ANTICANCER RESEARCH 32: 1791-1800 (2012)

Review The Interplay between Hemostasis and Malignancy: The Oral Cancer Paradigm

CHRISTOS YAPIJAKIS1,2, ATHANASIOS BRAMOS1, ALEXANDER MICHAEL NIXON1, VASSILIS RAGOS1 and ELEFTHERIOS VAIRAKTARIS1

1Department of Oral and Maxillofacial Surgery, University of Athens Medical School, Attikon Hospital, Athens, Greece; 2Department of Neurology, University of Athens Medical School, Eginition Hospital, Athens, Greece

Abstract. Accumulating evidence has revealed the role of and its cleavage to fibrin monomers result in the various components of the coagulatory system in different rapid formation of fibrin matrix. Furthermore, it is well stages of carcinogenesis including precancerous and initial documented that fibrinogen and cross-linked fibrin reside stages, tumor growth, , stroma generation, and inside the tumor stroma, facilitating its remodeling, metastasis of malignant cells. This comprehensive review angiogenesis, tumor growth and metastasis. In conclusion, discusses major points of evidence, in addition to recent the hemostatic system contributes to the development of the findings on specific factors associated with the paradigm of malignant phenotype acting on many different levels. oral squamous cell carcinoma. During carcinogenesis, angiogenesis is favored by local conditions of hypoxia, A bleeding diathesis and a systemic activation of the cell–to–cell interactions, and by expression of paracrine cascade contribute considerably to the morbidity growth factors and inflammatory cytokines. In the oral region and mortality of cancer patients (1-2). In some instances, the specifically, genetic association studies have revealed that disruption of hemostastic mechanisms, such as venous constitutively high gene expression of certain inflammatory thromboembolism, may even precede the clinical cytokines plays a major role in carcinogenesis. manifestations of malignancy (3). However, it is well (TF) has a physiological role in hemostasis, but it also established that the activation of coagulation is not merely an constitutes a notable procoagulant in many types of cancer, epiphenomenon of cancer (4-6). since it appears to be constitutively expressed by tumor cells. The present review discusses the role of various Furthermore, its pathway regulates mechanisms which components of the clotting system in different stages of involve and matrix metallo-proteinases, both of carcinogenesis, including precancerous and initial stages, which seem to be critical in oral carcinogenesis. Thrombin tumor growth, angiogenesis, stroma generation, and metastasis has a central role in hemostasis but it may also promote of malignant cells. Furthermore, specific findings of recent angiogenesis through pathways independently of fibrin genetic studies involving functional gene polymorphisms of generation. Thrombomodulin may act through attenuation of clotting system components, associated with the well studied the tumor-promoting properties of thrombin, but it also may paradigm of oral squamous cell carcinoma are presented. function as a cell–to–cell adhesion molecule, independently of its anticoagulant action. The activation of fibrinogen by Coagulatory System and Platelets

Coagulation is a complex cascade by which fibrin is produced forming the basis of a clot. Cancer-related Correspondence to: Professor Eleftherios Vairaktaris, MD, DDS, represents a complex imbalance of coagulation PhD. Department of Oral and Maxillofacial Surgery, University of and fibrinolysis, which also involves formation and Athens Medical School, Attikon Hospital, Rimini 1, Athens 12461, degradation of extracellular matrix (ECM) (Figure 1). As Greece. Tel: +30 2105831544, Fax: +30 2105831545, e-mail: new vessels within a tumor leak fibrin into the extravascular [email protected] space, they persistently activate the coagulatory system, both Key Words: Angiogenesis, carcinogenesis, fibrin, fibrinolysis, locally and systemically, producing the clinical appearance thrombin, thrombosis, review, tissue factor pathway. of a wound that does not heal (7).

0250-7005/2012 $2.00+.40 1791 ANTICANCER RESEARCH 32: 1791-1800 (2012)

Figure 1. Pathogenesis of the prothrombotic state in cancer patients (adapted from (1) with modifications). The central role of the tumor cell is illustrated, since: a) It directly interacts with platelets (P) inducing their aggregation; b) it interacts with vascular endothelial cells through several cytokines such as tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6) and vascular endothelial growth factor (VEGF), which induce the expression of tissue factor (TF) by the endothelial cells, as well as the release of von Willebrand’s factor (vWf), of leukocyte adhesion molecules, of platelet activating factor (PAF) and of plasminogen activator inhibitor type-1 (PAI-1), while at the same time down-regulating the expression of thrombomodulin, C (PC) receptor and tissue plasminogen activator (tPA); c) it stimulates leukocytes to produce TF and cytokines (TNF-α, IL-6); d) it directs the expression of procoagulants of TF, of cancer procoagulant (CP), of factor V receptor (FVr) and of mucin. The levels of IL-6, TNF-α, PAI-1 and VEGF (shown in bold) have been associated with carcinogenesis in the oral region by genetic association studies (12, 13, 42, 43).

The most essential components of the coagulation and Angiogenesis fibrinolysis systems are associated with the endothelial cell membrane, including tissue factor (TF), thrombin and The generation of new vessels from pre-existing vasculature urokinase receptors. They become exposed when vessels is an essential step for tumor survival, invariably observed in are injured and platelets bound to them at early stages of cancer pathology (8). Angiogenesis seems to be well either coagulation or inflammation. Consequently, platelets controlled during embryogenesis, the menstrual cycle, wound release vascular endothelial growth factor (VEGF) and healing, and inflammation, and a balance is eventually other growth factors into the circulation, thus attracting established between the various inhibitors and promoters of inflammatory cells to the site of injury (7). VEGF is known angiogenesis. In neoplasia, as the tumor develops, a pro- to promote both vascularization and tumor growth. On the angiogenic environment is favored by local conditions of other hand, in an effort to regulate the extent of tissue hypoxia, cell–to–cell interactions, expression of paracrine repair, platelets also contain hemostatic factors that inhibit growth factors and of inflammatory cytokines (9). This angiogenesis, such as plasminogen activator inhibitor-1 progressive nature of the tumor differs from the process of (PAI-1) (7). wound healing (10, 11).

