows only a macologically manipulate RGD-independent integrins, such marginal antiproliferative effect [316]. Other endogenous as the collagen- and laminin-binding integrins, as reviewed integrin αvβ3 inhibitors are the collagen XVIII fragment recently [287]. The collagen-binding subgroup of integrins endostatin [304], and the C-terminal fragment of the with their common A domain comprises interesting targets collagen IV α3-chain termed tumstatin [317], which also in the development of drugs against thrombosis, inflam- binds to integrin α6β1[318]. Tumstatin has two binding matory diseases, and cancer. TSPs-1 and -2 are naturally sites for integrin αvβ3. The N-terminal site mediates an occurring potent angiogenesis inhibitors, and their anti- antiangiogenic signal, whereas the C-terminal binding site angiogenic effects can be imitated by short-peptide mimetics is associated with the antitumor cellactivity [318, 319]. that among other targets bind to β1 integrins [288, 289]. Canstatin, the NC1 domain of the collagen IV α2 chain, An endogenous inhibitor, which blocks the interaction inhibits both integrins αvβ3and αvβ5[320] and seems of integrin α1β1 with collagen I and also binds to heparan to interact with integrin α3β1too [321]. A hemopexin- sulfate proteoglycans, is arresten, the C-terminal fragment like domain comprising C-terminal fragment of MMP-2, of the collagen IV α1 chain [290, 291]. Endorepellin, a termed PEX, also antagonizes integrin αvβ3 by preventing C-terminal fragment of perlecan specifically blocks the its binding to MMP-2 and thus inhibiting proteolytic activity function of integrin α2β1[292] and interestingly also on the cell surface, especially during vessel maturation [322, binds to endostatin, thus counteracting its antiangiogenic 323]. Fastatin and other FAS1 domains, which are present effect [293]. Additionally, integrin α1β1 can be specifically in the four human proteins periostin, FEEL1, FEEL2, and inhibited with obtustatin from the snake venom of Vipera βhig-h3, also function via integrin αvβ3asendogenous lebetina obtusa [294, 295]. The interaction of integrin α2β1 regulators of pathogenic angiogenesis [324]. Next to these with collagen can be specifically inhibited with the C- natural antagonists there is a variety of synthetic RGD— type lectin rhodocetin from the snake venom of Callose- containing peptide inhibitors that mimic a motif that occurs lasma rhodostoma [296, 297]. In addition, it can also be on many ECM molecules, such as fibronectin, vitronectin, selectively antagonized by the protein angiocidin, which fibrinogen, osteopontin, TSP, vWF, and partially degraded was first detected in lung carcinoma cells [298, 299]. The collagen. Most integrins of the αv subfamily and the integrins aromatic tetracyclic polyketides maggiemycin and anhydro- α5β1and αIIbβ3 bind to this motif. Therefore, adhesion and maggiemycin from Streptomyces, which have been described spreading of ECs to the ECM can be competitively inhib- as potential antitumor agents [300], inhibit collagen binding ited by RGD peptides, whereby anchorage-dependent ECs by blocking the A domain of the integrin subunits α1, undergo apoptosis [230]. To this group belong compounds, α2, α11, and to a lesser extent α10 while cell adhesion such as cilengitide/EMD121974 [325], S137 and S247 [326, to fibronectin, mediated by integrins α5β1, αvβ3, αvβ5, 327], the TSP-derived peptide TP508/chrysalin [328], and is unaffected [301]. Recently, the sulfonamide derivative several integrin αvβ3- and αvβ5-specific peptidomimetics, BTT-3016 has been described as a potent antithrombotic such as BCH-14661, which preferentially inhibits αvβ3and small-molecule inhibitor of integrin α2β1 with only slight BCH-15046, which blocks αvβ3, αvβ5, and α5β1[329], effect on other collagen-binding integrins and no effect on SCH221153 [330], and ST1646 [331]. Another inhibitor fibronectin- or vitronectin-binding integrins [302]. Another is the non-peptide antibiotic thiolutin, which intracellu- sulfonamide derivative, E7820, which does not interfere with larly blocks paxillin and thus, indirectly, integrin αvβ3- integrin-ligand interaction, reduces integrin α2 expression mediated adhesion to vitronectin [332]. Antibodies against on the mRNA level [303]. Angiogenesis can be inhibited with the β3 subunit inhibit contact of ECs to vitronectin and antibodies against the α subunits of the integrins α1β1and concomitantly VEGF-induced tyrosine phosphorylation of α2β1, whereas quiescent vessels are not affected [230]. VEGFR-2 in cell culture studies [333]. Moreover, integrin 12 Journal of Oncology ECM Matrikines Peptides Antibodies Small molecule inhibitors α β Adhesion Survival Migration Proliferation Figure 3: Options to manipulate integrin function. Essential cellular functions, such as adhesion, migration, proliferation, and survival, which all are regulated by integrins, can pharmacologically be manipulated with a panoply of matrikines, antibodies, peptides, and small molecule inhibitors, many of which are used as therapeutic tools in combination with conventional chemo- or radiotherapy to attack tumor cells and vasculature. Details are described in the text. αvβ3can be effectively antagonized with the monoclonal the RGD-containing inhibitors cilengitide and S 36578 alter antibody LM609/MEDI-552 and its humanized derivative the trafficking of integrins and VEGFR2 in tumor ECs, thus abegrin/etaracizumab/vitaxin [57, 334–337]. In contrast, stimulating angiogenesis and tumor growth [342]. the humanized anti-αv antibody CNTO95 targets both Current tumor therapy aims at vessel eradication in order integrins αvβ3and αvβ5[338]. The humanized Fab fragment to disrupt the connection of the tumor to the vascular 17E6/abciximab/ReoPro of the monoclonal antibody c7E3 system and thus cut off the supply of nutrients and oxygen. inhibits the integrins αvβ3 and also αMβ2/Mac-1 [339, This can be done with compounds that preferentially affect 340], whereas the human-specific monoclonal antibody tumor endothelia rather than normal cells, that is, (i) specific 17E6 targets all αv integrins [341]. Currently, humanized or angiogenesis inhibitors, (ii) tumor vessel toxins that attack chimeric integrin antibody antagonists of αvβ3, αvβ5, and inherent weaknesses in static tumor vessel endothelia and α5β1, and peptide inhibitors of these integrins are in clinical associated vascular structures, and (iii) dual-action com- trials as antiangiogenic agents [180]. pounds [343]. However, within the last years, a paradigm shift has taken place [344, 345]. Vessel normalization by pruning immature vessels and increasing pericytes and 5. Applications and Outlook BM coverage of the remaining vessels comes to the fore, Integrins and their binding partners are of special interest rather than vessel eradication, because mere antiangiogenic treatment can worsen malignancy [346]. A malformed as potential therapeutic targets, and several are already in clinical trials. However, the results fall short of the initial tumor vasculature creates and aggravates a hypoxic and expectations, pointing out that monotherapy with a single acidic milieu which hampers drug delivery and perfusion [347–349], and, due to its leaky endothelium, it promotes angiogenesis inhibitor is not sufficient to counteract the numerous angiogenic factors involved in tumor progres- tumor cell dissemination [346]. Therefore, chemotherapeu- sion [231]. Moreover, there are some caveats in aiming tic efficacy can be ameliorated by a concomitant vessel normalization therapy which improves delivery and efficacy at integrins as therapeutic targets. Obviously, integrins are expressed on virtually all cells under physiological as of cytotoxic drugs and also sensitizes the tumor cells to radiation [345, 350]. well as pathological conditions, and it is a major chal- lenge to target exclusively integrins on tumor or tumor- In vessel normalization, the interaction of cells with associated cells. Another problem is that low concentrations their surrounding ECM via integrins is of special impor- tance. However, many antiangiogenic compounds, for exam- of antagonists alter the signaling of integrins and other receptors. When administered in nanomolar concentrations, ple, ATN-161, endostatin, and integrin inhibitors, show Journal of Oncology 13 hormetic, that is, bell- or U-shaped, dose-response curves Additionally, integrins can be used as biomarkers to non- and thus present a challenge for clinical translation [351]. invasively assess the efficacy of chemotherapeutic and radio- Nanomolar concentrations of RGD-mimetic αvβ3and αvβ5 therapeutic drugs [12]. Integrin-targeted probes can be inhibitors (S 36578 and cilengitide) can paradoxically used to visualize tumor angiogenesis and the response to stimulate tumor growth and angiogenesis by altering the chemo- and radiotherapy by various imaging methods, trafficking of αvβ3 integrin and VEGFR2. Thus, they such as magnetic resonance imaging (MRI), positron emis- promote the migration of ECs towards VEGF, which has sion tomography (PET), and ultrasonography [360–362]. important implications for the use of RGD mimetics in Moreover, fluorescence labeling of integrin ligands allows tumor therapy [342]. Thus, depending on tumor type, dose, intraoperative fluorescence imaging, thus providing a tool and manner of application, the currently available-integrin to intraoperatively detect and remove metastases of sub- targeting compounds can act either anti- or proangiogenic. millimeter size [363]. A promising approach may be a combination therapy that In summary, the above data illustrate the importance blocks simultaneously angiogenic integrin αvβ3 and VEGFR of integrins and integrin-binding and signaling proteins in activities [352–355]. both physiological and pathological blood vessel formation. To circumvent these problems, instead of targeting the Thus, they may be potential targets for antiangiogenic tumor integrins, which are in principle present on both normal and therapy. Although our knowledge concerning this matter has malignant cells, another strategy aims at tumor-promoting increased remarkably within the last years, the understanding integrin ligands, such as ED-B fibronectin, tenascin-C, and is far from complete. tenascin W [252, 253, 255]. Invasive tumor cells partially degrade and denature their surrounding ECM, and the thereby released cryptic collagen IV epitope HU177 may also Abbreviations be a potential target for antiangiogenic and tumor-selective drug delivery [356]. Ang: Angiopoietin In comparison to a systemic administration of a chemo- BM: Basement membrane therapeutic agent, its therapeutic index can be increased CRDGK, Amino acid sequences in single letter by selectively targeting integrins that are overexpressed on CRGDKGPDC, code tumor cells [357]. Chemotherapeutic small molecules, pep- CRRETAWAC, tides, and proteins as well as nanoparticle-carried chemo- PHSCN, RGD: therapeutics, which are conjugated to ligands of integrins ECM: Extracellular matrix that are overexpressed on angiogenic ECs or tumor cells, can EC: Endothelial cell be selectively internalized after integrin binding [357]. Espe- FAK: Focal adhesion kinase cially