thromboplastin and Neoplasm-Metastasis

Tissue factor (TF) is the primary initiator of the coagulation cascade, though its effects extend well beyond hemostasis. When TF binds to Factor VII, the resulting TF:FVIIa complex can proteolytically cleave transmembrane G protein-coupled protease-activated receptors (PARs). In addition to activating PARs, TF:FVIIa complex can also activate receptor tyrosine kinases (RTKs) and integrins. These signaling pathways are utilized by tumors to increase cell proliferation, angiogenesis, metastasis, and cancer stem-like cell maintenance. Herein, we review in detail the regulation of TF expression, mechanisms of TF signaling, their pathological consequences, and how it is being targeted in experimental cancer therapeutics.

Topics: Amino Acid Sequence; Cell Hypoxia; Factor VIIa; Gene Expression Regulation, Neoplastic; Humans; Immunotherapy, Adoptive; Integrins; Molecular Sequence Data; Molecular Targeted Therapy; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasm Proteins; Neoplasms; Neoplastic Stem Cells; Neovascularization, Pathologic; Protein Conformation; Protein Domains; Protein Isoforms; Receptor Protein-Tyrosine Kinases; Receptors, Proteinase-Activated; Signal Transduction; Thrombophilia; Thromboplastin

There is a strong relationship between tissue factor (TF) and cancer. Many cancer cells express high levels of both full-length TF and alternatively spliced (as) TF. TF expression in cancer is associated with poor prognosis. In this review, the authors summarize the regulation of TF expression in cancer cells and the roles of TF and asTF in tumor growth and metastasis. A variety of different signaling pathways, transcription factors and micro ribonucleic acids regulate TF gene expression in cancer cells. The TF/factor VIIa complex enhances tumor growth by activating protease-activated receptor 2 signaling and by increasing the expression of angiogenic factors, such as vascular endothelial growth factor. AsTF increases tumor growth by enhancing integrin β1 signaling. TF and asTF also contribute to metastasis via multiple thrombin-dependent and independent mechanisms that include protecting tumor cells from natural killer cells. Finally, a novel anticancer therapy is using tumor TF as a target to deliver cytotoxic drugs to the tumor. TF may be useful in diagnosis, prognosis, and treatment of cancer.

Topics: Gene Expression Regulation, Neoplastic; Humans; Immunoconjugates; Molecular Targeted Therapy; Neoplasm Metastasis; Neoplasms; Prognosis; Signal Transduction; Thromboplastin; Vascular Endothelial Growth Factor A

Venous thromboembolism (VTE) and cancer are strongly associated, and present a major challenge in cancer patient treatment. Cancer patients have a higher risk of developing VTE, although the risk differs widely between tumour types. VTE prophylaxis is routinely given to cancer patients, in the form of vitamin K antagonists (VKA) or low molecular weight heparin (LMWH). Several studies have reported that cancer patients receiving anticoagulants show prolonged survival and this effect was more pronounced in patients with a good prognosis, although the mechanism is poorly understood. Tissue Factor (TF) is the initiator of extrinsic coagulation, but its non-haemostatic signalling via protease-activated receptors (PARs) is a potent driver of tumour angiogenesis. Furthermore, coagulation activation is strongly implicated in tumour cell migration and metastasis. This review discusses the effects of anticoagulants on cancer progression in patients, tumour cell behaviour, angiogenesis, and metastasis in in vitro and in vivo models. Inhibition of TF signalling shows great promise in curbing angiogenesis and in vivo tumour growth, but whether this translates to patients is not yet known. Furthermore, non-haemostatic properties of coagulation factors in cancer progression are discussed, which provide exciting opportunities on limiting oncologic processes without affecting blood coagulation.