1792 Yapijakis et al: Interplay between Hemostasis and Malignancy (Review)

Hemostatic mechanisms may directly or indirectly defective blood vessel development inside the yolk sac (18). influence angiogenesis through different pathways involving Similarly impaired vascular formation and death was the release of proangiogenic and antiangiogenic factors from observed in heterozygous VEGF-deficient embryos, pointing activated platelets and tumor cells, thus promoting the to the interplay between TF and VEGF (19). These non- formation of a fibrin-rich provisional ECM and thrombin hemostatic effects seem to be mediated through clotting- signaling via activation of G-protein-coupled dependent and -independent intracellular signaling pathways activated receptors on the endothelial cell surface (12). The that change the transcription rate of proangiogenic factors, interaction of angiogenesis-related factors, such as tumor especially of VEGF. In the clotting-dependent pathways TF, necrosis factor-alpha (TNF-α) with interleukins (IL) 6 and interacts with FVIIa, producing FXa and thrombin (Figure 1). 8, as well as their accumulated contribution to neoplasia have The secondary messengers involved in different cell types, been reported for oral cancer (13, 14). Functional DNA are intracellular calcium oscillations, activation of several polymorphisms affecting gene expression and the serum or mitogen-activating protein (MAP) kinases, transcription of the saliva levels of cytokines IL-6, IL-8, and TNF-α have genes such as EGR1 (early growth response protein 1) and been associated with increased risk for the development of activation of protease-activated receptors (PARs) (20). The oral squamous cell carcinoma (13, 14). There seems to be a latter are a novel family of seven transmembrane-spanning G- genetic predisposition for oral neoplasia in certain protein-coupled receptors up-regulated in various types of individuals who have genotypes which result in high cancer cells, including those of breast, prostate, colon and constitutive production of specific cytokines (13). According melanoma (21, 22). It is well documented that TF-FVIIa to these preliminary findings, such individuals may have 3- induces cell signaling via direct activation of PAR-2 that is 15 times higher risk of developing oral cancer than the independent of thrombin generation and that the ternary general population (13). complex of TF-FVIIa-FXa is more efficient than the binary complex, and activates both PAR-1 and PAR-2 (23, 24). The Tissue Factor TF cytoplasmic domain may exert a negative regulatory control over PAR-2 signaling. Genetic deletion of the TF cytoplasmic From the perspective of in vitro testing, the coagulatory domain in mice results in accelerated PAR-2-dependent cascade is artificially divided in the intrinsic or contact angiogenesis in synergy with the platelet-derived growth factor pathway and the extrinsic or tissue factor (TF) pathway (PDGF) but not with VEGF (25). In this context, cancer cells (Figure 1). There is a cross-talk between the two pathways expressing TF are likely to progressively regulate (through because both TF and thrombin can activate factor IX (FIX) distinct mechanisms) angiogenesis, primary tumor growth and (15). As the physiological initiator of coagulation, TF is metastasis via autocrine signaling (24, 26). In the coagulation- constitutively expressed on subendothelial fibroblasts and independent mechanisms, TF directly up-regulates the smooth muscle cells that are only exposed to blood upon expression of the proangiogenic factor VEGF and possibly vascular damage. TF is also expressed on peripheral blood other growth factors, whereas it diminishes the expression of monocytes and on vascular endothelia after exposure to the antiangiogenic peptide (27). In this case, activating or inflammatory stimuli such as endotoxin. the increased expression of VEGF is regulated via the Structurally, TF is a transmembrane glycoprotein which cytoplasmic tail of TF, with the use of protein kinase C (28) belongs to the class 2 cytokine-receptor superfamily (16). It and is independent of the TF ligand-binding extracellular has a 219-amino acid extracellular ligand-binding domain domain, as shown in some melanoma cell lines (29). which interacts with FVII/FVIIa, a 29-amino acid Since TF is expressed constitutively by tumor cells, in hydrophobic transmembrane region and a 21-amino acid contrast to normal tissues, it constitutes a notable intracellular tail (16). When plasma comes in contact with procoagulant in many types of cancer, including breast cells in extravascular sites that express TF, a complex is cancer (30), non-small lung cell carcinoma (31), head and formed on the cell surface with the circulating FVII, which is neck squamous cell carcinoma (32, 33), prostate carcinoma further activated to FVIIa by limited proteolysis. About 1% (34), malignant glioma (35), colon adenocarcinoma (36), and of FVII is already activated in circulation and can also bind pancreatic cancer (37). Other tumor types induce the directly to TF with high affinity (17). These initial TF/FVIIa expression of TF in host macrophages and endothelia (20). complexes are used for the downstream activation of The increased expression of TF appears to be the result of that will subsequently activate the bound FVII, oncogenic events occurring in cancer cells, caused by mutant forming a positive feedback loop. oncogenes K-RAS, EGFR (epidermal growth factor receptor), Beyond its physiological role in hemostasis, TF appears to PML-RARa (promyelocytic leukemia-retinoic acid receptor have important functions both in normal embryonic and alpha), as well as by the inactivation of the tumor suppressor tumor angiogenesis. The former has been demonstrated by the genes p53 or PTEN (38). In addition, high levels of TF (30), fact that TF-null mice die from abnormal circulation and and VEGF (39) seem to be associated with advanced cancer

1793 ANTICANCER RESEARCH 32: 1791-1800 (2012) and poor prognosis (36). TF aids in angiogenesis though tenase complex (complex of Ca2+ and factors VIIIa, IXa and induction of VEGF (40) and possibly in tumor invasion and X). This task is accomplished through binding of the TFPI/FXa metastasis by up-regulating the secretion of urokinase complex to the TF/FVIIa complex and by inactivating the latter plasminogen activator receptor (uPAR) (41). through this quaternary complex formation, thereby by shutting Interestingly, both VEGF and PAI-1 levels have been down the TF-induced tenase pathway (48). Both circulating associated with carcinogenesis in the oral region (42, 43). An rTFPI and tumor cell-associated TFPI reportedly reduced the association of functional DNA polymorphisms affecting the experimental lung metastasis of B16 murine melanoma by gene expression of VEGF and PAI-1 with increased risk for oral approximately 80% (49). cancer has been found (42, 43). A genetic predisposition for A second TF inhibitor, TFPI-2, which has similar overall development of early stages of oral neoplasia was detected in domain organization and considerable primary amino acid certain individuals with a genotype resulting in low production sequence homology to TFPI (50), is an emerging protease of VEGF (44). Those findings, in addition to the known limited inhibitor that seems to have an important role in normal ECM tumor-associated neo-angiogenesis in the oral mucosa because remodeling. Human TFPI-2 is synthesized by endothelia, of its rich vasculature, may indicate that the underlying fibroblasts and smooth muscle cells of the vasculature and is neoplasia mechanism might not involve angiogenesis but other then located in the subenthothelial ECM compartment. It VEGF-related functions such as thrombosis. On the other hand, directly inhibits plasmin and indirectly MMPs by inactivating individuals with a higher expressing genotype in the promoter serine proteases that convert proMMPs to their active form (51). region of the PAI-1 gene have a higher risk of developing early In this regard, TFPI-2 may reduce the invasive potential of stages of oral cancer (43). Possibly, increased levels of PAI-1 tumors. In the well studied paradigm of oral cancer, several promote initial development of oral cancer through regulation MMPs have been associated with tumor advancement of cell detachment by decreasing cellular adhesion. Yet, they (including MMP-1, -3, -7, -9 and -13) (52-56). In all cases, seem to have a protective role against later stages of malignancy functional polymorphisms in the promoter region of the MMP by interfering with the proteolytic activity of matrix genes, affecting their expression, seem to increase predisposition (MMPs), which remodel the tissue by of developing oral cancer in certain individuals (52-56). degrading the major components of the extracellular , The expression of TFPI-2 diminishes with the increasing including the type I collagen (43). degree of malignancy, as was demonstrated through There is interplay between TF and VEGF, one promoting immunohistochemical localization in a series of human the production of the other. Both are stimulated by hypoxia tumors (57). In addition to being anti-invasive, TFPI-2 has but different pathways are involved. VEGF can trigger TF in antiangiogenic properties because an anti-TFPI-2 IgG results endothelia (through EGR1, distinct from inflammatory in detachment of endothelia from the ECM (58). It also stimuli) (44), TF can also trigger the expression of VEGF in exhibits pro-apoptotic properties (59) and was recently various cells of tumor stroma and parenchyma, thereby recognized as a tumor-suppressor gene since its silencing is creating a vicious cycle (20). This close relationship is involved in melanoma metastasis (60). further indicated by the co-localization of TF and VEGF in various types of human cancer which correlates with Thrombin microvascular density (36, 45). Finally, recent reports underline the critical role that tumor cell-associated TF may Thrombin has a central role in hemostasis as it is the most have in metastatic spread, along with other hemostatic effective agonist for platelet activation and activates various components (46, 47). Through a fibrinogen- and-platelet zymogens and co-factors culminating in the formation of the dependent evasion of natural killer cell-mediated clearance fibrin clot. Thrombin acts on fribrinogen, factors XIII, V and of tumor cells, TF was linked with increased survival of VIII, platelet membrane GPV, protein S and C (61). It is micrometastases in the lung (47). generated by many different types of tumor cells suggesting The interplay of angiogenesis and hemostasis may impact additional tumor-promoting properties at different levels (11, progression of cancer locally and systemically. The more 12, 62). Like TF, thrombin may promote angiogenesis knowledge is accumulated to clarify these concepts the better through different ways, and independently of fibrin potential therapeutic management may be effected by combining generation. surgery, radiation, chemotherapy, and anti-angiogenic agents. Thrombin facilitates the detachment of endothelia by preventing their adhesion to collagen IV and to laminin TF Pathway Inhibitor (63). It also activates gelatinase A (MMP-2), which degrades collagen IV and releases tissue plasminogen (t-PA) Another protease inhibitor constitutively released from and PAI-1, further enhancing the migratory potential of endothelia is TF pathway inhibitor (TFPI). TFPI inhibits endothelia through basement membranes (64). Additional circulating FXa but mainly acts to down-regulate TF-induced important pro-angiogenic properties that thrombin exhibits