nanoparticles, such as micelles, liposomes, polymeric FGF: Fibroblast growth factor nanospheres, and polymersomes loaded with chemothera- G-CSF: Granulocyte colony-stimulating peutic or radiotherapeutic drugs and equipped with multi- factor valent integrin ligands show decreased systemic toxicity, pro- HGF: Hepatocyte growth factor longed half-life and passive retention in the tumor, improved HIF: Hypoxia-inducible factor binding affinity, and facilitated internalization, thus resulting IL: Interleukin in increased drug delivery [12, 357, 358]. A therapeutic MAPK: Mitogen-activated protein kinase strategy that targets several integrins and receptors by such MEK: MAPK/ERK kinase chemo-, radio-, and possibly gene therapeutic approaches MMP: Matrix metalloproteinase maybemoreeffective than a monotherapy [231, 357]. NC: Noncollagenous Coadministration of the αv integrin-targeting cyclic pep- NF-κB: Nuclear factor κ-light-chain tide iRGD (CRGDKGPDC), or structurally closely related enhancer of activated B cells peptides, with anticancer drugs considerably enhances their NRP-1: Neuropilin-1 efficacy and selectivity [359]. Upon binding to αv integrin- PDGF: Platelet-derived growth factor expressing tumor ECs, iRGD is proteolytically processed to PD-ECGF: Platelet-derived endothelial cell CRDGK with a much weaker integrin affinity, whereas this growth factor truncated peptide shows an increased affinity to neuropilin- PGE: Prostaglandin E 1 (NRP-1), thus increasing vascular and tissue perme- PI3K: Phosphatidylinositol-3 kinase ability in a tumor-specific and NRP-1-dependent manner PLC: Phospholipase C [359]. Interestingly, this coadministration does not require PlGF: Placenta-derived growth factor chemical conjugation of the drug with the iRGD peptide; Ras: Rat sarcoma protein that is, approved drugs could be used unmodified [359]. Src: Sarcoma oncogene Coadministration of such a tumor-penetrating peptide with TGF: Transforming growth factor either small molecules, such as doxorubicin, antibodies, such Tie: Tyrosine kinase with as trastuzumab, or nanoparticles, such as Nab-paclitaxel immunoglobulin-like and EGF-like (abraxane) or doxorubicin-loaded liposomes, resulted in domain equivalent or increased delivery and efficacy, and it improved TIMP: Tissue inhibitor of their therapeutic index by lowering the effective dose [359]. metalloproteinases 14 Journal of Oncology [15] J. A. Eble and S. Niland, “The extracellular matrix of blood vessels,” Current Pharmaceutical Design, vol. 15, no. 12, pp. TNF: Tumor necrosis factor 1385–1400, 2009. uPA(R): Urokinase-type plasminogen activator [16] V. S. LeBleu, B. MacDonald, and R. Kalluri, “Structure and (receptor) function of basement membranes,” Experimental Biology and VCAM: Vascular cell adhesion molecule Medicine, vol. 232, no. 9, pp. 1121–1129, 2007. VEGF(R): Vascular endothelial growth factor [17] R. V. Iozzo, J. J. Zoeller, and A. Nystrom, ¨ “Basement mem- (receptor) brane proteoglycans: modulators Par excellence of cancer VSMC: Vascular smooth muscle cell growth and angiogenesis,” Molecules and Cells, vol. 27, no. 5, vWF: Von Willebrand factor pp. 503–513, 2009. TSP: Thrombospondin. [18] P. D. Yurchenco, “Basement membranes: cell scaffoldings and signaling platforms,” Cold Spring Harbor Perspectives in Biology, vol. 3, no. 2, 2011. Acknowledgments [19] S. Astrof and R. O. Hynes, “Fibronectins in vascular morpho- genesis,” Angiogenesis, vol. 12, no. 2, pp. 165–175, 2009. The authors thank the Deutsche Forschungsgemeinschaft [20] S. Katsuda and T. Kaji, “Atherosclerosis and extracellular for financial support (SFB/TR23 A8 Eble). They sincerely matrix,” Journal of Atherosclerosis and Thrombosis, vol. 10, no. apologize to authors of important work not cited here for 5, pp. 267–274, 2003. reasons of space limitation. [21] G. A. M. Plenz, M. C. Deng,H.Robenek,and W. Volk ¨ er, “Vascular collagens: spotlight on the role of type VIII References collagen in atherogenesis,” Atherosclerosis, vol. 166, no. 1, pp. 1–11, 2003. [1] K. Kinzler and B. Vogelstein, The Genetic Basis of Human [22] K. Kuhn, ¨ “Basement membrane (type IV) collagen,” Matrix Cancer, McGraw-Hill, Medical Pub. Division, 2nd edition, Biology, vol. 14, no. 6, pp. 439–445, 1995. [23] E. Posc ¨ hl, U. Schlotz ¨ er-Schrehardt, B. Brachvogel, K. Saito, [2] G. Bergers and L. E. Benjamin, “Tumorigenesis and the Y. Ninomiya, and U. Mayer, “Collagen IV is essential for angiogenic switch,” Nature Reviews Cancer,vol. 3, no.6,pp. basement membrane stability but dispensable for initiation 401–410, 2003. of its assembly during early development,” Development, vol. [3] S. Rheingold, A. Neugut, and A. Meadows, “Secondary can- 131, no. 7, pp. 1619–1628, 2004. cers: incidence, risk factors, and management,” in Holland- [24] S. M. Mithieux and A. S. Weiss, “Elastin,” Advances in Protein Frei Cancer Medicine, D. Kufe, R. Pollock, and R. Weichsel- Chemistry, vol. 70, pp. 437–461, 2005. baum, Eds., p. 2399, B. C. Decker, Hamilton, Ont, Canada, [25] C. M. Kielty, “Elastic fibres in health and disease,” Expert Reviews in Molecular Medicine, vol. 8, no. 19, pp. 1–23, 2006. [4] D. Hanahan and R. A. Weinberg, “The hallmarks of cancer,” [26] A. Patel, B. Fine, M. Sandig, and K. Mequanint, “Elastin Cell, vol. 100, no. 1, pp. 57–70, 2000. biosynthesis: the missing link in tissue-engineered blood [5] D. Hanahan and R. A. Weinberg, “Hallmarks of cancer: the vessels,” Cardiovascular Research, vol. 71, no. 1, pp. 40–49, next generation,” Cell, vol. 144, no. 5, pp. 646–674, 2011. [6] D. Hanahan and J. Folkman, “Patterns and emerging mech- [27] A. Colombatti, R. Doliana, S. Bot et al., “The EMILIN protein anisms of the angiogenic switch during tumorigenesis,” Cell, family,” Matrix Biology, vol. 19, no. 4, pp. 289–301, 2000. vol. 86, no. 3, pp. 353–364, 1996. [28] B. S. Brooke, S. K. Karnik, and D. Y. Li, “Extracellular [7] B. Nico, E. Crivellato, D. Guidolin et al., “Intussusceptive matrix in vascular morphogenesis and disease: structure microvascular growth in human glioma,” Clinical and Exper- versus signal,” Trends in Cell Biology, vol. 13, no. 1, pp. 51– imental Medicine, vol. 10, no. 2, pp. 93–98, 2010. 56, 2003. [8] C. J. Avraamides, B. Garmy-Susini, and J. A. Varner, “Inte- [29] R. Timpl, T. Sasaki, G. Kostka, and M. L. Chu, “Fibulins: grins in angiogenesis and lymphangiogenesis,” Nature Re- a versatile family of extracellular matrix proteins,” Nature views Cancer, vol. 8, no. 8, pp. 604–617, 2008. Reviews Molecular Cell Biology, vol. 4, no. 6, pp. 479–489, [9] R. Rathinam and S. K. Alahari, “Important role of integrins in the cancer biology,” Cancer and Metastasis Reviews, vol. 29, [30] T. Nakamura, P. R. Lozano, Y. Ikeda et al., “Fibulin-5/DANCE no. 1, pp. 223–237, 2010. is essential for elastogenesis in vivo,” Nature, vol. 415, no. [10] G. Alghisi and C. Ruegg ¨ , “Vascular integrins in tumor angio- 6868, pp. 171–175, 2002. genesis: mediators and therapeutic targets,” Endothelium, vol. [31] M. Hirai, T. Ohbayashi, M. Horiguchi et al., “Fibulin- 13, no. 2, pp. 113–135, 2006. 5/DANCE has an elastogenic organizer activity that is [11] C. Ruegg ¨ and G. C. Alghisi, “Vascular integrins: therapeutic abrogated by proteolytic cleavage in vivo,” Journal of Cell and imaging targets of tumor angiogenesis,” Recent Results in Biology, vol. 176, no. 7, pp. 1061–1071, 2007. Cancer Research, vol. 180, pp. 83–101, 2010. [12] J. S. Desgrosellier and D. A. Cheresh, “Integrins in can- [32] R. Giltay, R. Timpl, and G. Kostka, “Sequence, recombinant expression and tissue localization of two novel extracellular cer: biological implications and therapeutic opportunities,” Nature Reviews Cancer, vol. 10, no. 1, pp. 9–22, 2010. matrix proteins, fibulin-3 and fibulin-4,” Matrix Biology, vol. 18, no. 5, pp. 469–480, 1999. [13] S.-H. Kim, J. Turnbull, and S. Guimond, “Extracellular matrix and cell signalling: the dynamic cooperation of [33] K. P. Dingemans, P. Teeling, J. H. Lagendijk, and A. E. integrin, proteoglycan and growth factor receptor,” Journal Becker, “Extracellular matrix of the human aortic media: of Endocrinology, vol. 209, no. 2, pp. 139–151, 2011. an ultrastructural histochemical and immunohistochemical [14] L. Gartner and J. Hiat, Color Atlas of Histology,Williams& study of the adult aortic media,” Anatomical Record, vol. 258, Wilkins, Baltimore, Md, USA, 1994. no. 1, pp. 1–14, 2000. Journal of Oncology 15 [34] A. G. Marneros and B. R. Olsen, “Physiological role of sarcomas,” American Journal of Pathology, vol. 142, no. 4, pp. collagen XVIII and endostatin,” FASEB Journal, vol. 19, no. 1009–1018, 1993. 7, pp. 716–728, 2005. [51] D. R. Senger, K. P. Claffey,J.E.Benes,C.A.Perruzzi, ¨ ¨ ¨ A. P. Sergiou, and M. Detmar, “Angiogenesis promoted by [35] S. Grassel, C. Unsold,H.Schacke, L. Bruckner-Tuderman, vascular endothelial growth factor: regulation through α1β1 and P. Bruckner, “Collagen XVI is expressed by human dermal fibroblasts and keratinocytes and is associated with and α2β1 integrins,” Proceedings of the National Academy of Sciences of the United States of America, vol. 94, no. 25, pp. the microfibrillar apparatus in the upper papillary dermis,” Matrix Biology, vol. 18, no. 3, pp. 309–317, 1999. 13612–13617, 1997. [52] D. R. Senger, C. A. Perruzzi, M. Streit, V. E. Koteliansky, A. [36] A. Kassner, U. Hansen, N. Miosge et al., “Discrete integration R. De Fougerolles, and M. Detmar, “The α1β1and α2β1 of collagen XVI into tissue-specific collagen fibrils or beaded integrins provide critical support for vascular endothelial microfibrils,” Matrix Biology, vol. 22, no. 2, pp. 131–143, growth factor signaling, endothelial cell migration, and tumor angiogenesis,” American Journal of Pathology, vol. 160, [37] E. P. Moiseeva, “Adhesion receptors of vascular smooth no. 1, pp. 195–204, 2002. muscle cells and their functions,” Cardiovascular Research, [53] K. Vuoriluoto, G. Hog ¨ nas, ¨ P. Meller, K. Lehti, and J. Ivaska, vol. 52, no. 3, pp. 372–386, 2001. “Syndecan-1 and -4 differentially regulate oncogenic K-ras [38] J. A. Eble, A. Kassner, S. Niland, M. Morgelin, ¨ J. Grifka, and S. dependent cell invasion into collagen through α2β1 integrin Grassel, ¨ “Collagen XVI harbors an integrin α1β1 recognition and MT1-MMP,” Matrix Biology, vol. 30, no. 3, pp. 207–217, site in its C-terminal domains,” Journal of Biological Chem- istry, vol. 281, no. 35, pp. 25745–25756, 2006. [54] R. O. Hynes, “Integrins: bidirectional, allosteric signaling [39] J. Thyberg, K. Blomgren, J. Roy, P. K. Tran, and U. Hedin, machines,” Cell, vol. 110, no. 6, pp. 673–687, 2002. “Phenotypic modulation of smooth muscle cells after arterial [55] J. T. Yang, H. Rayburn, and R. O. Hynes, “Cell adhesion injury is associated with changes in the distribution of events mediated by α4 integrins are essential in placental and laminin and fibronectin,” Journal of Histochemistry and cardiac development,” Development, vol. 121, no. 2, pp. 549– Cytochemistry, vol. 45, no. 6, pp. 837–846, 1997. 