Topics: Animals; Anticoagulants; Blood Coagulation; Coumarins; Heparin, Low-Molecular-Weight; Humans; Neoplasm Metastasis; Neoplasms; Neovascularization, Pathologic; Signal Transduction; Thromboplastin; Venous Thromboembolism; Vitamin K

Cancer-associated venous thromboembolism (VTE) constitutes the second cause of death after cancer. Many risk factors for cancer-associated VTE have been identified, among them soluble tissue factor and microparticles (MPs). Few data are available about the implication of MPs in cancer associated-VTE through animal model of cancer. The objective of the present review was to report the state of the current literature about MPs and cancer-associated VTE in animal model of cancer. Fourteen series have reported the role of MPs in cancer-associated VTE, through three main mouse models: ectopic or orthotopic tumor induction, experimental metastasis by intravenous injection of tumor cells into the lateral tail vein of the mouse. Pancreatic cancer is the most used animal model, due to its high rate of cancer-associated VTE. All the series reported that tumor cell-derived MPs can promote thrombus formation in TF-dependent manner. Some authors reported also the implication of phosphatidylserine and PSGL1 in the generation of thrombin. Moreover, MPs seem to be implicated in cancer progression through a coagulation-dependent mechanism secondary to thrombocytosis, or a mechanism implicating the regulation of the immune response. For these reasons, few authors have reported that antiplatelet and anticoagulant treatments may prevent tumor progression and the formation of metastases in addition of coagulopathy.

Topics: Animals; Anticoagulants; Blood Coagulation; Cell-Derived Microparticles; Disease Models, Animal; Disease Progression; Humans; Neoplasm Metastasis; Neoplasms; Platelet Aggregation Inhibitors; Risk Factors; Thromboplastin; Thrombosis; Venous Thromboembolism

Tissue Factor (TF) is an evolutionary conserved glycoprotein, which is of immense importance for a variety of biologic processes. TF is expressed in two naturally occurring protein isoforms, membrane-bound "full-length" (fl)TF and soluble alternatively spliced (as)TF. The TF isoform expression is differentially modulated on post-transcriptional level via regulatory factors, such as serine/arginine-rich (SR) proteins, SR protein kinases and micro (mi)RNAs. Both isoforms mediate a variety of physiologic- and pathophysiologic-relevant functions, such as thrombogenicity, angiogenesis, cell signaling, tumor cell proliferation and metastasis. In this review, we will depict the main mechanisms regulating the TF isoform expression in cancer and under other pathophysiologic-relevant conditions. Moreover, we will summarize and discuss the latest findings regarding the role of TF and its isoforms in cancer biology.

Topics: Alternative Splicing; Animals; Gene Expression Regulation, Neoplastic; Humans; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasms; Neovascularization, Pathologic; Protein Isoforms; RNA Interference; RNA Processing, Post-Transcriptional; Signal Transduction; Thromboplastin

Blood coagulation is one of the most profound factors that influence cancer progression. Especially the initiator of coagulation, tissue factor (TF), has been subject to many studies investigating the overlap between coagulation and cancer. It has been known for decades that TF is a risk factor for metastasis, and in mouse models, TF drives metastasis in a coagulation-dependent manner. However, TF also serves as a cellular receptor to drive primary tumor growth and tumor angiogenesis. Nevertheless, recent studies have indicated that TF plays more fundamental roles in cancer biology. TF regulates tumor cell dormancy, is associated with cancer stem cell behavior, epithelial-to-mesenchymal transition, and dictates establishment of the tumor cell premetastatic niche. Especially with regard to these recent roles attributed to TF, no clear idea exists on the exact molecular pathways that are initiated by TF. Finally, TF alternative splicing results in an isoform with different characteristics and functions in cancer. In this review, a summary will be given on both the established as well as the new aspects of TF function in cancer progression.

Topics: Alternative Splicing; Animals; Blood Coagulation; Gene Expression Regulation, Neoplastic; Humans; Mice; Neoplasm Metastasis; Neoplasm Proteins; Neoplasms; Neoplastic Stem Cells; Neovascularization, Pathologic; Protein Isoforms; Stem Cell Niche; Thromboplastin; Tumor Microenvironment

Although many studies have demonstrated that components of the hemostatic system may be involved in signaling leading to cancer progression, the potential mechanisms by which they contribute to cancer dissemination are not yet precisely understood. Among known coagulant factors, tissue factor (TF) and thrombin play a pivotal role in cancer invasion. They may be generated in the tumor microenvironment independently of blood coagulation and can induce cell signaling through activation of protease-activated receptors (PARs). PARs are transmembrane G-protein-coupled receptors (GPCRs) that are activated by a unique proteolytic mechanism. They play important roles in vascular physiology, neural tube closure, hemostasis, and inflammation. All of these agents (TF, thrombin, PARs-mainly PAR-1 and PAR-2) are thought to promote cancer invasion and metastasis at least in part by facilitating tumor cell migration, angiogenesis, and interactions with host vascular cells, including platelets, fibroblasts, and endothelial cells lining blood vessels. Here, we discuss the role of PARs and their activators in cancer progression, focusing on TF- and thrombin-mediated actions. Therapeutic options tailored specifically to inhibit PAR-induced signaling in cancer patients are presented as well.