1794 Yapijakis et al: Interplay between Hemostasis and Malignancy (Review) on vascular cells, mediated through PARs, is the increased affecting the high activity of TAFI was significantly reduced expression of VEGF from platelets and tumor cells, and the in patients with oral cancer compared to controls (71). The up-regulation of VEGF receptors on endothelial cells (63- prophylactic effect of the increased TAFI activity might 65). The latter effect results in potentiating the action of result from reduction of plasmin and inhibition of VEGF, which is a specific endothelial cell mitogen and a extracellular matrix dissolution. key angiogenic factor (66). Moreover, thrombin increases the expression of ανβ3 and may serve as an additional ligand for it. Integrin ανβ3 is considered a marker of the angiogenic phenotype of After cleavage from thrombin, antithrombin undergoes a endothelial cells in tumor vessels and interacts with ECM change in its structural conformation, which lacks proteins, including fibrinogen, , vitronectin, anticoagulant but exhibits antiangiogenic activity and and thrombospondin. The binding to ανβ3 antitumor growth properties. The antiangiogenic antithrombin promotes cell attachment, migration, proliferation and prevents exerts its effects through down-regulation of the (67, 68). Endothelial cells cultured on plates coated proangiogenic sulfate proteoglycan in endothelial with thrombin had a lower level of apoptosis, suggesting a cells (80), and may globally alter gene expression in thrombin-induced ανβ3-dependent endothelial cell survival endothelium, possibly through a putative endothelial cell (67, 68). Several thrombin-related factors have already been ligand-receptor signaling mechanism (81). Therefore, it is a associated with oral carcinogenesis (42, 43, 69-71). potent inhibitor of VEGF and beta fibroblast growth factor Besides endothelial cells, thrombin has diverse actions on (bFGF)-induced endothelial cells proliferation and platelets, tumor cells and inflammatory cells that result in angiogenesis (82). A prelatent conformation of antithrombin, tumor progression. It activates platelets, leading to the obtained by heat treatment, which has undergone only limited aggregation and release of their granules which possess conformational change and is still able to bind heparin and numerous pro- and anti-angiogenic factors. Thrombin is a inhibit thrombin, also exhibits potent antiangiogenic and potent mitogen inducing the proliferation of smooth muscle tumor inhibitory activities (83). cells as well as this of tumor cells, while it increases their Recently, it has been postulated that the binding of heparin adhesiveness favoring the formation of tumor-platelet to the antiangiogenic cleaved or latent forms of antithrombin aggregates, migration and metastasis (24, 72). The is crucial for the expression of its antiangiogenic activity multifunctional role of this protease is further demonstrated (84). However, in oral cancer in particular, antithrombin may by the fact that it has antiangiogenic properties. Prothrombin not have any significant antitumor growth properties. This kringle 2 domain as well as two prothombin fragments (F1 might be due to the fact that the oral region has a rich and F2) generated during activation of thrombin, inhibit vasculature and that low VEGF levels (instead of high ones) angiogenesis (73, 74). are strongly associated with oral carcinogenesis (43). Finally, thrombin inhibits clot lysis and modulates fibrinolysis through activation of thrombin activatable Thrombomodulin fibrinolysis inhibitor (TAFI), but the role of this factor in malignancy is poorly understood. TAFI cleaves C-terminal Thrombomodulin is an endothelial cell surface-associated lysine residues from fibrin, thereby preventing plasminogen, protein that forms a complex with thrombin leading to plasmin and t-PA from binding to fibrin (75). High TAFI activation of protein C (Figure 1). Activated protein C antigen levels have been reported in lung cancer patients cleaves non-platelet-associated FVa and FVIIa, thereby (76), while reduced TAFI activity, but not TAFI antigen, have inactivating the respective tenase and prothrombinase been documented in patients with acute promyelocytic complexes, thus down-regulating thrombin formation. In leukemia (77). Deficiency of TAFI in mice injected with addition to this anticoagulant effect, thrombomodulin different cancer lines did not affect the primary tumor reduces fibrinolysis by activating TAFI in plasma (85). growth, the tumor architecture or the invasive potential (78). The role of thrombomodulin in cancer biology seems to be In regard to neovascularization, TAFI reduced the formation multifaceted. A correlation between reduced thrombomodulin of capillary-like tubular structures in vitro, in human expression and shorter survival or increased metastasis in a microvascular endothelial cells possibly via a thrombin- variety of human tumors has been demonstrated (86, 87) and independent pathway (79). it has been shown that in vitro thrombomodulin reduces Interestingly, while prothrombin has not been found to be tumor cell proliferation and invasion (88, 89). The synthesis associated with risk for oral squamous cell carcinoma, of thrombomodulin and protein C is increased by VEGF, increased TAFI activity seems to inhibit tumor progression while many inflammatory cytokines, such as IL-1 and TGF- by reducing plasminogen activation (42, 69, 71). A genetic β, down-regulate thrombomodulin (90). Therefore, association study showed that a DNA polymorphism thrombomodulin may act through attenuation of the tumor-