560, 1995. [40] A. P. Hall, “Review of the pericyte during angiogenesis [56] M. R. Morgan, M. J. Humphries, and M. D. Bass, “Synergistic and its role in cancer and diabetic retinopathy,” Toxicologic control of cell adhesion by integrins and syndecans,” Nature Pathology, vol. 34, no. 6, pp. 763–775, 2006. Reviews Molecular Cell Biology, vol. 8, no. 12, pp. 957–969, [41] R. H. Adams and K. Alitalo, “Molecular regulation of angio- genesis and lymphangiogenesis,” Nature Reviews Molecular [57] P. C. Brooks,R.A.F.Clark,and D. A. Cheresh, “Requirement Cell Biology, vol. 8, no. 6, pp. 464–478, 2007. of vascular integrin α(v)β3 for angiogenesis,” Science, vol. [42] H. Gerhardt and H. Semb, “Pericytes: gatekeepers in tumour 264, no. 5158, pp. 569–571, 1994. cell metastasis?” Journal of Molecular Medicine, vol. 86, no. 2, [58] R. O. Hynes, “Cell-matrix adhesion in vascular develop- pp. 135–144, 2008. ment,” Journal of Thrombosis and Haemostasis,vol. 5, no.1, [43] Z. Isogai, A. Aspberg, D. R. Keene, R. N. Ono, D. P. Reinhardt, pp. 32–40, 2007. and L. Y. Sakai, “Versican interacts with fibrillin-1 and [59] C. N. Landen, T. J. Kim, Y. G. Lin et al., “Tumor-selective links extracellular microfibrils to other connective tissue response to antibody-mediated targeting of αvβ3 integrin in networks,” Journal of Biological Chemistry, vol. 277, no. 6, pp. ovarian cancer,” Neoplasia, vol. 10, no. 11, pp. 1259–1267, 4565–4572, 2002. [44] A. Aspberg, S. Adam, G. Kostka, R. Timpl, and D. Heinegar ˚ d, [60] G. E. Davis, “Affinity of integrins for damaged extracellular “Fibulin-1 is a ligand for the C-type lectin domains of matrix: α(v)β3 binds to denatured collagen type I through aggrecan and versican,” Journal of Biological Chemistry, vol. RGD sites,” Biochemical and Biophysical Research Communi- 274, no. 29, pp. 20444–20449, 1999. cations, vol. 182, no. 3, pp. 1025–1031, 1992. [45] A. I. Olin, M. Morgelin, ¨ T. Sasaki, R. Timpl, D. Heinegar ˚ d, [61] B. Nieswandt, M. Hafner, B. Echtenacher, and D. N. Mannel, ¨ and A. Aspberg, “The proteoglycans aggrecan and versican “Lysis of tumor cells by natural killer cells in mice is impeded form networks with fibulin-2 through their lectin domain by platelets,” Cancer Research, vol. 59, no. 6, pp. 1295–1300, binding,” Journal of Biological Chemistry, vol. 276, no. 2, pp. 1253–1261, 2001. [62] P. C. Brooks,S.Stromblad,R.Klemke, D. Visscher, F. H. [46] J. Heino and J. Kap ¨ yla, ¨ “Cellular receptors of extracellular Sarkar, and D. A. Cheresh, “Antiintegrin αvβ3blockshuman matrix molecules,” Current Pharmaceutical Design, vol. 15, breast cancer growth and angiogenesis in human skin,” no. 12, pp. 1309–1317, 2009. Journal of Clinical Investigation, vol. 96, no. 4, pp. 1815–1822, [47] L. Contois, A. Akalu, and P. C. Brooks, “Integrins as “func- tional hubs” in the regulation of pathological angiogenesis,” [63] G. H. Mahabeleshwar, W. Feng, D. R. Phillips, and T. Seminars in Cancer Biology, vol. 19, no. 5, pp. 318–328, 2009. V. Byzova, “Integrin signaling is critical for pathological [48] R. Zaidel-Bar, S. Itzkovitz, A. Ma’ayan, R. Iyengar, and B. angiogenesis,” Journal of Experimental Medicine, vol. 203, no. Geiger, “Functional atlas of the integrin adhesome,” Nature 11, pp. 2495–2507, 2006. Cell Biology, vol. 9, no. 8, pp. 858–867, 2007. [64] X. Huang, M. Griffiths, J. Wu, R. V. Farese, and D. Sheppard, [49] R. Silva, G. D’Amico, K. M. Hodivala-Dilke, and L. E. “Normal development, wound healing, and adenovirus Reynolds, “Integrins: the keys to unlocking angiogenesis,” susceptibility in β5- deficient mice,” Molecular and Cellular Arteriosclerosis, Thrombosis, and Vascular Biology, vol. 28, no. Biology, vol. 20, no. 3, pp. 755–759, 2000. 10, pp. 1703–1713, 2008. [65] B. L. Bader, H. Rayburn, D. Crowley, and R. O. Hynes, “Ex- [50] M. Miettinen, R. Castello, E. Wayner, and R. Schwarting, tensive vasculogenesis, angiogenesis, and organogenesis pre- “Distribution of VLA integrins in solid tumors: emergence cede lethality in mice lacking all αv integrins,” Cell, vol. 95, of tumor-type- related expression patterns in carcinomas and no. 4, pp. 507–519, 1998. 16 Journal of Oncology [66] J. H. McCarty, R. A. Monahan-Earley, L. F. Brown et al., of collagenase (MMP-1) and collagen α1(I) gene expression,” “Defective associations between blood vessels and brain Journal of Biological Chemistry, vol. 270, no. 22, pp. 13548– parenchyma lead to cerebral hemorrhage in mice lacking αv 13552, 1995. integrins,” Molecular and Cellular Biology, vol. 22, no. 21, pp. [82] O. Langholz, D. Roc ¨ kel, C. Mauch et al., “Collagen and colla- 7667–7677, 2002. genase gene expression in three-dimensional collagen lattices [67] J. H. McCarty, A. Lacy-Hulbert, A. Charest et al., “Selective are differentially regulated by α1β1and α2β1 integrins,” ablation of αv integrins in the central nervous system leads Journal of Cell Biology, vol. 131, no. 6, pp. 1903–1915, 1995. to cerebral hemorrhage, seizures, axonal degeneration and [83] H. Gardner, A. Broberg, A. Pozzi, M. Laato, and J. Heino, premature death,” Development, vol. 132, no. 1, pp. 165–176, “Absence of integrin α1β1 in the mouse causes loss of 2005. feedback regulation of collagen synthesis in normal and [68] L. E. Reynolds, L. Wyder, J. C. Lively et al., “Enhanced patho- wounded dermis,” Journal of Cell Science, vol. 112, no. 3, pp. logical angiogenesis in mice lacking β3 integrin or β3and β5 263–272, 1999. integrins,” Nature Medicine, vol. 8, no. 1, pp. 27–34, 2002. [84] F. Alves, W. Vogel, K. Mossie, B. Millauer, H. Hofler, and [69] F. D´ıaz-Gonzalez, ´ J. Forsyth, B. Steiner, and M. H. Ginsberg, A. Ullrich, “Distinct structural characteristics of discoidin “Trans-dominant inhibition of integrin function,” Molecular I subfamily receptor tyrosine kinases and complementary Biology of the Cell, vol. 7, no. 12, pp. 1939–1951, 1996. expression in human cancer,” Oncogene,vol. 10, no.3,pp. [70] K. M. Hodivala-Dilke, C. M. DiPersio, J. A. Kreidberg, and 609–618, 1995. R. O. Hynes, “Novel roles for α3β1integrinasaregulatorof [85] J. M. Auger, M. J. E. Kuijpers,Y.A.Senis,S.P.Watson, andJ. cytoskeletal assembly and as a trans-dominant inhibitor of W. M. Heemskerk, “Adhesion of human and mouse platelets integrin receptor function in mouse keratinocytes,” Journal to collagen under shear: a unifying model,” FASEB Journal, of Cell Biology, vol. 142, no. 5, pp. 1357–1369, 1998. vol. 19, no. 7, pp. 825–827, 2005. [71] D. G. Stupack, X. S. Puente, S. Boutsaboualoy, C. M. [86] L. Meyaard, “The inhibitory collagen receptor LAIR-1 Storgard, and D. A. Cheresh, “Apoptosis of adherent cells by (CD305),” Journal of Leukocyte Biology,vol. 83, no.4,pp. recruitment of caspase-8 to unligated integrins,” Journal of 799–803, 2008. Cell Biology, vol. 155, no. 4, pp. 459–470, 2001. [87] A. C. Curino, L. H. Engelholm, S. S. Yamada et al., “Intra- [72] J. Zhu, K. Motejlek, D. Wang, K. Zang, A. Schmidt, and cellular collagen degradation mediated by uPARAP/Endo180 L. F. Reichardt, “β8 Integrins are required for vascular is a major pathway of extracellular matrix turnover during morphogenesis in mouse embryos,” Development, vol. 129, malignancy,” Journal of Cell Biology, vol. 169, no. 6, pp. 977– no. 12, pp. 2891–2903, 2002. 985, 2005. [73] K. Venstrom and L. Reichardt, “Beta 8 integrins mediate [88] J. A. Kreidberg, M. J. Donovan, S. L. Goldstein et al., “Alpha interactions of chick sensory neurons with laminin-1, colla- 3 beta 1 integrin has a crucial role in kidney and lung gen IV, and fibronectin,” Molecular Biology of the Cell, vol. 6, organogenesis,” Development, vol. 122, no. 11, pp. 3537– no. 4, pp. 419–431, 1995. 3547, 1996. [74] R. Milner,J.B.Relvas, J. Fawcett, andC.Ffrench-Constant, [89] C. M. DiPersio,K.M.Hodivala-Dilke, R. Jaenisch,J.A. “Developmental regulation of αv integrins produces func- Kreidberg, andR.O.Hynes,“α3β1 integrin is required for tional changes in astrocyte behavior,” Molecular and Cellular normal development of the epidermal basement membrane,” Neuroscience, vol. 18, no. 1, pp. 108–118, 2001. Journal of Cell Biology, vol. 137, no. 3, pp. 729–742, 1997. [75] D. Mu,S.Cambier,L.Fjellbirkelandetal., “The integrin [90] U. Mayer, G. Saher, R. Fassler ¨ et al., “Absence of integrin α7 ανβ8 mediates epithelial homeostasis through MT1-MMP- causes a novel form of muscular dystrophy,” Nature Genetics, dependent activation of TGF-β1,” Journal of Cell Biology, vol. vol. 17, no. 3, pp. 318–323, 1997. 157, no. 3, pp. 493–507, 2002. [91] M. A. Stepp, S. Spurr-Michaud, A. Tisdale, J. Elwell, and I. [76] A. Kern, J. Eble, R. Golbik, and K. Kuhn, “Interaction of K. Gipson, “α6β4 integrin heterodimer is a component of type IV collagen with the isolated integrins α1β1and α2β1,” hemidesmosomes,” Proceedings of the National Academy of European Journal of Biochemistry, vol. 215, no. 1, pp. 151– Sciences of the United States of America, vol. 87, no. 22, pp. 159, 1993. 8970–8974, 1990. [77] M. Tulla, O. T. Pentikainen, T. Viitasalo et al., “Selective [92] O. Ibraghimov-Beskrovnaya, J. M. Ervasti, C. J. Leveille, C. binding of collagen subtypes by integrin α1I, α2I, and α10I A. Slaughter, S. W. Sernett, and K. P. Campbell, “Primary domains,” Journal of Biological Chemistry, vol. 276, no. 51, structure of dystrophin-associated glycoproteins linking dys- pp. 48206–48212, 2001. trophin to the extracellular matrix,” Nature, vol. 355, no. [78] M. M. Zutter and S. A. Santoro, “Widespread histologic dis- 6362, pp. 696–702, 1992. tribution of the α2β1 integrin cell-surface collagen receptor,” [93] T. Haenggi and J. M. Fritschy, “Role of dystrophin and utro- American Journal of Pathology, vol. 137, no. 1, pp. 113–120, phin for assembly and function of the dystrophin glycopro- 1990. tein complex in non-muscle tissue,” Cellular and Molecular [79] T. Bengtsson, A. Aszodi, C. Nicolae, E. B. Hunziker, E. Life Sciences, vol. 63, no. 14, pp. 1614–1631, 2006. Lundgren-Akerlund, and R. Fassler ¨ , “Loss of α10β1 integrin [94] J. Nelson, N. V. McFerran, G. Pivato et al., “The 67 kDa expression leads to moderate dysfunction of growth plate laminin receptor: structure, function and role in disease,” chondrocytes,” Journal of Cell Science, vol. 118, no. 5, pp. 929– Bioscience reports, vol. 28, no. 1, pp. 33–48, 2008. 936, 2005. [95] G. Fontanini, S. Vignati, S. Chine´ et al., “67-kilodalton [80] S. N. Popova, M. Barczyk, C. F. Tiger et al., “α11β1 integrin- laminin receptor expression correlates with worse prognostic dependent regulation of periodontal ligament function in the indicators in non-small cell lung carcinomas,” Clinical Cancer erupting mouse incisor,” Molecular and Cellular Biology, vol. Research, vol. 3, no. 2, pp. 227–231, 1997. 27, no. 12, pp. 4306–4316, 2007. [96] D. Waltregny, L. De Leval, S. Menar ´ d, J. De Leval, and V. [81] T. Riikonen, J. Westermarck, L. Koivisto, A. Broberg, V. M. Castronovo, “Independent prognostic value of the 67-kd Kahari, and J. Heino, “Integrin α2β1isapositive regulator laminin receptor in human prostate cancer,” Journal of the Journal of Oncology 17 National Cancer Institute, vol. 89, no. 16, pp. 1224–1227, [113] N. E. Vlahakis, B. A. Young, A. Atakilit et al., “Integrin α9β1 1997. directly binds to vascular endothelial growth factor (VEGF)- A and contributes to VEGF-A-induced angiogenesis,” Journal [97] E. Ardini, E. Tagliabue, A. Magnifico et al., “Co-regulation of Biological Chemistry, vol. 282, no. 20, pp. 15187–15196, and physical association of the 67-kDa monomeric laminin receptor and the α6β4 integrin,” Journal of Biological Chem- istry, vol. 272, no. 4, pp. 2342–2345, 1997. [114] N. E. Vlahakis, B. A. Young, A. Atakilit, and D. Sheppard, “The lymphangiogenic vascular endothelial growth factors [98] T. Ogawa, Y. Tsubota, J. Hashimoto, Y. Kariya, and K. VEGF-C and -D are ligands for the integrin α9β1,” Journal Miyazaki, “The short arm of laminin γ2 chain of laminin- of Biological Chemistry, vol. 280, no. 6, pp. 4544–4552, 2005. 