Topics: Blood Platelets; Endothelial Cells; Enzyme Activation; Humans; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasms; Neovascularization, Pathologic; Receptor, PAR-1; Receptor, PAR-2; Thrombin; Thromboplastin; Tumor Microenvironment

Inhibition of coagulation greatly limits cancer metastasis in many experimental models. Cancer cells trigger coagulation, through expression of tissue factor or P-selectin ligands that have correlated with worse prognosis in human clinical studies. Cancer cells also affect coagulation through expression of thrombin and release of microparticles that augment coagulation. In the cancer-bearing host, coagulation facilitates tumour progression through release of platelet granule contents, inhibition of Natural Killer cells and recruitment of macrophages. We are revisiting this literature in the light of recent studies in which treatment of clinical cohorts with anticoagulant drugs led to diminished metastasis.

Topics: Animals; Anticoagulants; Blood Coagulation; Blood Platelets; Cysteine Endopeptidases; Cytoplasmic Granules; Hirudins; Humans; Killer Cells, Natural; Lung Neoplasms; Macrophages; Mice; Neoplasm Metastasis; Neoplasm Proteins; Neoplasms, Experimental; Neoplastic Cells, Circulating; Neuraminidase; P-Selectin; Platelet Activation; Platelet Aggregation Inhibitors; Rats; Thrombin; Thrombophilia; Thromboplastin

Topics: Blood Coagulation; Gene Expression Regulation, Neoplastic; Lung Neoplasms; Neoplasm Metastasis; Thromboplastin

Topics: Anticoagulants; Antineoplastic Agents; Blood Coagulation Factors; Clinical Trials as Topic; Disease Progression; Glucuronidase; Heparin; Heparin, Low-Molecular-Weight; Humans; Neoplasm Metastasis; Neoplasms; Neovascularization, Pathologic; Selectins; Thrombin; Thromboembolism; Thrombophilia; Thromboplastin

Diverse oncogenic transformations result in the constitutive expression of tissue factor (TF) in cancer cells. The local and systemic activation of the coagulation cascade has long been a recognized hallmark for aggressive cancer, but genetic mouse models and new experimental therapeutics have only recently demonstrated crucial roles for TF initiated cell signaling in the pathogenesis of cancer. On tumor cells, the TF-VIIa binary complex mediates activation of protease activated receptor (PAR) 2 and thereby shapes the tumor microenvironment by inducing an array of proangiogenic and immune modulating cytokines, chemokines, and growth factors. PAR2 also uniquely triggers tumor cell migration by G protein-independent pathways through beta-arrestin scaffolding. Metastatic tumor cells use additional signaling networks of the coagulation cascade by activating PAR1 through thrombin or the ternary TF-VIIa-Xa signaling complex in the vascular and potentially lymphatic system. Selective antagonists of TF-VIIa-PAR2 signaling may be used as antiangiogenic therapy without increasing the risk of bleeding, whereas coagulation and associated signaling pathways on platelets and other host cells may be targeted for therapeutic benefit in advanced cancer and metastatic disease.

Topics: Animals; Blood Coagulation; Factor VIIa; Female; Humans; Male; Mice; Models, Biological; Neoplasm Metastasis; Neoplasms; Receptor, PAR-2; Signal Transduction; Thrombin; Thromboplastin

The activation of the coagulation cascade in the tumor microenvironment is a key feature of advanced malignancies. On tumor cells, tissue factor (TF) plays a central role to initiate cross-talk through the release of procoagulant microparticles or through direct, protease-activated receptor (PAR)-mediated cell signaling that leads to the production of soluble cytokines and angiogenic growth factors. In addition, the hemostatic system in the host compartment sustains crucial circuits that promote metastasis and support tumor growth and angiogenesis. Experimental tumor and genetic models have defined specific pathways that are supported by tumor cell and host TF and have identified potential therapeutic modalities to specifically interrupt TF signaling in tumor biology without impairment of hemostatic functions.