1795 ANTICANCER RESEARCH 32: 1791-1800 (2012) promoting properties of thrombin. Independently of its natural cadherin (101), on the endothelial surface and possibly trigger anticoagulant action however, thrombomodulin can function vascular development (102). The latter interaction may as a cell–to–cell adhesion molecule through a Ca2+-dependent account for the increased angiogenesis and tumor growth interaction of its lectin-like domain, which restrains tumor observed in mice lacking β3 or both β3 and β5 (103). growth in vivo (91). There are reports suggesting that Fibrin mesh is further implicated in metastasis. Aggregates thrombomodulin is an alternative useful clinical predictor of of tumor cells and platelets are rich in fibrin, enhancing tumor prognosis (87, 92), however, no genetic association of further platelet adhesion to circulating tumor cells and functional polymorphisms affecting the activity of endothelial cells, thereby, facilitating metastasis (104). Fibrin thrombomodulin, of FV or of IL-1 with oral carcinogenesis and fibrinogen, however, do not appear to play a critical role has been detected (69, 93, 94). in the growth of established metastases as demonstrated in studies of fibrinogen-deficient mice (105, 106). In lung Fibrinogen and Cross-linked Fibrin carcinoma and melanoma cell lines, fibrinogen deficiency strongly diminished, but did not prevent, the development of After activation by thrombin, fibrinogen is cleaved to fibrin lung metastases, while angiogenesis and tumor stroma monomers that are rapidly combined to form a fibrin matrix. formation was equivalent both in normal controls and in Thrombin also activates FXIII that converts the soluble fibrin deficient mice (105). These results indicate that fibrin into an insoluble fibrin polymer. In addition, by linking α2- deposition may not be essential for the tumor establishment plasmin inhibitor to fibrin activated FXIII protects the clot and show how complex the interactions between the various against fibrinolysis (95). components of the ECM and the tumor cells are, during Besides the critical role of fibrinogen in hemostasis, this tumor growth and invasion. In addition, fibrin, along with protein is also important in tumor biology. It is well platelets, shelters tumor cells from the host’s antitumor cell documented that fibrinogen and the cross-linked fibrin reside immune surveillance mechanisms (46). inside the tumor stroma. This deposition is indicative of the extravascular activation of the hemostatic mechanism, a Fibrinolysis process favored by the increased vascular permeability of the newly formed tumor vessels (8). The fibrin gel provides a The aforementioned provisional matrix finally undergoes provisional matrix, enriched in growth factors, that promotes remodeling and is transformed into mature connective angiogenesis and cancer cell growth, as well as provides a tissue stroma, which resembles the stroma of healing surface for the prothrombinase assembly, a function normally wounds in its constituent elements (8, 10). In fact, the carried out by platelets in intravascular clotting. Various deposition of fibrin gel in tumors is balanced between growth factors, such as VEGF, bFGF and insulin-like growth deposition from clotting activation and dissolution through factor-1 (IGF-1), are sequestered and protected from activation of the fibrinolytic pathways (8). Angiogenesis degradation in fibrin (20), and some tumor cells are capable involves a sequence of key events that include focal of controling their own microenviroment by endogenously detachment of endothelial cells from the basement synthesizing and secreting fibrinogen (96). membrane, localized proteolytic degradation of the In addition, fibrin can induce the expression of the basement membrane and ECM invasion, leading to proangiogenic IL-8 in vitro, in oral and pharyngeal tumor capillary tube formation and vascular remodeling (107). cells but not in a non-tumorigenic oral cell line, which points A breach of the basement membrane is the critical initial to the specificity of fibrin in eliciting this response (97). step in tumor invasion. Malignant cells along with stromal Furthermore, fibrin matrices support endothelial adhesion cells secrete a variety of proteases that degrade one or more and promote the migration and survival of various types of components of the basement membrane. Such are cells, including transformed cells, macrophages and the urokinase-type plasminogen activator (uPA) and the fibroblasts (98). Interestingly, genetic association studies MMPs (108). Fibrinolytic activity depends on the balance indicate that the thinner the fibrin polymer made by FXIII, between plasminogen activators, such as tPA and uPA, and the higher the risk of initiation of oral carcinogenesis, while their inhibitors, such as PAI-1 and α2-antiplasmin. In increased levels of IL-8 also promote formation of oral addition, the contact system activators, kallikrein, high- squamous cell carcinoma (70, 99). molecular-weight kininogen and FXIIa convert plasminogen Cells interact with fibrin via integrin binding sites located at to plasmin. Among plasminogen activators the one strongly the amino acids 95-98 (RGDF) and 572-575 (RGDS) of associated with tumor progression is uPA. Among the above human the fibrinogen (100), while other proteins found inside mentioned factors, genetic association studies have shown fibrin, such as fibronectin, also offer additional binding sites that PAI-1 is strongly associated with initial stages of oral (8). Non-integrin binding sites of fibrinogen, such as the N- carcinogenesis, while FXII seems to plays no role in mouth terminal β15-42 sequence, bind to vascular endothelial (VE)- tumors (42, 70).