5 (laminin-332) binds syndecan-1 and regulates cellular [115] T. R. Carlson, Y. Feng, P. C. Maisonpierre, M. Mrksich, and A. adhesion and migration by suppressing phosphorylation of O. Morla, “Direct cell adhesion to the angiopoietins mediated integrin β4 chain,” Molecular Biology of the Cell, vol. 18, no. by integrins,” Journal of Biological Chemistry, vol. 276, no. 28, 5, pp. 1621–1633, 2007. pp. 26516–26525, 2001. [99] R. G. Da Silva, B. Tavora, S. D. Robinson et al., “Endothelial [116] S. J. Leu, S. C. T. Lam, and L. F. Lau, “Pro-angiogenic α3β1-integrin represses pathological angiogenesis and sus- activities of CYR61 (CCN1) mediated through integrins tains endothelial-VEGF,” American Journal of Pathology, vol. αvβ3and α6β1 in human umbilical vein endothelial cells,” 177, no. 3, pp. 1534–1548, 2010. Journal of Biological Chemistry, vol. 277, no. 48, pp. 46248– [100] R. Van der Neut, P. Krimpenfort, J. Calafat, C. M. Niessen, 46255, 2002. and A. Sonnenberg, “Epithelial detachment due to absence of [117] S. J. Leu, Y. Liu, N. Chen, C. C. Chen, S. C. T. Lam, and L. F. hemidesmosomes in integrin β null mice,” Nature Genetics, Lau, “Identification of a novel integrin α6β1 binding site in vol. 13, no. 3, pp. 366–369, 1996. the angiogenic inducer CCN1 (CYR61),” Journal of Biological [101] S. N. Nikolopoulos, P. Blaikie, T. Yoshioka, W. Guo, and Chemistry, vol. 278, no. 36, pp. 33801–33808, 2003. F. G. Giancotti, “Integrin β4 signaling promotes tumor [118] S. Mori, C. Y. Wu, S. Yamaji et al., “Direct binding of integrin angiogenesis,” Cancer Cell, vol. 6, no. 5, pp. 471–483, 2004. αvβ3 to FGF1 plays a role in FGF1 signaling,” Journal of [102] T. S. Hiran, J. E. Mazurkiewicz, P. Kreienberg, F. L. Rice, and Biological Chemistry, vol. 283, no. 26, pp. 18066–18075, 2008. S. E. LaFlamme, “Endothelial expression of the α6β4 integrin [119] K. Suzuki, T. Okuno, M. Yamamoto et al., “Semaphorin is negatively regulated during angiogenesis,” Journal of Cell 7A initiates T-cell-mediated inflammatory responses through Science, vol. 116, no. 18, pp. 3771–3781, 2003. α1β1 integrin,” Nature, vol. 446, no. 7136, pp. 680–684, 2007. [103] S. M. Frisch and H. Francis, “Disruption of epithelial [120] J. T. Chao, L. A. Martinez-Lemus, S. J. Kaufman, G. A. cell-matrix interactions induces apoptosis,” Journal of Cell Meininger, K. S. Ramos, and E. Wilson, “Modulation of Biology, vol. 124, no. 4, pp. 619–626, 1994. α7-integrin-mediated adhesion and expression by platelet- [104] S. A. Wickstrom, ¨ K. Radovanac, and R. Fassler ¨ , “Genetic derived growth factor in vascular smooth muscle cells,” analyses of integrin signaling,” Cold Spring Harbor Perspec- American Journal of Physiology, vol. 290, no. 4, pp. C972– tives in Biology, vol. 3, no. 2, 2011. C980, 2006. [105] F. G. Giancotti and E. Ruoslahti, “Integrin signaling,” Science, [121] N. L. Flintoff-Dye, J. Welser, J. Rooney et al., “Role for vol. 285, no. 5430, pp. 1028–1032, 1999. the α7β1 integrin in vascular development and integrity,” [106] M. A. Arnaout, B. Mahalingam, and J. P. Xiong, “Integrin Developmental Dynamics, vol. 234, no. 1, pp. 11–21, 2005. structure, allostery, and bidirectional signaling,” Annual [122] Y. Taooka, J. Chen, T. Yednock, and D. Sheppard, “The Review of Cell and Developmental Biology, vol. 21, pp. 381– integrin α9β1 mediates adhesion to activated endothelial 410, 2005. cells and transendothelial neutrophil migration through [107] H. Methe, S. Hess, and E. R. Edelman, “Endothelial immu- interaction with vascular cell adhesion molecule-1,” Journal nogenicity—a matter of matrix microarchitecture,” Throm- of Cell Biology, vol. 145, no. 2, pp. 413–420, 1999. bosis and Haemostasis, vol. 98, no. 2, pp. 278–282, 2007. [123] I. Staniszewska, S. Zaveri, L. D. Valle et al., “Interaction of [108] Y. Wallez and P. Huber, “Endothelial adherens and tight α9β1 integrin with thrombospondin-1 promotes angiogen- junctions in vascular homeostasis, inflammation and angio- esis,” Circulation Research, vol. 100, no. 9, pp. 1308–1316, genesis,” Biochimica et Biophysica Acta, vol. 1778, no. 3, pp. 794–809, 2008. [124] D. Bouvard, C. Brakebusch, E. Gustafsson et al., “Functional [109] A. Orpana, V. Ranta, T. Mikkola, L. Viinikka, and O. Yliko- consequences of integrin gene mutations in mice,” Circula- rkala, “Inducible nitric oxide and prostacyclin productions tion Research, vol. 89, no. 3, pp. 211–223, 2001. are differently controlled by extracellular matrix and cell [125] R. Fassler ¨ and M. Meyer, “Consequences of lack of β1 integrin density in human vascular endothelial cells,” Journal of gene expression in mice,” Genes and Development, vol. 9, no. Cellular Biochemistry, vol. 64, no. 4, pp. 538–546, 1997. 15, pp. 1896–1908, 1995. [110] M. A. Schwartz and D. W. DeSimone, “Cell adhesion [126] L. E. Stephens, A. E. Sutherland, I. V. Klimanskaya et al., receptors in mechanotransduction,” Current Opinion in Cell “Deletion of β1 integrins in mice results in inner cell mass Biology, vol. 20, no. 5, pp. 551–556, 2008. failure and peri-implantation lethality,” Genes and Develop- [111] J. Ivaska and J. Heino, “Interplay between cell adhesion and ment, vol. 9, no. 15, pp. 1883–1895, 1995. growth factor receptors: from the plasma membrane to the [127] T. R. Carlson, H. Hu, R. Braren, Y. H. Kim, and R. A. Wang, endosomes,” Cell and Tissue Research, vol. 339, no. 1, pp. 111– “Cell-autonomous requirement for β1 integrin in endothelial 120, 2010. cell adhesion, migration and survival during angiogenesis in [112] H. Hutchings, N. Ortega, and J. Plouet, ¨ “Extracellular mice,” Development, vol. 135, no. 12, pp. 2193–2202, 2008. [128] L. Lei, D. Liu, Y. Huang et al., “Endothelial expression of matrix-bound vascular endothelial growth factor promotes endothelial cell adhesion, migration, and survival through β1 integrin is required for embryonic vascular patterning integrin ligation,” The FASEB Journal, vol. 17, no. 11, pp. and postnatal vascular remodeling,” Molecular and Cellular 1520–1522, 2003. Biology, vol. 28, no. 2, pp. 794–802, 2008. 18 Journal of Oncology [129] H. Tanjore, E. M. Zeisberg, B. Gerami-Naini, and R. Kalluri, [146] J. P. Xiong, T. Stehle, B. Diefenbach et al., “Crystal structure “β1 integrin expression on endothelial cells is required of the extracellular segment of integrin αVβ3,” Science, vol. for angiogenesis but not for vasculogenesis,” Developmental 294, no. 5541, pp. 339–345, 2001. Dynamics, vol. 237, no. 1, pp. 75–82, 2008. [147] J. P. Xiong, T. Stehle, S. L. Goodman, and M. A. Arnaout, [130] A. C. Zovein, A. Luque, K. A. Turlo et al., “β1 integrin “New insights into the structural basis of integrin activation,” establishes endothelial cell polarity and arteriolar lumen Blood, vol. 102, no. 4, pp. 1155–1159, 2003. formation via a Par3-dependent mechanism,” Developmental [148] J.-P. Xiong, T. Stehle, R. Zhang et al., “Crystal structure of Cell, vol. 18, no. 1, pp. 39–51, 2010. the extracellular segment of integrin αVβ3incomplex with [131] A. Pozzi, P. E. Moberg, L. A. Miles, S. Wagner, P. Soloway, an Arg-Gly-Asp ligand,” Science, vol. 296, no. 5565, pp. 151– and H. A. Gardner, “Elevated matrix metalloprotease and 155, 2002. angiostatin levels in integrin α1 knockout mice cause reduced [149] M. J. Humphries, E. J. H. Symonds, and A. P. Mould, “Map- tumor vascularization,” Proceedings of the National Academy ping functional residues onto integrin crystal structures,” of Sciences of the United States of America, vol. 97, no. 5, pp. Current Opinion in Structural Biology, vol. 13, no. 2, pp. 236– 2202–2207, 2000. 243, 2003. [132] Z. Zhang, N. E. Ramirez, T. E. Yankeelov et al., “α2β1 integrin [150] A. P. Mould, E. J. Koper, A. Byron, G. Zahn, and M. J. expression in the tumor microenvironment enhances tumor Humphries, “Mapping the ligand-binding pocket of integrin angiogenesis in a tumor cell-specific manner,” Blood, vol. 111, α5β1 using a gain-of-function approach,” Biochemical Jour- no. 4, pp. 1980–1988, 2008. nal, vol. 424, no. 2, pp. 179–189, 2009. [133] J. T. Yang, H. Rayburn, and R. O. Hynes, “Embryonic meso- [151] D. A. Calderwood, “Integrin activation,” Journal of Cell dermal defects in α5 integrin-deficient mice,” Development, Science, vol. 117, no. 5, pp. 657–666, 2004. vol. 119, no. 4, pp. 1093–1105, 1993. [152] S. J. Shattil, C. Kim, and M. H. Ginsberg, “The final steps of [134] S. E. Francis, K. L. Goh, K. Hodivala-Dilke et al., “Central integrin activation: the end game,” Nature Reviews Molecular roles of α5β1 integrin and fibronectin in vascular develop- Cell Biology, vol. 11, no. 4, pp. 288–300, 2010. ment in mouse embryos and embryoid bodies,” Arteriosclero- [153] A. R. Ramjaun and K. Hodivala-Dilke, “The role of cell sis, Thrombosis, and Vascular Biology, vol. 22, no. 6, pp. 927– adhesion pathways in angiogenesis,” International Journal of 933, 2002. Biochemistry and Cell Biology, vol. 41, no. 3, pp. 521–530, [135] P. Parsons-Wingerter, I. M. Kasman, S. Norberg et al., “Uni- form overexpression and rapid accessibility of α5β1 integrin [154] K. L. Wegener, A. W. Partridge, J. Han et al., “Structural Basis on blood vessels in tumors,” American Journal of Pathology, of Integrin Activation by Talin,” Cell, vol. 128, no. 1, pp. 171– vol. 167, no. 1, pp. 193–211, 2005. 182, 2007. [136] E. Georges-Labouesse, N. Messaddeq, G. Yehia, L. Cadalbert, [155] M. Moser, B. Nieswandt, S. Ussar, M. Pozgajova, and R. A. Dierich, and M. Le Meur, “Absence of integrin α6leads Fassler ¨ , “Kindlin-3 is essential for integrin activation and to epidermolysis bullosa and neonatal death in mice,” Nature platelet aggregation,” Nature Medicine, vol. 14, no. 3, pp. 325– Genetics, vol. 13, no. 3, pp. 370–373, 1996. 