Topics: Animals; Anticoagulants; Blood Coagulation; Colorectal Neoplasms; Disease Progression; Factor VIIa; Humans; Mice; Neoplasm Metastasis; Neoplasms; Receptor, PAR-1; Receptor, PAR-2; Signal Transduction; Thromboplastin

There is a persuasive body of evidence suggesting that tissue factor (TF) is a major determinant of tumor progression. In addition to its "traditional" function as the initiator of hemostasis, TF may support tumor progression through signaling mechanisms involving either direct signal transduction through the TF cytoplasmic domain or TF:F VIIa-mediated and TF/F VIIa/F Xa-mediated activation of protease-activated receptors. Whereas TF-mediated signaling events uncoupled from hemostasis may play an important role in tumor dissemination in some contexts, TF-mediated thrombin generation appears to be the major mechanism linking tumor cell-associated TF to metastasis. At least one mechanism coupling thrombin generation to metastatic potential involves the most distal components of the hemostatic system (i.e., platelets, fibrinogen, and factor XIII) and leads to a restriction in natural killer cell-mediated lysis of newly formed micrometastases. A detailed understanding of the mechanisms linking TF and circulating hemostatic system components to tumor progression may lead to novel therapeutic targets for cancer treatment.

Topics: Animals; Hemostasis; Humans; Mice; Neoplasm Metastasis; Signal Transduction; Thromboplastin

Tissue factor (TF) is the primary initiator of the coagulation cascade and plays an essential role in hemostasis. TF also contributes to many diseases, including cancer. The correlation between thrombosis and cancer has been recognized for more than a century. However, it is only in the past two decades that we have begun to understand the role of TF in tumor biology. TF expression is upregulated on both tumor and host cells in cancer patients as well as in the circulation. Clinical observations indicate a direct correlation between the levels of tumor cell TF expression and poor prognosis for cancer patients. The role of TF in tumor biology has been extensively studied using various mouse tumor models. It has been demonstrated that tumor cell TF contributes to tumor metastasis, growth, and angiogenesis. The role of host TF in tumor progression is less clear. Recently developed mouse models with altered levels of TF may be useful in further analysis of the role of host cell TF in cancer.

Topics: Animals; Disease Models, Animal; Mice; Neoplasm Metastasis; Neoplasm Transplantation; Neoplasms; Neovascularization, Pathologic; Thromboplastin; Up-Regulation

Heparanase is an endo-beta- D-glucuronidase that is capable of cleaving heparan sulfate side chains of heparan sulfate proteoglycans on cell surfaces and the extracellular matrix, activity that is strongly implicated in tumor metastasis and angiogenesis. Evidence was provided that heparanase overexpression in human leukemia, glioma, and breast carcinoma cells results in a marked increase in tissue factor (TF) levels. Likewise, TF was induced by exogenous addition of recombinant heparanase to tumor cells and primary endothelial cells, induction that was mediated by p38 phosphorylation and correlated with enhanced procoagulant activity. TF induction was further confirmed in heparanase-overexpressing transgenic mice and correlated with heparanase expression levels in leukemia patients. Heparanase was also found to be involved in the regulation of tissue factor pathway inhibitor (TFPI). It was shown that heparanase overexpression or exogenous addition induces two- to threefold increase of TFPI expression. Similarly, heparanase stimulated accumulation of TFPI in the cell culture medium. Extracellular accumulation exceeded, however, the observed increase in TFPI at the protein level and appeared to be independent of heparan sulfate and heparanase enzymatic activity. Instead, a physical interaction between heparanase and TFPI was demonstrated, suggesting a mechanism by which secreted heparanase interacts with TFPI on the cell surface, leading to dissociation of TFPI from the cell membrane and increased coagulation activity, thus further supporting the local prothrombotic function of heparanase. As heparins are strong inhibitors of heparanase, in view of the effect of heparanase on TF/TFPI pathway, the role of heparins' anticoagulant activity may potentially be expanded.