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10 Dvorak HF: Tumors: wounds that do not heal. Similarities The Hemostatic System as a Target between tumor stroma generation and wound healing. N Engl for Anticancer Therapy J Med 315: 1650-1659, 1986. 11 Wojtukiewicz MZ, Sierko E and Rak J: Contribution of the hemostatic system to angiogenesis in cancer. Semin Thromb Τhe hemostatic system contributes to the development of the Hemost 30: 5-20, 2004. malignant phenotype acting on many different levels (Figure 12 Coughlin SR: Thrombin signalling and protease-activated 1). Common events in cancer biology, namely the activation receptors. Nature 407: 258-264, 2000. of oncogenes, the inactivation of tumor suppressor genes and 13 Vairaktaris E, Yapijakis C, Serefoglou Z, Avgoustidis D, the changes in tumor microenvironment, such as hypoxia, Critselis E, Spyridonidou S, Vylliotis A, Derka S, Vassiliou S, exposure to cytokines and altered cellular adhesion, Nkenke E and Patsouris E: Gene expression polymorphisms of culminate in the induction of TF, uPAR, PAI-1 and PARs interleukins-1beta, -4, -6, -8, -10, and tumor necrosis factors- alpha, -beta: regression analysis of their effect upon oral (38). The result is complex interactions affecting aggressive squamous cell carcinoma. J Cancer Res Clin Oncol 134: 821- local growth, cellular migration, invasion, angiogenesis, 832, 2008. metastasis and tumor coagulopathy. 14 Yapijakis C, Serefoglou Z, Vylliotis A, Nkenke E, Derka S, The relationship between hemostasis and carcinogenesis Vassiliou S, Avgoustidis D, Neukam FW, Patsouris E and may be exploited therapeutically. Several drugs affecting the Vairaktaris E: Association of polymorphisms in tumor necrosis coagulation process may have promising anticancer activity, factor alpha and beta genes with increased risk for oral cancer. including heparin, aspirin, warfarin, angiostatin, Anticancer Res 29(6): 2379-2386, 2009. 15 Gomez K and McVey JH: Tissue factor initiated blood and non-steroidal anti-inflammatory substances. coagulation. Front Biosci 11: 1349-1359, 2006. In conclusion, accumulating evidence over the past two 16 Peppelenbosch MP and Versteeg HH: Cell biology of tissue decades has illuminated the important role of various factor, an unusual member of the cytokine receptor family. components of the coagulatory system in different stages of Trends Cardiovasc Med 11: 335-339, 2001. carcinogenesis. Based on the paradigm of oral cancer, there 17 Nemerson Y and Repke D: Tissue factor accelerates the seems to be an emerging notion of genetic predisposition activation of coagulation factor VII: the role of a bifunctional towards the development of the malignant phenotype in some coagulation . Thromb Res 40: 351-358, 1985. individuals, whose genetically-determined level of 18 Carmeliet P, Mackman N, Moons L, Luther T, Gressens P, Van Vlaenderen I, Demunck H, Kasper M, Breier G, Evrard P, Müller constitutive gene expression leads to biologically important M, Risau W, Edgington T and Collen D: Role of tissue factor in levels of certain factors associated with both hemostasis and embryonic blood vessel development. Nature 383: 73-75, 1996. tumor progression, such as cytokines IL-6 and TNF-α. 19 Carmeliet P, Ferreira V, Breier G, Pollefeyt S, Kieckens L, Gertsenstein M, Fahrig M, Vandenhoeck A, Harpal K, References Eberhardt C, Declercq C, Pawling J, Moons L, Collen D, Risau W and Nagy A: Abnormal blood vessel development and 1 Elice F, Jacoub J, Rickles FR, Falanga A and Rodeghiero F: lethality in embryos lacking a single VEGF allele. Nature 380: Hemostatic complications of angiogenesis inhibitors in cancer 435-439, 1996b. patients. Am J Hematol 83: 862-870, 2008. 20 Fernandez PM, Patierno SR and Rickles FR: Tissue factor and 2 Sutherland DE, Weitz IC and Liebman HA: Thromboembolic fibrin in tumor angiogenesis. Semin Thromb Hemost 30: 31-44, complications of cancer: Epidemiology, pathogenesis, 2004. diagnosis, and treatment. Am J Hematol 72: 43-52, 2003. 21 Arora P, Ricks TK and Trejo J: Protease-activated receptor 3 Monreal M and Trujillo-Santos J: Screening for occult cancer signalling, endocytic sorting and dysregulation in cancer. J Cell in patients with acute venous thromboembolism. Curr Opin Sci 120: 921-928, 2007. Pulm Med 13: 368-671, 2007. 22 Yin YJ, Salah Z, Grisaru-Granovsky S, Cohen I, Even-Ram SC, 4 De Cicco M: The prothrombotic state in cancer: pathogenic Maoz M, Uziely B, Peretz T and Bar-Shavit R: Human mechanisms. Crit Rev Oncol Hematol 50: 187-196, 2004. protease-activated receptor 1 expression in malignant epithelia: 5 Falanga A, Panova-Noeva M and Russo L: Procoagulant a role in invasiveness. Arterioscler Thromb Vasc Biol 23: 940- mechanisms in tumour cells. Best Pract Res Clin Haematol 22: 944, 2003. 49-60, 2009. 23 Rao LV and Pendurthi UR: Tissue factor-factor VIIa signaling. 6 Franchini M, Montagnana M, Targher G, Manzato F and Lippi Arterioscler Thromb Vasc Biol 25: 47-56, 2005. G: Pathogenesis, clinical and laboratory aspects of thrombosis 24 Ruf W and Mueller BM: Thrombin generation and the pathogenesis in cancer. J Thromb Thrombolysis 24: 29-38, 2007. of cancer. Semin Thromb Hemost 32(S1): 61-68, 2006. 7 Nash GF, Walsh DC and Kakkar AK: The role of the 25 Belting M, Dorrell MI, Sandgren S, Aguilar E, Ahamed J, coagulation system in tumour angiogenesis. Lancet Oncol 2: Dorfleutner A, Carmeliet P, Mueller BM, Friedlander M and 608-613, 2001. Ruf W: Regulation of angiogenesis by tissue factor cytoplasmic 8 Dvorak HF: Rous-Whipple Award Lecture. How tumors make bad domain signaling. Nat Med 10: 502-509, 2004. blood vessels and stroma. Am J Pathol 162: 1747-1757, 2003. 26 Belting M, Ahamed J and Ruf W: Signaling of the tissue factor 9 Carmeliet P: Angiogenesis in health and disease. Nat Med 9: coagulation pathway in angiogenesis and cancer. Arterioscler 653-660, 2003. Thromb Vasc Biol 25: 1545-1550, 2005.