330, 2008. [137] J. V. Welser, N. D. Lange, N. Flintoff-Dye, H. R. Burkin, and [156] J. Zhu, C. V. Carman, M. Kim, M. Shimaoka, T. A. Springer, D. J. Burkin, “Placental Defects in α7 Integrin Null Mice,” andB.H.Luo,“Requirementof α and β subunit transmem- Placenta, vol. 28, no. 11-12, pp. 1219–1228, 2007. brane helix separation for integrin outside-in signaling,” [138] R. Fassler, E. Georges-Labouesse, and E. Hirsch, “Genetic Blood, vol. 110, no. 7, pp. 2475–2483, 2007. analyses of integrin function in mice,” Current Opinion in [157] T. Xiao, J. Takagi, B. S. Coller, J. H. Wang, and T. A. Springer, Cell Biology, vol. 8, no. 5, pp. 641–646, 1996. “Structural basis for allostery in integrins and binding to [139] X. Z. Huang, J. F. Wu, R. Ferrando et al., “Fatal bilateral fibrinogen-mimetic therapeutics,” Nature, vol. 432, no. 7013, chylothorax in mice lacking the integrin α9β1,” Molecular pp. 59–67, 2004. and Cellular Biology, vol. 20, no. 14, pp. 5208–5215, 2000. [158] N. Nishida, C. Xie, M. Shimaoka, Y. Cheng, T. Walz, and T. A. [140] K. M. Hodivala-Dilke, K. P. McHugh, D. A. Tsakiris et al., Springer, “Activation of leukocyte β2 integrins by conversion “β3-integrin-deficient mice are a model for Glanzmann from bent to extended conformations,” Immunity, vol. 25, no. thrombasthenia showing placental defects and reduced sur- 4, pp. 583–594, 2006. vival,” Journal of Clinical Investigation, vol. 103, no. 2, pp. [159] K. R. Legate, E. Montanez, ˜ O. Kudlacek, and R. Fassler ¨ , “ILK, 229–238, 1999. PINCH and parvin: the tIPP of integrin signalling,” Nature [141] Y. Takada, X. Ye, and S. Simon, “The integrins,” Genome Reviews Molecular Cell Biology, vol. 7, no. 1, pp. 20–31, 2006. Biology, vol. 8, no. 5, article 215, pp. 211–219, 2007. [160] M. Zoller ¨ , “Tetraspanins: push and pull in suppressing and [142] M. H. Ginsberg, A. Partridge, and S. J. Shattil, “Integrin promoting metastasis,” Nature Reviews Cancer, vol. 9, no. 1, regulation,” Current Opinion in Cell Biology, vol. 17, no. 5, pp. 40–55, 2009. pp. 509–516, 2005. [161] J. H. Park, J. M. Ryu, and H. J. Han, “Involvement of [143] M. A. Arnaout, S. L. Goodman, and J. P. Xiong, “Structure caveolin-1 in fibronectin-induced mouse embryonic stem and mechanics of integrin-based cell adhesion,” Current cell proliferation: role of FAK, RhoA, PI3K/Akt, and ERK 1/2 Opinion in Cell Biology, vol. 19, no. 5, pp. 495–507, 2007. pathways,” Journal of Cellular Physiology, vol. 226, no. 1, pp. [144] B. H. Luo, C. V. Carman, and T. A. Springer, “Structural 267–275, 2011. basis of integrin regulation and signaling,” Annual Review of [162] S. H. Lee, Y. J. Lee, S. W. Park, H. S. Kim, and H. J. Immunology, vol. 25, pp. 619–647, 2007. Han, “Caveolin-1 and integrin β1 regulate embryonic stem [145] J. Takagi, K. Strokovich, T. A. Springer, and T. Walz, cell proliferation via p38 MAPK and FAK in high glucose,” “Structure of integrin α5β1 in complex with fibronectin,” Journal of Cellular Physiology, vol. 226, no. 7, pp. 1850–1859, EMBO Journal, vol. 22, no. 18, pp. 4607–4615, 2003. 2011. Journal of Oncology 19 [163] A. Byron, M. R. Morgan, and M. J. Humphries, “Adhesion [178] B. P. Eliceiri, X. S. Puente, J. D. Hood et al., “Src-mediated signalling complexes,” Current Biology, vol. 20, no. 24, pp. coupling of focal adhesion kinase to integrin αvβ5invascular R1063–R1067, 2010. endothelial growth factor signaling,” Journal of Cell Biology, vol. 157, no. 1, pp. 149–160, 2002. [164] M. A. Del Pozo, N. B. Alderson, W. B. Kiosses, H. H. Chiang, R. G. W. Anderson, and M. A. Schwartz, “Integrins regulate [179] S. A. Wickstrom, ¨ K. Alitalo, and J. Keski-Oja, “Endostatin rac targeting by internalization of membrane domains,” associates with integrin α5β1 and caveolin-1, and activates Science, vol. 303, no. 5659, pp. 839–842, 2004. Src via a tyrosyl phosphatase-dependent pathway in human endothelial cells,” Cancer Research, vol. 62, no. 19, pp. 5580– [165] I. J. Salanueva, A. Cerezo, M. C. Guadamillas, and M. A. 5589, 2002. Del Pozo, “Integrin regulation of caveolin function: caveolae review series,” Journal of Cellular and Molecular Medicine, vol. [180] A. Aiyer and J. Varner, “The role of integrins in tumor 11, no. 5, pp. 969–980, 2007. angiogenesis,” in Cancer Drug Discovery Development—Anti- angiogenic Agents in Cancer Therapy,B.A.Teicher andL.M. [166] I. Bethani, S. S. Skanland, ˚ I. Dikic, and A. Acker-Palmer, “Spatial organization of transmembrane receptor signalling,” Ellis, Eds., pp. 49–73, Humana Press, Totowa, NJ, USA, 2008. EMBO Journal, vol. 29, no. 16, pp. 2677–2688, 2010. [181] C. Chandra Kumar, “Signaling by integrin receptors,” Onco- gene, vol. 17, no. 11, pp. 1365–1373, 1998. [167] R. W. Tilghman and J. T. Parsons, “Focal adhesion kinase as a regulator of cell tension in the progression of cancer,” [182] K. I. Nagashima, A. Endo, H. Ogita et al., “Adaptor protein Seminars in Cancer Biology, vol. 18, no. 1, pp. 45–52, 2008. Crk is required for ephrin-B1-induced membrane ruffling and focal complex assembly of human aortic endothelial [168] M. C. Brown, L. A. Cary, J. S. Jamieson, J. A. Cooper, cells,” Molecular Biology of the Cell, vol. 13, no. 12, pp. 4231– and C. E. Turner, “Src and FAK kinases cooperate to phos- 4242, 2002. phorylate paxillin kinase linker, stimulate its focal adhesion localization, and regulate cell spreading and protrusiveness,” [183] F. Paulhe, C. Racaud-Sultan, A. Ragab et al., “Differential Molecular Biology of the Cell, vol. 16, no. 9, pp. 4316–4328, regulation of phosphoinositide metabolism by α vβ3and 2005. αvβ5 integrins upon smooth muscle cell migration,” Journal of Biological Chemistry, vol. 276, no. 45, pp. 41832–41840, [169] E. G. Arias-Salgado, S. Lizano, S. Sarkar, J. S. Brugge, M. H. Ginsberg, and S. J. Shattil, “Src kinase activation by direct interaction with the integrin β cytoplasmic domain,” [184] V. Carloni, R. G. Romanelli, M. Pinzani, G. Laffi,and P. Proceedings of the National Academy of Sciences of the United Gentilini, “Focal adhesion kinase and phospholipase Cγ States of America, vol. 100, no. 23, pp. 13298–13302, 2003. involvement in adhesion and migration of human hepatic stellate cells,” Gastroenterology, vol. 112, no. 2, pp. 522–531, [170] S. K. Hanks, M. B. Calalb, M. C. Harper, and S. K. Patel, “Focal adhesion protein-tyrosine kinase phosphorylated in 1997. response to cell attachment to fibronectin,” Proceedings of the [185] X. Zhang, A. Chattopadhyay, Q. S. Ji et al., “Focal adhesion National Academy of Sciences of the United States of America, kinase promotes phospholipase C-γ1 activity,” Proceedings vol. 89, no. 18, pp. 8487–8491, 1992. of the National Academy of Sciences of the United States of America, vol. 96, no. 16, pp. 9021–9026, 1999. [171] A. Papapetropoulos, D. Fulton, K. Mahboubi et al., “Angi- opoietin-1 inhibits endothelial cell apoptosis via the Akt/ [186] L. Bi, I. Okabe, D. J. Bernard, A. Wynshaw-Boris, and R. survivin pathway,” Journal of Biological Chemistry, vol. 275, L. Nussbaum, “Proliferative defect and embryonic lethality no. 13, pp. 9102–9105, 2000. in mice homozygous for a deletion in the p110α subunit of phosphoinositide 3-kinase,” Journal of Biological Chemistry, [172] S. H. Kim and S. H. Kim, “Antagonistic effect of EGF vol. 274, no. 16, pp. 10963–10968, 1999. on FAK phosphorylation/dephosphorylation in a cell,” Cell Biochemistry and Function, vol. 26, no. 5, pp. 539–547, 2008. [187] E. Lelievre, P. M. Bourbon, L. J. Duan, R. L. Nussbaum, and G. H. Fong, “Deficiency in the p110α subunit of PI3K results [173] J. K. Slack-Davis, S. T. Eblen, M. Zecevic et al., “PAK1 in diminished Tie2 expression and Tie2-/–like vascular phosphorylation of MEK1 regulates fibronectin-stimulated defects in mice,” Blood, vol. 105, no. 10, pp. 3935–3938, 2005. MAPK activation,” Journal of Cell Biology, vol. 162, no. 2, pp. 281–291, 2003. [188] M. Graupera, J. Guillermet-Guibert, L. C. Foukas et al., [174] M. L. Edin and R. L. Juliano, “Raf-1 serine 338 phospho- “Angiogenesis selectively requires the p110α isoform of PI3K to control endothelial cell migration,” Nature, vol. 453, no. rylation plays a key role in adhesion-dependent activation 7195, pp. 662–666, 2008. of extracellular signal-regulated kinase by epidermal growth factor,” Molecular and Cellular Biology, vol. 25, no. 11, pp. [189] A. Sudhakar, H. Sugimoto, C. Yang, J. Lively, M. Zeisberg, and 4466–4475, 2005. R. Kalluri, “Human tumstatin and human endostatin exhibit [175] T. L. Shen, A. Y. J. Park, A. Alcaraz et al., “Conditional distinct antiangiogenic activities mediated by αvβ and α5β1 integrins,” Proceedings of the National Academy of Sciences of knockout of focal adhesion kinase in endothelial cells reveals its role in angiogenesis and vascular development in late the United States of America, vol. 100, no. 8, pp. 4766–4771, embryogenesis,” Journal of Cell Biology, vol. 169, no. 6, pp. 941–952, 2005. [190] S. Gupta, A. R. Ramjaun, P. Haiko et al., “Binding of ras to [176] D. L. Courter, L. Lomas, M. Scatena, and C. M. Giachelli, “Src phosphoinositide 3-kinase p110α is required for ras-driven tumorigenesis in mice,” Cell, vol. 129, no. 5, pp. 957–968, kinase activity is required for integrin αvβ 3-mediated acti- vation of nuclear factor-κB,” Journal of Biological Chemistry, 2007. vol. 280, no. 13, pp. 12145–12151, 2005. [191] M. S. Roberts, A. J. Woods, P. E. Shaw, and J. C. Norman, “ERK1 associates with αvβ3 integrin and regulates cell [177] J. Zaric and C. Ruegg ¨ , “Integrin-mediated adhesion and spreading on vitronectin,” Journal of Biological Chemistry, soluble ligand binding stabilize COX-2 protein levels in vol. 278, no. 3, pp. 1975–1985, 2003. endothelial cells by inducing expression and preventing degradation,” Journal of Biological Chemistry, vol. 280, no. 2, [192] S. M. Short, G. A. Talbott, and R. L. Juliano, “Integrin- pp. 1077–1085, 2005. mediated signaling events in human endothelial cells,” 20 Journal of Oncology Molecular Biology of the Cell, vol. 9, no. 8, pp. 1969–1980, [208] V. Samanna, H. Wei, D. Ego-Osuala, and M. A. Chellaiah, 1998. “Alpha-V-dependent outside-in signaling is required for the regulation of CD44 surface expression, MMP-2 secretion, [193] M. Huser ¨ , J. Luckett, A. Chiloeches et al., “MEK kinase and cell migration by osteopontin in human melanoma activity is not necessary for Raf-1 function,” EMBO Journal, cells,” Experimental Cell Research, vol. 312, no. 12, pp. 2214– vol. 20, no. 8, pp. 1940–1951, 2001. 2230, 2006. [194] S. Giroux, M. Tremblay, D. Bernard et al., “Embryonic [209] M. M. Zutter, S. A. Santoro, W. D. Staatz, and Y. L. Tsung, death of Mek1-deficient mice reveals a role for this kinase “Re-expression of the α2β1 integrin abrogates the malignant in angiogenesis in the labyrinthine region of the placenta,” phenotype of breast carcinoma cells,” Proceedings of the Current Biology, vol. 9, no. 7, pp. 369–372, 1999. National Academy of Sciences of the United States of America, [195] J. D. Hood, R. Frausto, W. B. Kiosses, M. A. Schwartz, and vol. 92, no. 16, pp. 7411–7415, 1995. D. A. Cheresh, “Differential αv integrin-mediated Ras-ERK [210] A. Kren, V. Baeriswyl, F. Lehembre et al., “Increased tumor signaling during two pathways of angiogenesis,” Journal of cell dissemination and cellular senescence in the absence of Cell Biology, vol. 162, no. 5, pp. 933–943, 2003. β1-integrin function,” EMBO Journal, vol. 26, no. 12, pp. [196] K. K. Wary, F. Mainiero, S. J. Isakoff,E.E.Marcantonio,and 2832–2842, 2007. F. G. Giancotti, “The adaptor protein Shc couples a class of [211] H. Zhao, F. P. Ross, and S. L. Teitelbaum, “Unoccupied integrins to the control of cell cycle progression,” Cell, vol. 87, αvβ3 integrin regulates osteoclast apoptosis by transmitting a no. 4, pp. 733–743, 1996. positive death signal,” Molecular Endocrinology, vol. 19, no. 3, [197] A. Pozzi, K. K. Wary, F. G. Giancotti, and H. A. Gardner, pp. 771–780, 2005. “Integrin α1β1 mediates a unique collagen-dependent pro- [212] S. M. Frisch and R. A. Screaton, “Anoikis mechanisms,” liferation pathway in vivo,” Journal of Cell Biology, vol. 142, Current Opinion in Cell Biology, vol. 13, no. 5, pp. 555–562, no. 2, pp. 587–594, 1998. [198] A. K. Fournier, L. E. Campbell, P. Castagnino et al., “Rac- [213] D. G. Stupack, T. Teitz, M. D. Potter et al., “Potentiation of dependent cyclin D1 gene expression regulated by cadherin- neuroblastoma metastasis by loss of caspase-8,” Nature, vol. and integrin-mediated adhesion,” Journal of Cell Science, vol. 439, no. 7072, pp. 95–99, 2006. 