Topics: Breast Neoplasms; Cell Line, Tumor; Endothelial Cells; Glucuronidase; Heparin; Humans; Lipoproteins; Neoplasm Metastasis; Neoplasms; Neovascularization, Pathologic; Thromboplastin

The clinical link between cancer and thrombosis has been recognized by Armand Trousseau in 1865. It has now become clear that clotting activation in malignancy not only plays an important role in the evolution of venous thromboembolism (VTE) or systemic coagulation disorders such as disseminated intravascular coagulation, but that multiple components of the haemostatic and fibrinolytic systems are directly involved in tumour progression. In particular, tissue factor (TF) appears to be involved in several pathways relevant to cancer growth and metastasis. Increasing evidence emerges that haemostatic perturbances in cancer patients are, at least in part, controlled by defined genetic events in molecular tumourigenesis including activating and inactivating mutations of oncogenes and tumour suppressor genes, respectively. Long-term therapy with low-molecular-weight heparin (LMWH) is considered as standard treatment for cancer-associated VTE. However, several experimental studies and clinical trials suggest that LMWH may also be beneficial as an adjunct in the treatment of patients with malignant disease. This article provides an overview on the significance, pathogenesis and treatment of cancer-related clotting disorders as well as on the cellular and molecular mechanisms, by which haemostatic components such as TF, platelets and fibrin(ogen) drive tumour progression.

Topics: Hemostasis; Hemostatic Disorders; Humans; Neoplasm Metastasis; Neoplasms; Thromboplastin; Thrombosis; Venous Thromboembolism

Tissue factor (TF), the key regulator of haemostasis and angiogenesis, is also involved in the pathology of several diseases, including cardiovascular, inflammatory and neoplastic conditions. In the latter, TF is upregulated by cancer cells, as well as by certain host cells, and it is the interactions between these distinct pools of TF-expressing cells that likely influence tumour progression in several ways. Furthermore, the release of TF microparticles into the circulation is thought to contribute to the systemic coagulopathies commonly observed in cancer patients. The direct regulation of TF by oncogenic events has provided a plausible explanation for the relatively common overexpression of TF in various cancers and its involvement in tumour growth, angiogenesis, metastasis and coagulopathy. However, this constitutive influence is modified by the tumour microenvironment, cellular interactions and host factors rendering TF expression patterns complex and heterogeneous. It appears that in many biological contexts TF plays a central role in disease progression and thereby potentially constitutes an attractive therapeutic target, especially in scenarios where the risk of bleeding can be avoided by selecting appropriate medications, refined dosing or by targeting the signalling component of TF activity. The efficacy and safety of such approaches still awaits clinical verification.

Topics: Animals; Antineoplastic Agents; Blood Coagulation Factors; Blood Platelets; Cell Transformation, Neoplastic; Cell-Derived Microparticles; Drug Delivery Systems; Genes, Tumor Suppressor; Hemostasis; Humans; Mice; Mice, Knockout; Models, Biological; Neoplasm Metastasis; Neoplasm Proteins; Neoplasms; Neovascularization, Pathologic; Oncogenes; Organ Specificity; Platelet Activation; Thrombophilia; Thromboplastin

Clotting activation occurs frequently in cancer. Tissue factor (TF), the most potent initiator of coagulation, is expressed aberrantly in many types of malignancy and is involved not only in tumor-associated hypercoagulability but also in promoting tumor angiogenesis and metastasis via coagulation-dependent and coagulation-independent (signaling) mechanisms. Tissue factor pathway inhibitor (TFPI) is the natural inhibitor of TF coagulant and signaling activities. Studies have shown that TFPI exhibits antiangiogenic and antimetastatic effects in vitro and in vivo. In animal models of experimental metastasis, both circulating and tumor cell-associated TFPI are shown to significantly reduce tumor cell-induced coagulation activation and lung metastasis. Heparins and heparin derivatives, which induce the release of TFPI from the vascular endothelium, also exhibit antitumor effects, and TFPI may contribute significantly to those effects. Indeed, a non-anticoagulant low-molecular-weight heparin with intact TFPI-releasing capacity has been shown to have significant antimetastatic effect in a similar experimental mouse model. The evidence supporting the dual inhibitory functions on TF-driven coagulation and signaling strengthen the rationale for considering TFPI as a potential anticancer agent. This article primarily summarizes the evidence for antiangiogenic and antimetastatic effects of TFPI and describes its potential mechanisms of action. The possible application of TFPI and other inhibitors of TF as potential anticancer agents is described, and information regarding potential antitumor properties of TFPI-2 (which has structural similarities to TFPI) is also included.