1797 ANTICANCER RESEARCH 32: 1791-1800 (2012)

27 Zhang Y, Deng Y, Luther T, Müller M, Ziegler R, Waldherr R, 41 Taniguchi T, Kakkar AK, Tuddenham EG, Williamson RC and Stern DM and Nawroth PP: Tissue factor controls the balance Lemoine NR: Enhanced expression of urokinase receptor of angiogenic and antiangiogenic properties of tumor cells in induced through the tissue factor-factor VIIa pathway in human mice. J Clin Invest 94: 1320-1327, 1994. pancreatic cancer. Cancer Res 58: 4461-4467, 1998. 28 Abe K, Shoji M, Chen J, Bierhaus A, Danave I, Micko C, 42 Vairaktaris E, Yapijakis C, Serefoglou Z, Vylliotis A, Ries J, Casper K, Dillehay DL, Nawroth PP and Rickles FR: Nkenke E, Wiltfang J, Derka S, Vassiliou S, Springer I, Kessler Regulation of vascular endothelial growth factor production and P and Neukam FW: Plasminogen activator inhibitor-1 angiogenesis by the cytoplasmic tail of tissue factor. Proc Natl polymorphism is associated with increased risk for oral cancer. Acad Sci USA 96: 8663-8668, 1999. Oral Oncol 42: 888-892, 2006. 29 Bromberg ME, Sundaram R, Homer RJ, Garen A and 43 Yapijakis C, Vairaktaris E, Vassiliou S, Vylliotis A, Nkenke E, Konigsberg WH: Role of tissue factor in metastasis: functions Nixon AM, Derka S, Spyridonidou S, Vorris E, Neukam F and of the cytoplasmic and extracellular domains of the molecule. Patsouris E: The low VEGF production allele of the +936C/T Thromb Haemost 82: 88-92, 1999. polymorphism is strongly associated with increased risk for oral 30 Ueno T, Toi M, Koike M, Nakamura S and Tominaga T: Tissue cancer. J Cancer Res Clin Oncol 133: 787-791, 2007. factor expression in breast cancer tissues: its correlation with 44 Mechtcheriakova D, Wlachos A, Holzmuller, Binder BR and prognosis and plasma concentration. Br J Cancer 83: 164-170, Hofer E: Vascular endothelial cell growth factor-induced tissue 2000. factor expression in endothelial cells is mediated by EGR-1. 31 Minamiya Y, Matsuzaki I, Sageshima, Saito H, Taguchi K, Blood 93: 3811-3823, 1999. Nakagawa T and Ogawa J: Expression of tissue factor mRNA 45 Shoji M, Hancock WW, Abe K, Micko C, Casper KA, Baine and invasion of blood vessels by tumor cells in non-small cell RM, Wilcox JN, Danave I, Dillehay DL, Matthews E, Contrino lung cancer. Surg Today 34: 1-5, 2004. J, Morrissey JH, Gordon S, Edgington TS, Kudryk B, Kreutzer 32 Buchs AE, Zehavi S, Sher O, Yeheskely E, Muggia-Sulam M, DL and Rickles FR: Activation of coagulation and angiogenesis Sherman Y and Rapoport MJ: Heparanase, galectin-3, and in cancer: immunohistochemical localization in situ of clotting tissue factor mRNA are expressed in benign neoplasms of the proteins and vascular endothelial growth factor in human thyroid. Endocrine 22: 81-84, 2003. cancer. Am J Pathol 152: 399-411, 1998. 33 Wojtukiewicz MZ, Zacharski LR, Rucinska M, Zimnoch L, 46 Palumbo JS, Talmage KE, Massari JV, La Jeunesse CM, Flick Jaromin J, Rózañska-Kudelska M, Kisiel W and Kudryk BJ: MJ, Kombrinck KW, Jirousková M and Degen JL: Platelets and Expression of tissue factor and tissue factor pathway inhibitor fibrin(ogen) increase metastatic potential by impeding natural in situ in laryngeal carcinoma. Thromb Haemost 82: 1659- killer cell-mediated elimination of tumor cells. Blood 105: 178- 1662, 1999. 185, 2005. 34 Abdulkadir SA, Carvalhal GF, Kaleem Z, Kisiel W, Humphrey 47 Palumbo JS, Talmage KE, Massari JV, La Jeunesse CM, Flick MJ, PA, Catalona WJ and Milbrandt J: Tissue factor expression and Kombrinck KW, Hu Z, Barney KA and Degen JL: Tumor cell- angiogenesis in human prostate carcinoma. Hum Pathol 31: associated tissue factor and circulating hemostatic factors cooperate 443-447, 2000. to increase metastatic potential through natural killer cell-dependent 35 Rong Y, Hu F, Huang R, Mackman N, Horowitz JM, Jensen RL, and -independent mechanisms. Blood 110: 133-141, 2007. Durden DL, Van Meir EG and Brat DJ: Early growth response 48 Crawley JT and Lane DA: The haemostatic role of tissue factor gene-1 regulates hypoxia-induced expression of tissue factor in pathway inhibitor. Arterioscler Thromb Vasc Biol 28(2): 233- glioblastoma multiforme through hypoxia-inducible factor-1- 242, 2008. independent mechanisms. Cancer Res 66: 7067-7074, 2006. 49 Amirkhosravi A, Meyer T, Chang JY, Amaya M, Siddiqui F, 36 Nakasaki T, Wada H, Shigemori C, Miki C, Gabazza EC, Nobori Desai H and Francis JL: Tissue factor pathway inhibitor reduces T, Nakamura S and Shiku H: Expression of tissue factor and experimental lung metastasis of B16 melanoma. Thromb vascular endothelial growth factor is associated with angiogenesis Haemost 87: 930-936, 2002. in colorectal cancer. Am J Hematol 69: 247-254, 2002. 50 Sprecher CA, Kisiel W, Mathewes S and Foster DC: Molecular 37 Khorana AA, Ahrendt SA, Ryan CK, Francis CW, Hruban RH, cloning, expression, and partial characterization of a second Hu YC, Hostetter G, Harvey J and Taubman MB: Tissue factor human tissue-factor-pathway inhibitor. Proc Natl Acad Sci USA expression, angiogenesis, and thrombosis in pancreatic cancer. 91: 3353-3357, 1994. Clin Cancer Res 13: 2870-2875, 2007. 51 Chand HS, Foster DC and Kisiel W: Structure, function and 38 Rak J, Milsom C, May L, Klement P and Yu J: Tissue factor in biology of tissue factor pathway inhibitor-2. Thromb Haemost cancer and angiogenesis: the molecular link between genetic 94: 1122-1130, 2005. tumor progression, tumor neovascularization, and cancer 52 Vairaktaris E, Serefoglou Z, Yapijakis C, Vylliotis A, Nkenke coagulopathy. Semin Thromb Hemost 32: 54-70, 2006. E, Derka S, Vassiliou S, Avgoustidis D, Neukam FW and 39 Altomare DF, Rotelli MT, Pentimone A, Rossiello MR, Patsouris E: High gene expression of matrix - Martinelli E, Guglielmi A, De Fazio M, Marino F, Memeo V, 7 is associated with early stages of oral cancer. Anticancer Res Colucci M and Semeraro N: Tissue factor and vascular 27(4B): 2493-2498, 2007. endothelial growth factor expression in colorectal cancer: relation 53 Vairaktaris E, Yapijakis C, Derka S, Serefoglou Z, Vassiliou S, with cancer recurrence. Colorectal Dis 9: 133-138, 2007. Nkenke E, Ragos V, Vylliotis A, Spyridonidou S, Tsigris C, 40 Ollivier V, Bentolila S, Chabbat J, Hakim J and de Prost D: Yannopoulos A, Tesseromatis C, Neukam FW and Patsouris E : Tissue factor-dependent vascular endothelial growth factor Association of matrix metalloproteinase-1 (-1607 1G/2G) production by human fibroblasts in response to activated factor polymorphism with increased risk for oral squamous cell VII. Blood 91: 2698-2703, 1998. carcinoma. Anticancer Res 27(1A): 459-464, 2007.