121, no. 2, pp. 226–233, 2008. [214] J. S. Desgrosellier, L. A. Barnes, D. J. Shields et al., “An [199] E. A. Klein, L. Yin, D. Kothapalli et al., “Cell-cycle control by integrin α(v)β(3)-c-Src oncogenic unit promotes anchorage- physiological matrix elasticity and in vivo tissue stiffening,” independence and tumor progression,” Nature Medicine, vol. Current Biology, vol. 19, no. 18, pp. 1511–1518, 2009. 15, no. 10, pp. 1163–1169, 2009. [200] S. Klein, A. R. De Fougerolles, P. Blaikie et al., “α5β1 integrin [215] M. L. Matter and E. Ruoslahti, “A signaling pathway from the activates an NF-κB-dependent program of gene expression α5β1and αvβ3 integrins that elevates bcl-2 transcription,” important for angiogenesis and inflammation,” Molecular Journal of Biological Chemistry, vol. 276, no. 30, pp. 27757– and Cellular Biology, vol. 22, no. 16, pp. 5912–5922, 2002. 27763, 2001. [201] M. Reidy, P. Zihlmann, J. A. Hubbell, and H. Hall, “Acti- [216] F. Aoudjit and K. Vuori, “Matrix attachment regulates Fas- vation of cell-survival transcription factor NFκBinL1Ig6- induced apoptosis in endothelial cells: a role for c-Flip and stimulated endothelial cells,” Journal of Biomedical Materials implications for anoikis,” Journal of Cell Biology, vol. 153, no. Research Part A, vol. 77, no. 3, pp. 542–550, 2006. 3, pp. 633–643, 2001. [202] M. Scatena, M. Almeida, M. L. Chaisson, N. Fausto, R. F. [217] W. Bao and S. Stromblad, ¨ “Integrin αv-mediated inactivation Nicosia, and C. M. Giachelli, “NF-κB mediates αvβ3 integrin- of p53 controls a MEK1-dependent melanoma cell survival induced endothelial cell survival,” Journal of Cell Biology, vol. pathway in three-dimensional collagen,” Journal of Cell 141, no. 4, pp. 1083–1093, 1998. Biology, vol. 167, no. 4, pp. 745–756, 2004. [203] O. Dormond, M. Bezzi, A. Mariotti, and C. Ruegg ¨ , “Pros- [218] A. Alavi, J. D. Hood, R. Frausto, D. G. Stupack, and D. A. taglandin E2 promotes integrin αvβ3-dependent endothelial Cheresh, “Role of Raf in vascular protection from distinct cell adhesion, Rac-activation, and spreading through cAMP/ apoptotic stimuli,” Science, vol. 301, no. 5629, pp. 94–96, PKA-dependent signaling,” Journal of Biological Chemistry, vol. 277, no. 48, pp. 45838–45846, 2002. [219] E. Ruoslahti, “Specialization of tumour vasculature,” Nature [204] C. S. Boosani, A. P. Mannam, D. Cosgrove et al., “Regulation Reviews Cancer, vol. 2, no. 2, pp. 83–90, 2002. of COX-2-mediated signaling by α3typeIVnoncollagenous [220] N. Alam, H. L. Goel, M. J. Zarif et al., “The integrin—growth domain in tumor angiogenesis,” Blood, vol. 110, no. 4, pp. factor receptor duet,” Journal of Cellular Physiology, vol. 213, 1168–1177, 2007. no. 3, pp. 649–653, 2007. [205] T. Kisseleva, L. Song, M. Vorontchikhina, N. Feirt, J. Kitajew- [221] G. Serini, L. Napione, M. Arese, and F. Bussolino, “Besides ski, and C. Schindler, “NF-κB regulation of endothelial cell adhesion: new perspectives of integrin functions in angio- function during LPS-induced toxemia and cancer,” Journal of genesis,” Cardiovascular Research, vol. 78, no. 2, pp. 213–222, Clinical Investigation, vol. 116, no. 11, pp. 2955–2963, 2006. [206] H. Lahlou, V. Sanguin-Gendreau, D. Zuo et al., “Mammary [222] E. S. Wijelath, S. Rahman, M. Namekata et al., “Heparin-II epithelial-specific disruption of the focal adhesion kinase domain of fibronectin is a vascular endothelial growth factor- blocks mammary tumor progression,” Proceedings of the binding domain: enhancement of VEGF biological activity National Academy of Sciences of the United States of America, by a singular growth factor/matrix protein synergism,” vol. 104, no. 51, pp. 20302–20307, 2007. Circulation Research, vol. 99, no. 8, pp. 853–860, 2006. [207] Y. Pylayeva, K. M. Gillen, W. Gerald, H. E. Beggs, L. F. [223] C. Q. Zhu, S. N. Popova, E. R. S. Brown et al., “Integrin α11 regulates IGF2 expression in fibroblasts to enhance Reichardt, and F. G. Giancotti, “Ras- and PI3K-dependent breast tumorigenesis in mice and humans requires focal tumorigenicity of human non-small-cell lung cancer cells,” Proceedings of the National Academy of Sciences of the United adhesion kinase signaling,” Journal of Clinical Investigation, vol. 119, no. 2, pp. 252–266, 2009. States of America, vol. 104, no. 28, pp. 11754–11759, 2007. Journal of Oncology 21 [224] A. Orecchia, P. M. Lacal, C. Schietroma, V. Morea, G. [241] A. C. Aplin, E. Fogel, and R. F. Nicosia, “MCP-1 promotes Zambruno, and C. M. Failla, “Vascular endothelial growth mural cell recruitment during angiogenesis in the aortic ring factor receptor-1 is deposited in the extracellular matrix by model,” Angiogenesis, vol. 13, no. 3, pp. 219–226, 2010. endothelial cells and is a ligand for the α5β1 integrin,” Journal [242] E. Iivanainen, V. M. Kah ¨ ar ¨ i, J. Heino, and K. Elenius, of Cell Science, vol. 116, no. 17, pp. 3479–3489, 2003. “Endothelial cell-matrix interactions,” Microscopy Research and Technique, vol. 60, no. 1, pp. 13–22, 2003. [225] K. Kajiya, S. Hirakawa, B. Ma, I. Drinnenberg, and M. [243] J. E. Rundhaug, “Matrix metalloproteinases and angiogene- Detmar, “Hepatocyte growth factor promotes lymphatic vessel formation and function,” EMBO Journal, vol. 24, no. sis,” Journal of Cellular and Molecular Medicine, vol. 9, no. 2, pp. 267–285, 2005. 16, pp. 2885–2895, 2005. [244] G. Murphy and H. Nagase, “Localizing matrix metallo- [226] M. Murakami, A. Elfenbein, and M. Simons, “Non-canonical proteinase activities in the pericellular environment,” FEBS fibroblast growth factor signalling in angiogenesis,” Cardio- Journal, vol. 278, no. 1, pp. 2–15, 2011. vascular Research, vol. 78, no. 2, pp. 223–231, 2008. [245] G. O. Ahn and J. M. Brown, “Matrix metalloproteinase-9 is [227] G. Camenisch, M. T. Pisabarro, D. Sherman et al., “ANGPTL3 required for tumor vasculogenesis but not for angiogenesis: stimulates endothelial cell adhesion and migration via inte- role of bone marrow-derived myelomonocytic cells,” Cancer grin αvβ3 and induces blood vessel formation in vivo,” Cell, vol. 13, no. 3, pp. 193–205, 2008. Journal of Biological Chemistry, vol. 277, no. 19, pp. 17281– [246] H. M. Eilken and R. H. Adams, “Dynamics of endothelial cell 17290, 2002. behavior in sprouting angiogenesis,” Current Opinion in Cell [228] S. M. Dallabrida, N. Ismail, J. R. Oberle, B. E. Himes, and M. Biology, vol. 22, no. 5, pp. 617–625, 2010. A. Rupnick, “Angiopoietin-1 promotes cardiac and skeletal [247] W. Risau, “Mechanisms of angiogenesis,” Nature, vol. 386, myocyte survival through integrins,” Circulation research, vol. no. 6626, pp. 671–674, 1997. 96, no. 4, pp. e8–e24, 2005. [248] J. Folkman, “Looking for a good endothelial address,” Cancer [229] E. C. Finger and A. J. Giaccia, “Hypoxia, inflammation, and Cell, vol. 1, no. 2, pp. 113–115, 2002. the tumor microenvironment in metastatic disease,” Cancer [249] T. Asahara, T. Murohara, A. Sullivan et al., “Isolation and Metastasis Reviews, vol. 29, no. 2, pp. 285–293, 2010. of putative progenitor endothelial cells for angiogenesis,” [230] A. Billioux, U. Modlich, and R. Bicknell, “Angiogenesis,” in Science, vol. 275, no. 5302, pp. 964–967, 1997. The Cancer Handbook, M. Alison, Ed., vol. 1, pp. 144–154, [250] F. Brellier, R. P. Tucker, and R. Chiquet-Ehrismann, John Wiley & Sons, 2007. “Tenascins and their implications in diseases and tissue [231] P. Carmeliet, “Angiogenesis in health and disease,” Nature mechanics,” Scandinavian Journal of Medicine and Science in Medicine, vol. 9, no. 6, pp. 653–660, 2003. Sports, vol. 19, no. 4, pp. 511–519, 2009. [232] J. Folkman, K. Watson, D. Ingber, and D. Hanahan, “Induc- [251] M. Kaspar, L. Zardi, and D. Neri, “Fibronectin as target for tion of angiogenesis during the transition from hyperplasia tumor therapy,” International Journal of Cancer, vol. 118, no. to neoplasia,” Nature, vol. 339, no. 6219, pp. 58–61, 1989. 6, pp. 1331–1339, 2006. [233] N. Weidner, J. P. Semple, W. R. Welch, and J. Folkman, [252] M. Midulla, R. Verma, M. Pignatelli, M. A. Ritter, N. S. “Tumor angiogenesis and metastasis—correlation in invasive Courtenay-Luck, and A. J. T. George, “Source of oncofetal breast carcinoma,” New England Journal of Medicine, vol. 324, ED-B-containing fibronectin: implications of production of no. 1, pp. 1–8, 1991. both tumor and endothelial cells,” Cancer Research, vol. 60, [234] J. Kandel, E. Bossy-Wetzel, F. Radvanyi, M. Klagsbrun, J. no. 1, pp. 164–169, 2000. Folkman, and D. Hanahan, “Neovascularization is associated [253] K. S. Midwood and G. Orend, “The role of tenascin-C in tis- with a switch to the export of bFGF in the multistep sue injury and tumorigenesis,” Journal of Cell Communication development of fibrosarcoma,” Cell, vol. 66, no. 6, pp. 1095– and Signaling, vol. 3, no. 3-4, pp. 287–310, 2009. 1104, 1991. [254] M. Degen, F. Brellier, S. Schenk et al., “Tenascin-W, a new [235] E. Y. Lin and J. W. Pollard, “Tumor-associated macrophages marker of cancer stroma, is elevated in sera of colon and press the angiogenic switch in breast cancer,” Cancer breast cancer patients,” International Journal of Cancer, vol. Research, vol. 67, no. 11, pp. 5064–5066, 2007. 122, no. 11, pp. 2454–2461, 2008. [236] M. C. Schmid and J. A. Varner, “Myeloid cell trafficking and [255] E. Martina, R. Chiquet-Ehrismann, and F. Brellier, tumor angiogenesis,” Cancer Letters, vol. 250, no. 1, pp. 1–8, “Tenascin-W: an extracellular matrix protein associated with osteogenesis and cancer,” International Journal of Biochem- [237] N. Ferrara, H. P. Gerber, and J. LeCouter, “The biology of istry and Cell Biology, vol. 42, no. 9, pp. 1412–1415, 2010. VEGF and its receptors,” Nature Medicine,vol. 9, no.6,pp. [256] B. Dome, ¨ M. J. C. Hendrix, S. Paku, J. Tov ´ ar ´ i, and J. 669–676, 2003. T´ımar ´ , “Alternative vascularization mechanisms in cancer: [238] E. C. Keeley, B. Mehrad, and R. M. Strieter, “Chemokines pathology and therapeutic implications,” American Journal of as mediators of tumor angiogenesis and neovascularization,” Pathology, vol. 170, no. 1, pp. 1–15, 2007. Experimental Cell Research, vol. 317, no. 5, pp. 685–690, 2011. [257] F. Hillen and A. W. Griffioen, “Tumour vascularization: [239] K. H. Hong, J. Ryu, and K. H. Han, “Monocyte chemoattrac- sprouting angiogenesis and beyond,” Cancer and Metastasis tant protein-1-induced angiogenesis is mediated by vascular Reviews, vol. 26, no. 3-4, pp. 489–502, 2007. endothelial growth factor-A,” Blood, vol. 105, no. 4, pp. 1405– [258] D. Lyden, K. Hattori, S. Dias et al., “Impaired recruitment 1407, 2005. of bone-marrow-derived endothelial and hematopoietic pre- cursor cells blocks tumor angiogenesis and growth,” Nature [240] J. Niu, A. Azfer, O. Zhelyabovska, S. Fatma, and P. E. Kolat- Medicine, vol. 7, no. 11, pp. 1194–1201, 2001. tukudy, “Monocyte chemotactic protein (MCP)-1 promotes angiogenesis via a novel transcription factor, MCP-1-induced [259] D. Ribatti, “The involvement of endothelial progenitor cells in tumor angiogenesis,” Journal of Cellular and Molecular protein (MCPIP),” Journal of Biological Chemistry, vol. 283, no. 21, pp. 14542–14551, 2008. Medicine, vol. 8, no. 3, pp. 294–300, 2004. 