Topics: Animals; Anticoagulants; Fibrinolytic Agents; Hemostasis; Heparin, Low-Molecular-Weight; Humans; Lipoproteins; Neoplasm Metastasis; Neoplasms; Neovascularization, Pathologic; Serine Proteinase Inhibitors; Thromboplastin

Angiogenesis, the growth of new blood vessels from preexisting ones, is a necessary component of embryogenesis, wound healing, and the proliferative phase of the female reproductive cycle. Angiogenesis also plays a critical role in important pathologic processes such as cancer and cardiovascular complications. In addition, clinical, laboratory, and pharmacologic evidence has shown a link between angiogenesis, coagulation, hemostasis, and thrombosis in the settings of these pathologies. Recent studies in our laboratory revealed that thrombin has a significant stimulatory effect on angiogenesis. This effect of thrombin is independent of fibrin formation and can be attributed mainly to the activation of protease-activated receptor-1 (PAR-1). PAR-1 is widely expressed in vascular cells and is involved in cardiovascular complications such as atherosclerosis, restenosis, and neointimal formation. It is also expressed in many cancer cells contributing to induction of tumor growth and metastasis. In this review, we will summarize our present-day understanding of the role of thrombin and PAR-1 in angiogenesis and the potential therapeutic utility of targeting PAR-1 in angiogenesis-related disease, such as atherosclerosis, restenosis, and cancer.

Topics: Animals; Apoptosis; Blood Coagulation; Blood Vessels; Cardiovascular Diseases; Endothelium, Vascular; Humans; Neoplasm Metastasis; Neoplasms; Neovascularization, Pathologic; Neovascularization, Physiologic; Receptor, PAR-1; Thrombin; Thromboplastin

Tissue factor (TF), the primary cellular initiator of blood coagulation, is also involved in cancer-related processes such as hypercoagulability (Trousseau syndrome), tumor growth, angiogenesis, and metastasis. Indeed, elevated TF expression by cancer cells and their associated endothelial cells has been reported frequently. Oncogenic events in cancer cells (e.g., expression of mutant K- ras, EGFR, PTEN or p53) lead to an increase in TF levels and activity, and thereby promote tumor aggressiveness, angiogenesis, and hypercoagulability. Like TF, thrombin receptor (protease-activated receptor-1) is also upregulated in cancer cells expressing oncogenic K -ras. Pharmacological antagonists of some of these transforming genes (e.g., epidermal growth factor receptor inhibitors) could diminish TF expression, both locally and systemically, and hence these targeted agents could be viewed as potential indirect and cancer-specific anticoagulants, in addition to their direct antitumor effects. We postulate that levels of circulating TF may be useful in monitoring the biological activity of these agents. Although TF is essential for vascular development, its expression by tumor-associated endothelium appears to play a subtle and seemingly dispensable role. Thus, TF is a pivotal element of the tumor-vascular interface, is involved in many cancer-related processes, and may well constitute a promising new target for anticancer combination therapies in some disease settings.

Topics: Animals; Blood Coagulation; Disease Progression; Enzyme Inhibitors; Gene Expression Regulation, Neoplastic; Humans; Neoplasm Metastasis; Neoplasm Proteins; Neoplasms; Neovascularization, Pathologic; Signal Transduction; Thrombin; Thromboplastin