1798 Yapijakis et al: Interplay between Hemostasis and Malignancy (Review)

54 Vairaktaris E, Yapijakis C, Nkenke E, Serefoglou ZC, 69 Vairaktaris E, Yapijakis C, Wiltfang J, Ries J, Vylliotis A, Derka Chatzitheofylaktou A, Vassiliou S, Derka S, Vylliotis A, Perrea S, Vasiliou S and Neukam FW: Are factor V and prothrombin D, Neukam FW and Patsouris E: A metalloproteinase-13 mutations associated with increased risk of oral cancer? polymorphism affecting its gene expression is associated with Anticancer Res 25(3C): 2561-2565, 2005. advanced stages of oral cancer. Anticancer Res 27(6B): 4027- 70 Vairaktaris E, Vassiliou S, Yapijakis C, Spyridonidou S, 4030, 2007. Vylliotis A, Derka S, Nkenke E, Fourtounis G, Neukam FW 55 Vairaktaris E, Yapijakis C, Vasiliou S, Derka S, Nkenke E, and Patsouris E: Increased risk for oral cancer is associated Serefoglou Z, Vorris E, Vylliotis A, Ragos V, Neukam FW and with coagulation factor XIII but not with factor XII. Oncol Rep Patsouris E: Association of -1171 promoter polymorphism of 18: 1537-1543, 2007. matrix metalloproteinase-3 with increased risk for oral cancer. 71 Vairaktaris E, Yapijakis C, Nkenke E, Vassiliou S, Vylliotis A, Anticancer Res 27(6B): 4095-4100, 2007. Nixon AM, Derka S, Ragos V, Spyridonidou S, Tsigris C, 56 Vairaktaris E, Vassiliou S, Nkenke E, Serefoglou Z, Derka S, Neukam FW and Patsouris E: The 1040C/T polymorphism Tsigris C, Vylliotis A, Yapijakis C, Neukam FW and Patsouris influencing thermal stability and activity of thrombin E: A metalloproteinase-9 polymorphism which affects its activatable fibrinolysis inhibitor is associated with risk for oral expression is associated with increased risk for oral squamous cancer. Am J Hematol 82: 1010-1012, 2007. cell carcinoma. Eur J Surg Oncol 34: 450-455, 2008. 72 Nierodzik ML and Karpatkin S: Thrombin induces tumor growth, 57 Wojtukiewicz MZ, Sierko E, Zimnoch L, Kozlowski L and metastasis, and angiogenesis: Evidence for a thrombin-regulated Kisiel W: Immunohistochemical localization of tissue factor dormant tumor phenotype. Cancer Cell 10: 355-362, 2006. pathway inhibitor-2 in human tumor tissue. Thromb Haemost 73 Lee TH, Rhim T and Kim SS: Prothrombin kringle-2 domain 90: 140-146, 2003. has a growth inhibitory activity against basic fibroblast growth 58 Iino M, Foster DC and Kisiel W: Quantification and factor-stimulated capillary endothelial cells. J Biol Chem 273: characterization of human endothelial cell-derived tissue factor 28805-28812, 1998. pathway inhibitor-2. Arterioscler Thromb Vasc Biol 18: 40-46, 74 Rhim TY, Park CS, Kim E and Kim SS: Human prothrombin 1998. fragment 1 and 2 inhibit bFGF-induced BCE cell growth. 59 Tasiou A, Konduri SD, Yanamandra N, Dinh DH, Olivero WC, Biochem Biophys Res Commun 252: 513-516, 1998. Gujrati M, Obeyesekere M and Rao JS: A novel role of tissue 75 Bouma BN and Mosnier LO: Thrombin activatable fibrinolysis factor pathway inhibitor-2 in apoptosis of malignant human inhibitor (TAFI) – Ηow does thrombin regulate fibrinolysis? gliomas. Int J Oncol 19: 591-597, 2001. Ann Med 38: 378-388, 2006. 60 Nobeyama Y, Okochi-Takada E, Furuta J, Miyagi Y, Kikuchi K, 76 Hataji O, Taguchi O, Gabazza EC, Yuda H, D'Alessandro- Yamamoto A, Nakanishi Y, Nakagawa H and Ushijima T: Gabazza CN, Fujimoto H, Nishii Y, Hayashi T, Suzuki K and Silencing of tissue factor pathway inhibitor-2 gene in malignant Adachi Y: Increased circulating levels of thrombin-activatable melanomas. Int J Cancer 121: 301-307, 2007. fibrinolysis inhibitor in lung cancer patients. Am J Hematol 76: 61 Crawley JT, Zanardelli S, Chion CK and Lane DA: The central 214-219, 2004. role of thrombin in hemostasis. J Thromb Haemost 5(S1): 95- 77 Meijers JC, Oudijk EJ, Mosnier LO, Bos R, Bouma BN, 101, 2007. Nieuwenhuis HK and Fijnheer R: Reduced activity of TAFI 62 Zacharski LR, Memoli VA, Morain WD, Schlaeppi JM and (thrombin-activatable fibrinolysis inhibitor) in acute Rousseau SM: Cellular localization of enzymatically active promyelocytic leukaemia. Br J Haematol 108: 518-523, 2000. thrombin in intact human tissues by hirudin binding. Thromb 78 Reijerkerk A, Meijers JC, Havik SR, Bouma BN, Voest EE and Haemost 73: 793-797, 1995. Gebbink MF: Tumor growth and metastasis are not affected in 63 Tsopanoglou NE and Maragoudakis ME: On the mechanism of thrombin-activatable fibrinolysis inhibitor-deficient mice. J thrombin-induced angiogenesis. Potentiation of vascular Thromb Haemost 2: 769-779, 2004. endothelial growth factor activity on endothelial cells by up- 79 Guimaraes AH, Laurens N, Weijers EM, Koolwijk P, van regulation of its receptors. J Biol Chem 274: 23969-23976, 1999. Hinsbergh VW and Rijken DC: TAFI and pancreatic 64 Maragoudakis ME, Kraniti N, Giannopoulou E, Alexopoulos K carboxypeptidase B modulate in vitro capillary tube formation and Matsoukas J: Modulation of angiogenesis and progelatinase by human microvascular endothelial cells. Arterioscler Thromb a by thrombin receptor mimetics and antagonists. Endothelium Vasc Biol 27: 2157-2162, 2007. 8: 195-205, 2001. 80 Zhang W, Chuang YJ, Swanson R, Li J, Seo K, Leung L, Lau LF 65 Mohle R, Green D, Moore MA, Nachman RL and Rafii S: and Olson ST: Antiangiogenic antithrombin down-regulates the Constitutive production and thrombin-induced release of expression of the proangiogenic heparan sulfate proteoglycan, vascular endothelial growth factor by human megakaryocytes , in endothelial cells. Blood 103: 1185-1191, 2004. and platelets. Proc Natl Acad Sci USA 94: 663-668, 1997. 81 Zhang W, Chuang YJ, Jin T, Swanson R, Xiong Y, Leung L and 66 Ferrara N, Gerber HP and LeCouter J: The biology of VEGF Olson ST: Antiangiogenic antithrombin induces global changes and its receptors. Nat Med 9: 669-676, 2003. in the gene expression profile of endothelial cells. Cancer Res 67 Tsopanoglou NE, Andriopoulou P and Maragoudakis ME: On 66: 5047-5055, 2006. the mechanism of thrombin-induced angiogenesis: involvement 82 Zhang W, Swanson R, Xiong Y, Richard B and Olson ST: of αvβ3-integrin. Am J Physiol Cell Physiol 283: C1501-1510, Antiangiogenic antithrombin blocks the heparan sulfate-dependent 2002. binding of proangiogenic growth factors to their endothelial cell 68 Tsopanoglou NE and Maragoudakis ME: Role of thrombin in receptors: evidence for differential binding of antiangiogenic and angiogenesis and tumor progression. Semin Thromb Hemost anticoagulant forms of antithrombin to proangiogenic heparan 30: 63-69, 2004. sulfate domains. J Biol Chem 281: 37302-37310, 2006.