22 Journal of Oncology [260] M. Reyes, A. Dudek, B. Jahagirdar, L. Koodie, P. H. Marker, withdrawal,” Journal of Clinical Investigation, vol. 103, no. 2, and C. M. Verfaillie, “Origin of endothelial progenitors pp. 159–165, 1999. in human postnatal bone marrow,” Journal of Clinical [276] A. J. Maniotis, R. Folberg, A. Hess et al., “Vascular channel Investigation, vol. 109, no. 3, pp. 337–346, 2002. formation by human melanoma cells in vivo and in vitro: [261] M. L. Iruela-Arispe and G. E. Davis, “Cellular and molecular vasculogenic mimicry,” American Journal of Pathology, vol. mechanisms of vascular lumen formation,” Developmental 155, no. 3, pp. 739–752, 1999. Cell, vol. 16, no. 2, pp. 222–231, 2009. [277] R. Folberg and A. J. Maniotis, “Vasculogenic mimicry,” Acta [262] B. Strilic, ´ T. Kucer ˇ a, J. Eglinger et al., “The molecular basis Pathologica, Microbiologica. et Immunologica Scandinavica, of vascular lumen formation in the developing mouse aorta,” vol. 112, no. 7-8, pp. 508–525, 2004. Developmental Cell, vol. 17, no. 4, pp. 505–515, 2009. [278] A. J. G. Potgens, ¨ M. C. Van Altena, N. H. Lubsen, D. J. Ruiter, [263] C. M. Ghajar, S. C. George, and A. J. Putnam, “Matrix met- and R. M. W. De Waal, “Analysis of the tumor vasculature and alloproteinase control of capillary morphogenesis,” Critical metastatic behavior of xenografts of human melanoma cell Reviews in Eukaryotic Gene Expression, vol. 18, no. 3, pp. 251– lines transfected with vascular permeability factor,” American 278, 2008. Journal of Pathology, vol. 148, no. 4, pp. 1203–1217, 1996. [264] R. Hildenbrand, H. Allgayer, A. Marx, and P. Stroebel, [279] R. Clarijs, I. Otte-Holler ¨ , D. J. Ruiter, and R. M. W. De Waal, “Modulators of the urokinase-type plasminogen activation “Presence of a fluid-conducting meshwork in xenografted system for cancer,” Expert Opinion on Investigational Drugs, cutaneous and primary human uveal melanoma,” Investiga- vol. 19, no. 5, pp. 641–652, 2010. tive Ophthalmology and Visual Science, vol. 43, no. 4, pp. 912– 918, 2002. [265] F. Bougatef, C. Quemener, S. Kellouche et al., “EMMPRIN promotes angiogenesis through hypoxia-inducible factor-2α- [280] T. Kucer ˇ a and E. Lammert, “Ancestral vascular tube forma- mediated regulation of soluble VEGF isoforms and their tion and its adoption by tumors,” Biological Chemistry, vol. receptor VEGFR-2,” Blood, vol. 114, no. 27, pp. 5547–5556, 390, no. 10, pp. 985–994, 2009. 2009. [281] W. Ruf, E. A. Seftor, R. J. Petrovan et al., “Differential role [266] P. C. Brooks, S. Stromblad, ¨ L. C. Sanders et al., “Localization of tissue factor pathway inhibitors 1 and 2 in melanoma of matrix metalloproteinase MMP-2 to the surface of invasive vasculogenic mimicry,” Cancer Research, vol. 63, no. 17, pp. cells by interaction with integrin αvβ3,” Cell, vol. 85, no. 5, 5381–5389, 2003. pp. 683–693, 1996. [282] S. Anand, B. K. Majeti, L. M. Acevedo et al., “MicroRNA- [267] V. Djonov, M. Schmid, S. A. Tschanz, and P. H. Burri, 132-mediated loss of p120RasGAP activates the endothelium “Intussusceptive angiogenesis. Its role in embryonic vascular to facilitate pathological angiogenesis,” Nature Medicine, vol. network formation,” Circulation Research, vol. 86, no. 3, pp. 16, no. 8, pp. 909–914, 2010. 286–292, 2000. [283] S. Anand and D. A. Cheresh, “MicroRNA-mediated regula- [268] H. Kurz, P. H. Burri, and V. G. Djonov, “Angiogenesis tion of the angiogenic switch,” Current Opinion in Hematol- and vascular remodeling by intussusception: from form to ogy, vol. 18, no. 3, pp. 171–176, 2011. function,” News in Physiological Sciences,vol. 18, no.2,pp. [284] G. Bellon, L. Martiny, and A. Robinet, “Matrix metallopro- 65–70, 2003. teinases and matrikines in angiogenesis,” Critical Reviews in [269] A. N. Makanya, R. Hlushchuk, and V. G. Djonov, “Intussus- Oncology/Hematology, vol. 49, no. 3, pp. 203–220, 2004. ceptive angiogenesis and its role in vascular morphogenesis, [285] P. Nyberg, L. Xie, and R. Kalluri, “Endogenous inhibitors patterning, and remodeling,” Angiogenesis,vol. 12, no.2,pp. of angiogenesis,” Cancer Research, vol. 65, no. 10, pp. 3967– 113–123, 2009. 3979, 2005. [270] D. J. Brat and E. G. Van Meir, “Glomeruloid microvascular [286] M. Shimaoka and T. A. Springer, “Therapeutic antagonists proliferation orchestrated by VPF/VEGF: a new world of and conformational regulation of integrin function,” Nature angiogenesis research,” American Journal of Pathology, vol. Reviews Drug Discovery, vol. 2, no. 9, pp. 703–716, 2003. 158, no. 3, pp. 789–796, 2001. [287] J. A. Eble and J. Haier, “Integrins in cancer treatment,” [271] O. Straume, P. O. Chappuis, H. B. Salvesen et al., “Prognostic Current Cancer Drug Targets, vol. 6, no. 2, pp. 89–105, 2006. importance of glomeruloid microvascular proliferation indi- [288] S. M. Short, A. Derrien, R. P. Narsimhan, J. Lawler, D. cates an aggressive angiogenic phenotype in human cancers,” E. Ingber, and B. R. Zetter, “Inhibition of endothelial cell Cancer Research, vol. 62, no. 23, pp. 6808–6811, 2002. migration by thrombospondin-1 type-1 repeats is mediated [272] J. Holash, P. C. Maisonpierre, D. Compton et al., “Vessel by β1 integrins,” Journal of Cell Biology, vol. 168, no. 4, pp. cooption, regression, and growth in tumors mediated by 643–653, 2005. angiopoietins and VEGF,” Science, vol. 284, no. 5422, pp. [289] X. Zhang and J. Lawler, “Thrombospondin-based antiangio- 1994–1998, 1999. genic therapy,” Microvascular Research,vol. 74, no.2-3,pp. [273] B. Dome, ¨ S. Paku, B. Somlai, and J. Timar, “Vascularization 90–99, 2007. of cutaneous melanoma involves vessel co-option and has [290] P. C. Colorado, A. Torre, G. Kamphaus et al., “Anti- clinical significance,” Journal of Pathology, vol. 197, no. 3, pp. angiogenic cues from vascular basement membrane colla- 355–362, 2002. gen,” Cancer Research, vol. 60, no. 9, pp. 2520–2526, 2000. [274] M. Scharpfenecker, U. Fiedler, Y. Reiss, and H. G. Augustin, [291] P. Nyberg, L. Xie, H. Sugimoto et al., “Characterization of “The Tie-2 ligand angiopoietin-2 destabilizes quiescent the anti-angiogenic properties of arresten, an α1β1 integrin- endothelium through an internal autocrine loop mecha- dependent collagen-derived tumor suppressor,” Experimental nism,” Journal of Cell Science, vol. 118, no. 4, pp. 771–780, Cell Research, vol. 314, no. 18, pp. 3292–3305, 2008. [292] B. P. Woodall, A. Nystro ¨m,R.A.Iozzo et al., “Integrin α2β1 [275] L. E. Benjamin, D. Golijanin, A. Itin, D. Pode, and E. Keshet, is the required receptor for endorepellin angiostatic activity,” “Selective ablation of immature blood vessels in established Journal of Biological Chemistry, vol. 283, no. 4, pp. 2335– human tumors follows vascular endothelial growth factor 2343, 2008. Journal of Oncology 23 [293] M. Mongiat, S. M. Sweeney, J. D. San Antonio, J. Fu, and [308] S. A. Wickstrom, ¨ K. Alitalo, and J. Keski-Oja, “An endostatin- R. V. Iozzo, “Endorepellin, a novel inhibitor of angiogenesis derived peptide interacts with integrins and regulates actin derived from the C terminus of perlecan,” Journal of Biologi- cytoskeleton and migration of endothelial cells,” Journal of cal Chemistry, vol. 278, no. 6, pp. 4238–4249, 2003. Biological Chemistry, vol. 279, no. 19, pp. 20178–20185, 2004. [294] C. Marcinkiewicz, P. H. Weinreb, J. J. Calvete et al., “Obtu- [309] M. E. Cianfrocca, K. A. Kimmel, J. Gallo et al., “Phase 1 trial statin: a potent selective inhibitor of α1β1 integrin in vitro of the antiangiogenic peptide ATN-161 (Ac-PHSCN-NH 2), and angiogenesis in vivo,” Cancer Research, vol. 63, no. 9, pp. a beta integrin antagonist, in patients with solid tumours,” 2020–2023, 2003. British Journal of Cancer, vol. 94, no. 11, pp. 1621–1626, 2006. [295] M. C. Brown, I. Staniszewska, L. Del Valle, G. P. Tuszynski, [310] A. P. Mould, L. Burrows, and M. J. Humphries, “Iden- and C. Marcinkiewicz, “Angiostatic activity of obtustatin as tification of amino acid residues that form part of the α1β1 integrin inhibitor in experimental melanoma growth,” ligand- binding pocket of integrin α5β1,” Journal of Biological International Journal of Cancer, vol. 123, no. 9, pp. 2195– Chemistry, vol. 273, no. 40, pp. 25664–25672, 1998. 2203, 2008. [311] L. Marinelli, A. Meyer, D. Heckmann, A. Lavecchia, E. Nov- [296] J. A. Eble, B. Beermann, H.-J. Hinz, and A. Schmidt- ellino, and H. Kessler, “Ligand binding analysis for human Hederich, “α2β1 integrin is not recognized by rhodocytin but α5β1 integrin: strategies for designing new α5β1 integrin is the specific, high affinity target of rhodocetin, an RGD- antagonists,” Journal of Medicinal Chemistry, vol. 48, no. 13, independent disintegrin and potent inhibitor of cell adhesion pp. 4204–4207, 2005. to collagen,” Journal of Biological Chemistry, vol. 276, no. 15, [312] N. Umeda, S. Kachi, H. Akiyama et al., “Suppression pp. 12274–12284, 2001. and regression of choroidal neovascularization by systemic [297] J. A. Eble, S. Niland, A. Dennes, A. Schmidt-Hederich, P. administration of an α5β1 integrin antagonist,” Molecular Bruckner, and G. Brunner, “Rhodocetin antagonizes stromal Pharmacology, vol. 69, no. 6, pp. 1820–1828, 2006. tumorinvasioninvitro andother α2β1 integrin-mediated [313] S. K. Kuwada, “Volociximab, an angiogenesis-inhibiting cell functions,” Matrix Biology, vol. 21, no. 7, pp. 547–558, chimeric monoclonal antibody,” Current Opinion in Molec- ular Therapeutics, vol. 9, no. 1, pp. 92–98, 2007. [298] J. Zhou, V. L. Rothman, I. Sargiannidou et al., “Cloning and [314] M. L. Wahl, T. L. Moser, and S. V. Pizzo, “Angiostatin and characterization of angiocidin, a tumor cell binding protein anti-angiogenic therapy in human disease,” Recent Progress for thrombospondin-1,” Journal of Cellular Biochemistry, vol. in Hormone Research, vol. 59, pp. 73–104, 2004. 92, no. 1, pp. 125–146, 2004. [315] D. Zhang, P. L. Kaufman, G. Gao, R. A. Saunders, and J. [299] Y. Sabherwal, V. L. Rothman, S. Dimitrov et al., “Integrin X. Ma, “Intravitreal injection of plasminogen kringle 5, an α2β1 mediates the anti-angiogenic and anti-tumor activities endogenous angiogenic inhibitor, arrests retinal neovascu- of angiocidin, a novel tumor-associated protein,” Experimen- larization in rats,” Diabetologia, vol. 44, no. 6, pp. 757–765, tal Cell Research, vol. 312, no. 13, pp. 2443–2453, 2006. [300] R. C. Pandey, M. W. Toussaint, J. C. McGuire, and [316] W.R. Ji, F. J. Castellino, Y. Chang et al., “Characterization of M. C. Thomas, “Maggiemycin and anhydromaggiemycin: kringle domains of angiostatin as antagonists of endothelial two novel anthracyclinone antitumor antibiotics—isolation, cell migration, an important process in angiogenesis,” FASEB structures, partial synthesis and biological properties,” Jour- Journal, vol. 12, no. 15, pp. 1731–1738, 1998. nal of Antibiotics, vol. 42, no. 11, pp. 1567–1577, 1989. [317] Y. Hamano and R. Kalluri, “Tumstatin, the NC1 domain of [301] J. Kap ¨ yla, ¨ O. T. Pentikainen, ¨ T. Nyronen ¨ et al., “Small α3 chain of type IV collagen, is an endogenous inhibitor molecule designed to target metal binding site in the α2I of pathological angiogenesis and suppresses tumor growth,” domain inhibits integrin function,” Journal of Medicinal Biochemical and Biophysical Research Communications, vol. Chemistry, vol. 50, no. 11, pp. 2742–2746, 2007. 