Advanced cancer is associated with a hypercoagulable state that is triggered by tissue factor (TF). TF-initiated thrombin generation is crucial for metastasis through fibrin and platelet deposition, as well as thrombin-dependent protease-activated receptor (PAR) 1 signaling. Surprisingly, PAR2, which is not cleaved by thrombin, appears to cosignal with PAR1 to elicit thrombin effects in metastatic tumor cells. In contrast to TF-driven thrombin pathways in metastasis, direct TF signaling plays a role in angiogenesis-dependent tumor growth. In TF cytoplasmic-domain-deleted mice, PAR2-dependent angiogenesis and tumor growth is enhanced, demonstrating a role for host cell TF signaling. In tumor cells, TF-factor VIIa (FVIIa) activates PAR2 and thereby regulates proangiogenic growth factor expression as well as integrins involving crosstalk with the TF cytoplasmic domain. In addition to thrombin-PAR signaling in metastasis and TF-FVIIa-PAR2 signaling in tumor growth, it is likely that additional protease pathways will prove to be crucial activators of PARs in cancer. Transmembrane serine proteases as well as matrix metalloproteinase are prime candidates for accessory pathways to regulate metastasis, tumor expansion, and angiogenesis dependent on specific features of the local tumor microenvironment.

Topics: Animals; Humans; Neoplasm Metastasis; Neoplasms; Neovascularization, Pathologic; Receptors, Proteinase-Activated; Thrombin; Thromboplastin

In the last few years, it has become clear that the processes of tumor angiogenesis, metastasis and invasiveness are highly dependent on components of the blood coagulation cascade. One of the key proteins in coagulation is tissue factor (TF). In addition, TF is also known as a mediator of intracellular signaling events that can alter gene expression patterns and cell behavior. TF significantly participates in tumor-associated angiogenesis and its expression levels have been correlated with the metastatic potential of many types of hematological malignancies. Signaling pathways initiated by both, tissue factor-activated factor VII (TF-FVIIa) protease activation of protein-activated receptors (PARs), and phosphorylation of the TF-cytoplasmic domain, appear to regulate these tumoral functions. Advances in antiangiogenic therapies and preclinical studies with TF-targeted therapeutics are hopeful in the control of tumor growth and metastasis, but continued studies on the regulation of TF are still needed. In the last few years, the use of approaches of functional genomics and proteomics has allowed the discovery of new proteins involved in the origin of the neoplasia and their participation in the development of the disease. This review attempts to establish a cellular and molecular causal link between cancer coagulopathy, angiogenesis and tumor progression in hematological malignancies.

Topics: Animals; Disease Progression; Hematologic Neoplasms; Humans; Leukemia; Neoplasm Metastasis; Neovascularization, Pathologic; Proteomics; Signal Transduction; Thromboplastin; Vascular Endothelial Growth Factor A

Topics: Anticoagulants; Blood Coagulation; Chemotaxis; Factor VIIa; Humans; Integrins; Lipoproteins; Models, Biological; Neoplasm Metastasis; Neoplasms; Neovascularization, Pathologic; Signal Transduction; Thromboplastin; Thrombosis; Vascular Endothelial Growth Factor A

As early as the nineteenth century, it was recognized that tissues exhibit clot-promoting activity. The realization that a single protein was responsible for this procoagulant effect led to the discovery of tissue factor (TF), but for many years it was thought that TF activity was restricted to the activation of an auxiliary pathway that had little biological relevance. Research over recent decades has greatly improved our understanding of TF function, and today the protein is recognized to be the primary biological initiator of the coagulation cascade. Furthermore, it is now clear that TF is a major player in several other physiological processes. We present recent data suggesting that TF is vital not only for effective haemostasis, but also for angiogenesis and tumour metastasis, for embryonic development, and for induction of a pro-inflammatory response. These advances in our knowledge of TF function will improve our understanding of several pathophysiological conditions, and may expand the therapeutic options available for their treatment.

Topics: Animals; Embryonic Development; Hemostasis; Humans; Inflammation; Neoplasm Metastasis; Neovascularization, Pathologic; Thromboplastin

The possible role of tissue factor (TF) in colorectal cancer (CRC) is reviewed. A correlation between TF expression and advanced stages of malignancy, and a correlation between TF expression and overall survival have been suggested in CRC. This is supported by experimental studies indicating that TF plays a key role in growth, invasion and dissemination of tumour cells, and in tumour related angiogenesis as well. In addition, the activation of TF in CRC patients in relation to the surgical trauma, perioperative blood transfusion and development of postoperative bacterial infectious complications are discussed. Finally, future directions for the development of anticancer modalities directed against regulation of TF are considered.