1799 ANTICANCER RESEARCH 32: 1791-1800 (2012)

83 Larsson H, Akerud P, Nordling K, Raub-Segall E, Claesson- 97 Lalla RV, Goralnick SJ, Tanzer ML and Kreutzer DL: Fibrin Welsh L and Björk I: A novel anti-angiogenic form of induces IL-8 expression from human oral squamous cell antithrombin with retained proteinase binding ability and carcinoma cells. Oral Oncol 37: 234-242, 2001. heparin affinity. J Biol Chem 276: 11996-20002, 2001. 98 Dvorak HF, Nagy JA, Berse B, Brown LF, Yeo KT, Yeo TK, 84 Zhang W, Swanson R, Izaguirre G, Xiong Y, Lau LF and Olson Dvorak AM, van de Water L, Sioussat TM and Senger DR: ST: The heparin- of antithrombin is crucial for Vascular permeability factor, fibrin, and the pathogenesis of antiangiogenic activity. Blood 106: 1621-1628, 2005. tumor stroma formation. Ann NY Acad Sci 667: 101-111, 1992. 85 Koutsi A, Papapanagiotou A and Papavassiliou AG: 99 Vairaktaris E, Yapijakis C, Serefoglou Z, Derka S, Vassiliou S, Thrombomodulin: From haemostasis to inflammation and Nkenke E, Vylliotis A, Wiltfang J, Avgoustidis D, Critselis E, tumourigenesis. Int J Biochem Cell Biol 40: 1669-1673, 2008. Neukam FW and Patsouris E: The interleukin-8 (-251A/T) 86 Hanly AM, Hayanga A, Winter DC and Bouchier-Hayes DJ: polymorphism is associated with increased risk for oral squamous Thrombomodulin: tumour biology and prognostic implications. cell carcinoma. Eur J Surg Oncol 33(4): 504-507, 2007. Eur J Surg Oncol 31: 217-220, 2005. 100 Henschen A: On the structure of functional sites in fibrinogen. 87 Hanly AM, Redmond M, Winter DC, Brophy S, Deasy JM, Thromb Res S5: 27-33, 1983. Bouchier-Hayes DJ and Kay EW: Thrombomodulin expression 101 Bach TL, Barsigian C, Yaen CH and Martinez J: Endothelial in colorectal carcinoma is protective and correlates with cell VE-cadherin functions as a receptor for the beta15-42 survival. Br J Cancer 94: 1320-1325, 2006. sequence of fibrin. J Biol Chem 273: 30719-30728, 1998. 88 Hosaka Y, Higuchi T, Tsumagari M and Ishii H: Inhibition of 102 Cavallaro U, Liebner S and Dejana E: Endothelial cadherins invasion and experimental metastasis of murine melanoma cells and tumor angiogenesis. Exp Cell Res 312: 659-667, 2006. by human soluble thrombomodulin. Cancer Lett 161: 231-240, 103 Reynolds LE, Wyder L, Lively JC, Taverna D, Robinson SD, 2000. Huang X, Sheppard D, Hynes RO and Hodivala-Dilke KM: 89 Zhang Y, Weiler-Guettler H, Chen J, Wilhelm O, Deng Y, Qiu F, Enhanced pathological angiogenesis in mice lacking beta3 Nakagawa K, Klevesath M, Wilhelm S, Böhrer H, Nakagawa integrin or beta3 and beta5 integrins. Nat Med 8: 27-34, 2002. M, Graeff H, Martin E, Stern DM, Rosenberg RD, Ziegler R 104 Biggerstaff JP, Seth N, Amirkhosravi A, Amaya M, Fogarty S, and Nawroth PP: Thrombomodulin modulates growth of tumor Meyer TV, Siddiqui F and Francis JL: Soluble fibrin augments cells independent of its anticoagulant activity. J Clin Invest 101: platelet/tumor cell adherence in vitro and in vivo, and enhances 1301-1309, 1998. experimental metastasis. Clin Exp Metastasis 17: 723-730, 90 Calnek DS and Grinnell BW: Thrombomodulin-dependent 1999. anticoagulant activity is regulated by vascular endothelial 105 Palumbo JS, Kombrinck KW, Drew AF, Grimes TS, Kiser JH, growth factor. Exp Cell Res 238: 294-298, 1998. Degen JL and Bugge TH: Fibrinogen is an important 91 Huang HC, Shi GY, Jiang SJ, Shi CS, Wu CM, Yang HY and determinant of the metastatic potential of circulating tumor Wu HL: Thrombomodulin-mediated cell adhesion: involvement cells. Blood 96: 3302-3309, 2000. of its lectin-like domain. J Biol Chem 278: 46750-46759, 2003. 106 Palumbo JS, Potter JM, Kaplan LS, Talmage K, Jackson DG 92 Iino S, Abeyama K, Kawahara K, Yamakuchi M, Hashiguchi T, and Degen JL: Spontaneous hematogenous and lymphatic Matsukita S, Yonezawa S, Taniguchi S, Nakata M, Takao S, metastasis, but not primary tumor growth or angiogenesis, is Aikou T and Maruyama I: The antimetastatic role of diminished in fibrinogen-deficient mice. Cancer Res 62: 6966- thrombomodulin expression in islet cell-derived tumors and its 6972, 2002. diagnostic value. Clin Cancer Res 10: 6179-6188, 2004. 107 Engelse MA, Hanemaaijer R, Koolwijk P and van Hinsbergh 93 Vairaktaris E, Serefoglou Z, Yapijakis C, Nkenke E, Vassiliou VW: The fibrinolytic system and matrix metalloproteinases in S, Spyridonidou S, Vylliotis A, Nixon AM, Neukam FW and angiogenesis and tumor progression. Semin Thromb Hemost Patsouris E: Coagulation-related factors, thrombomodulin and 30: 71-82, 2004. protein Z, are not associated with risk for oral cancer. 108 Rabbani SA: Metalloproteases and urokinase in angiogenesis Anticancer Res 27(4B): 2449-2451, 2007. and tumor progression. In Vivo 12: 135-42, 1998. 94 Vairaktaris E, Serefoglou Z, Yapijakis C, Stathopoulos P, Vassiliou S, Derka S, Nkenke E, Vylliotis A, Ragos V, Neukam FW and Patsouris E: The interleukin-1 beta gene polymorphism +3953 C/T is not associated with risk for oral cancer. Anticancer Res 27(6B): 3981-3986, 2007. 95 Mosesson MW: Fibrinogen and fibrin structure and functions. J Thromb Haemost 3: 1894-1904, 2005. 96 Sahni A, Simpson-Haidaris PJ, Sahni SK, Vaday GG and Francis CW: Fibrinogen synthesized by cancer cells augments Received February 13, 2012 the proliferative effect of FGF-2. J Thromb Haemost 6: 176- Revised April 5, 2012 183, 2008. Accepted April 6, 2012

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