333, no. 2, pp. 292–298, 2005. [302] L. Nissinen, O. T. Pentikainen, ¨ A. Jouppila et al., “A small- [318] Y. Maeshima, P. C. Colorado, and R. Kalluri, “Two RGD- molecule inhibitor of integrin α2β1 introduces a new strategy independent α(v)β3 integrin binding sites on tumstatin for antithrombotic therapy,” Thrombosis and Haemostasis, regulate distinct anti-tumor properties,” Journal of Biological vol. 103, no. 2, pp. 387–397, 2010. Chemistry, vol. 275, no. 31, pp. 23745–23750, 2000. [303] Y. Funahashi, N. H. Sugi, T. Semba et al., “Sulfonamide [319] N. Floquet, S. Pasco, L. Ramont et al., “The antitumor derivative, E7820, is a unique angiogenesis inhibitor sup- properties of the α3(IV)-(185–203) peptide from the NC1 pressing an expression of integrin α2 subunit on endothe- domain of type IV collagen (tumstatin) are conformation- lium,” Cancer Research, vol. 62, no. 21, pp. 6116–6123, 2002. dependent,” Journal of Biological Chemistry, vol. 279, no. 3, [304] M. S. O’Reilly, T. Boehm, Y. Shing et al., “Endostatin: an pp. 2091–2100, 2004. endogenous inhibitor of angiogenesis and tumor growth,” [320] C. Magnon, A. Galaup, B. Mullan et al., “Canstatin acts Cell, vol. 88, no. 2, pp. 277–285, 1997. on endothelial and tumor cells via mitochondrial damage [305] R. S. Herbst, K. R. Hess, H. T. Tran et al., “Phase I study initiated through interaction with αvβ3and αvβ5 integrins,” of recombinant human endostatin in patients with advanced Cancer Research, vol. 65, no. 10, pp. 4353–4361, 2005. solid tumors,” Journal of Clinical Oncology, vol. 20, no. 18, pp. [321] E. Petitclerc, A. Boutaud, A. Prestayko et al., “New functions 3792–3803, 2002. for non-collagenous domains of human collagen type IV. [306] J. Dixelius, H. Larsson, T. Sasaki et al., “Endostatin-induced Novel integrin ligands inhibiting angiogenesis and tumor tyrosine kinase signaling through the Shb adaptor protein growth in vivo,” Journal of Biological Chemistry, vol. 275, no. regulates endothelial cell apoptosis,” Blood, vol. 95, no. 11, 11, pp. 8051–8061, 2000. pp. 3403–3411, 2000. [322] P. C. Brooks, S. Silletti, T. L. Von Schalscha, M. Friedlander, [307] S. A. Karumanchi, V. Jha, R. Ramchandran et al., “Cell and D. A. Cheresh, “Disruption of angiogenesis by PEX, surface glypicans are low-affinity endostatin receptors,” a noncatalytic metalloproteinase fragment with integrin Molecular Cell, vol. 7, no. 4, pp. 811–822, 2001. binding activity,” Cell, vol. 92, no. 3, pp. 391–400, 1998. 24 Journal of Oncology [323] L. Bello, V. Lucini, G. Carrabba et al., “Simultaneous inhibi- integrin alpha(v)beta(3), + or - dacarbazine in patients with tion of glioma angiogenesis, cell proliferation, and invasion stage IV metastatic melanoma,” Cancer, vol. 116, no. 6, pp. by a naturally occurring fragment of human metallopro- 1526–1534, 2010. teinase-2,” Cancer Research, vol. 61, no. 24, pp. 8730–8736, [338] S. J. O’Day, A. C. Pavlick, M. R. Albertini et al., “Clinical and pharmacologic evaluation of two dose levels of intetumumab (CNTO 95) in patients with melanoma or angiosarcoma,” [324] J.-O. Nam, H.-W. Jeong, B.-H. Lee, R.-W. Park, and I.-S. Kim, “Regulation of tumor angiogenesis by fastatin, the fourth Investigational New Drugs. In press. FAS1 domain of βig-h3, via αvβ3 integrin,” Cancer Research, [339] J. A. Varner, M. T. Nakada, R. E. Jordan, and B. S. Coller, vol. 65, no. 10, pp. 4153–4161, 2005. “Inhibition of angiogenesis and tumor growth by murine [325] C. Mas-Moruno, F. Rechenmacher, and H. Kessler, “Cilengi- 7E3, the parent antibody of c7E3 Fab (abciximab; ReoPro),” Angiogenesis, vol. 3, no. 1, pp. 53–60, 1999. tide: the first anti-angiogenic small molecule drug candidate. Design, synthesis and clinical evaluation,” Anti-Cancer Agents [340] M. T. Nakada, G. Cao, P. M. Sassoli, and H. M. DeLisser, in Medicinal Chemistry, vol. 10, no. 10, pp. 753–768, 2010. “c7E3 Fab inhibits human tumor angiogenesis in a SCID mouse human skin xenograft model,” Angiogenesis, vol. 9, no. [326] K. E. Shannon, J. L. Keene, S. L. Settle et al., “Anti-metastatic properties of RGD-peptidomimetic agents S137 and S247,” 4, pp. 171–176, 2006. Clinical and Experimental Metastasis, vol. 21, no. 2, pp. 129– [341] F. Mitjans, T. Meyer, C. Fittschen et al., “In vivo therapy 138, 2004. of malignant melanoma by means of antagonists of αv [327] A. Abdollahi, D. W. Griggs, H. Zieher et al., “Inhibition of integrins,” International Journal of Cancer, vol. 87, no. 5, pp. 716–723, 2000. αvβ3 integrin survival signaling enhances antiangiogenic and antitumor effects of radiotherapy,” Clinical Cancer Research, [342] A. R. Reynolds, I. R. Hart, A. R. Watson et al., “Stimulation vol. 11, no. 17, pp. 6270–6279, 2005. of tumor growth and angiogenesis by low concentrations of RGD-mimetic integrin inhibitors,” Nature Medicine, vol. 15, [328] N. E. Tsopanoglou, M. E. Papaconstantinou, C. S. Flordellis, no. 4, pp. 392–400, 2009. and M. E. Maragoudakis, “On the mode of action of thrombin-induced angiogenesis: thrombin peptide, TP508, [343] D. Hanahan, “A flanking attack on cancer,” Nature Medicine, mediates effects in endothelial cells via α vβ3 integrin,” vol. 4, no. 1, pp. 13–14, 1998. Thrombosis and Haemostasis, vol. 92, no. 4, pp. 846–857, [344] R. K. Jain, “Normalization of tumor vasculature: an emerging concept in antiangiogenic therapy,” Science, vol. 307, no. 5706, pp. 58–62, 2005. [329] K. Meerovitch, F. Bergeron, L. Leblond et al., “A novel RGD antagonist that targets both αvβ3and α5β1 induces apoptosis [345] G. Huang and L. Chen, “Tumor vasculature and microenvi- of angiogenic endothelial cells on type I collagen,” Vascular ronment normalization: a possible mechanism of antiangio- Pharmacology, vol. 40, no. 2, pp. 77–89, 2003. genesis therapy,” Cancer Biotherapy and Radiopharmaceuti- [330] C. C. Kumar, M. Malkowski, Z. Yin et al., “Inhibition of cals, vol. 23, no. 5, pp. 661–667, 2008. angiogenesis and tumor growth by SCH 221153, a dual αvβ3 [346] G. Bergers and D. Hanahan, “Modes of resistance to anti- and αvβ5 integrin receptor antagonist,” Cancer Research, vol. angiogenic therapy,” Nature Reviews Cancer, vol. 8, no. 8, pp. 61, no. 5, pp. 2232–2238, 2001. 592–603, 2008. [331] L. Belvisi, T. Riccioni, M. Marcellini et al., “Biological [347] P. Fraisl, M. Mazzone, T. Schmidt, and P. Carmeliet, and molecular properties of a new αvβ3/αvβ5 integrin “Regulation of angiogenesis by oxygen and metabolism,” antagonist,” Molecular Cancer Therapeutics, vol. 4, no. 11, pp. Developmental Cell, vol. 16, no. 2, pp. 167–179, 2009. 1670–1680, 2005. [348] A. Rapisarda and G. Melillo, “Role of the hypoxic tumor [332] K. Minamiguchi, H. Kumagai, T. Masuda, M. Kawada, M. microenvironment in the resistance to anti-angiogenic thera- Ishizuka, and T. Takeuchi, “Thiolutin, an inhibitor of huvec pies,” Drug Resistance Updates, vol. 12, no. 3, pp. 74–80, 2009. adhesion to vitronectin, reduces paxillin in huvecs and [349] S. J. Lunt, N. Chaudary, and R. P. Hill, “The tumor microen- suppresses tumor cell-induced angiogenesis,” International vironment and metastatic disease,” Clinical and Experimental Journal of Cancer, vol. 93, no. 3, pp. 307–316, 2001. Metastasis, vol. 26, no. 1, pp. 19–34, 2009. [333] R. Soldi, S. Mitola, M. Strasly, P. Defilippi, G. Tarone, and [350] K. De Bock, S. Cauwenberghs, and P. Carmeliet, “Vessel F. Bussolino, “Role of α(v)β3 integrin in the activation abnormalization: another hallmark of cancer? Molecular of vascular endothelial growth factor receptor-2,” EMBO mechanisms and therapeutic implications,” Current Opinion Journal, vol. 18, no. 4, pp. 882–892, 1999. in Genetics and Development, vol. 21, no. 1, pp. 73–79, 2010. [334] J. C. Gutheil, T. N. Campbell, P. R. Pierce et al., “Targeted [351] A. R. Reynolds, “Potential relevance of bell-shaped and antiangiogenic therapy for cancer using vitaxin: a humanized u-shaped dose-responses for the therapeutic targeting of monoclonal antibody to the integrin α(v)β3,” Clinical Cancer angiogenesis in cancer,” Dose-Response, vol. 8, no. 3, pp. 253– Research, vol. 6, no. 8, pp. 3056–3061, 2000. 284, 2010. [335] D. G. McNeel, J. Eickhoff,F.T.Lee et al., “Phase Itrial of [352] S. De, O. Razorenova, N. P. McCabe, T. O’Toole, J. Qin, a monoclonal antibody specific for α vβ3 integrin (MEDI- and T. V. Byzova, “VEGF—integrin interplay controls tumor 522) in patients with advanced malignancies, including an growth and vascularization,” Proceedings of the National assessment of effect on tumor perfusion,” Clinical Cancer Academy of Sciences of the United States of America, vol. 102, Research, vol. 11, no. 21, pp. 7851–7860, 2005. no. 21, pp. 7589–7594, 2005. [336] D. Zhang, T. Pier, D. G. McNeel, G. Wilding, and A. Friedl, [353] G. H. Mahabeleshwar, J. Chen, W. Feng, P. R. Somanath, O. V. “Effects of a monoclonal anti-αvβ3 integrin antibody on Razorenova, and T. V. Byzova, “Integrin affinity modulation blood vessels—a pharmacodynamic study,” Investigational in angiogenesis,” Cell Cycle, vol. 7, no. 3, pp. 335–347, 2008. New Drugs, vol. 25, no. 1, pp. 49–55, 2007. [354] P. R. Somanath, A. Ciocea, and T. V. Byzova, “Integrin [337] P. Hersey, J. Sosman, S. O’Day et al., “A randomized phase and growth factor receptor alliance in angiogenesis,” Cell 2 study of etaracizumab, a monoclonal antibody against Biochemistry and Biophysics, vol. 53, no. 2, pp. 53–64, 2009. Journal of Oncology 25 [355] P. R. Somanath, N. L. Malinin, and T. V. Byzova, “Cooper- ation between integrin ανβ3 and VEGFR2 in angiogenesis,” Angiogenesis, vol. 12, no. 2, pp. 177–185, 2009. [356] A. Cretu, J. M. Roth, M. Caunt et al., “Disruption of endothe- lial cell interactions with the novel Hu177 cryptic collagen epitope inhibits angiogenesis,” Clinical Cancer Research, vol. 13, no. 10, pp. 3068–3078, 2007. [357] K. Chen and X. Chen, “Integrin targeted delivery of chemotherapeutics,” Theranostics, vol. 1, pp. 189–200, 2011. [358] Z. Wang, W. K. Chui, and P. C. Ho, “Integrin targeted drug and gene delivery,” Expert Opinion on Drug Delivery, vol. 7, no. 2, pp. 159–171, 2010. [359] K. N. Sugahara, T. Teesalu, P. Prakash Karmali et al., “Coad- ministration of a tumor-penetrating peptide enhances the efficacy of cancer drugs,” Science, vol. 328, no. 5981, pp. 1031–1035, 2010. [360] G. J. Strijkers, E. Kluza, G. A. F. Van Tilborg et al., “Param- agnetic and fluorescent liposomes for target-specific imaging and therapy of tumor angiogenesis,” Angiogenesis, vol. 13, no. 2, pp. 161–173, 2010. [361] A. J. Beer, H. Kessler, H. J. Wester, and M. Schwaiger, “PET Imaging of Integrin alphaVbeta3 expression,” Theranostics, vol. 1, pp. 48–57, 2011. [362] F. Kiessling, J. Gaetjens, and M. Palmowski, “Application of molecular ultrasound for imaging integrin expression,” Theranostics, vol. 1, pp. 127–134, 2011. [363] E. Mery, E. Jouve, S. Guillermet et al., “Intraoperative flu- orescence imaging of peritoneal dissemination of ovarian carcinomas. A preclinical study,” Gynecologic Oncology, vol. 122, no. 1, pp. 155–162, 2011. 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