Topics: Animals; Biomarkers, Tumor; Colorectal Neoplasms; Endothelial Growth Factors; Humans; Neoplasm Invasiveness; Neoplasm Metastasis; Neovascularization, Pathologic; Prognosis; Thromboembolism; Thromboplastin

There is now considerable evidence that the blood coagulation system plays an important role in the biology of malignant tumors. This evidence has been derived from a combination of clinical, biochemical, histological, and pharmacological observations that point to the possibility of favorably affecting the course of malignant disease with agents that interfere with blood coagulation pathways. For a number of years our laboratory has used experimental models of blood-borne metastasis to study the events that follow the introduction of procoagulant-bearing tumor cells into the circulating blood. This article summarizes our experience with these models, which suggests that intravascular coagulation is a necessary prelude to lung tumor formation and that interruption of coagulation pathways in various ways may be an effective antimetastatic strategy. We have shown that anticoagulation with commonly used agents such as unfractionated heparin and warfarin (Coumadin) prevent tumor formation by limiting the ability of tumor cells to be retained in the pulmonary microvasculature. Binding of fibrin-coated tumor cells to activated platelets is essential for this retention and, therefore, treatment with potent antiplatelet agents such as abciximab is also effective. The predominant tumor procoagulant is tissue factor (TF), and direct targeting of this protein with concanavalin A, monoclonal antibodies, and tissue factor pathway inhibitor (TFPI) has provided compelling evidence that TF is an important determinant of tumor seeding in these experimental models. Collectively, our data provide strong support for the concept that some form of anticoagulant therapy would be a useful adjunct to existing cancer treatments.

Topics: Animals; Anticoagulants; Blood Coagulation Factors; Disease Models, Animal; Hemostasis; Humans; Neoplasm Metastasis; Neoplasms, Experimental; Thromboplastin

Tissue factor is a cellular receptor for plasma clotting factor VII. In health, tissue factor is constitutively expressed in many cells, such as fibroblasts and keratinocytes, but is absent in vascular cells that come in contact with blood. However, tissue factor is induced in vascular cells in certain pathophysiological conditions. Thus, vessel wall injury or a disease condition allows blood to come in contact with tissue factor, resulting in factor VII binding to tissue factor. Once native factor VII complexed with tissue factor is converted to the enzyme factor VIIa, the complex triggers the clotting cascade that ultimately leads to fibrin formation. In addition to its established role in coagulation, molecular links between factor VIIa/tissue factor and various biological processes, such as development, inflammation, and tumor metastasis, are also evident. Recent studies suggest that factor VIIa/tissue factor affects various cellular processes by inducing intracellular signaling. Emerging evidence suggest that factor VIIa/tissue factor participates in cell signaling by two distinct mechanisms: proteolysis-independent signaling via the cytoplasmic domain of tissue factor, and proteolysis-dependent signaling, which is independent of tissue factor's cytoplasmic tail. In proteolysis-independent signaling, filamin 1 is recruited to tissue factor upon its ligation, and this probably provides an essential intracellular link in transmitting signals from tissue factor. In proteolysis-dependent signaling, factor VIIa/tissue factor activates one or more protease-activated receptors, which couple to G proteins, to impact multiple signaling pathways. In this chapter, we review various nonhemostatic functions attributed to factor VIIa and tissue factor, describe signaling mechanisms initiated upon factor VIIa binding to tissue factor, and discuss how factor VIIa/tissue factor-induced signaling could contribute to various pathophysiological processes. The relationship between increased clotting and manifestation of various diseases is well recognized. Although aberrant activation of the coagulation pathway is primarily the consequence of a disease, activation of the coagulation pathway during the disease process, in turn, could contribute to pathogenesis of the disease. Further, recent transgenic studies in mouse suggest that the coagulation system also plays a role in embryogenesis and development. Then the question arises, how do proteins involved in the clotting

Topics: Animals; Blood Coagulation; Factor VIIa; Humans; Inflammation; Neoplasm Metastasis; Neovascularization, Pathologic; Signal Transduction; Thromboplastin

Tissue factor (TF) is the primary cellular initiator of blood coagulation. At sites of vascular injury, formation of a TF:FVIIa complex leads to the generation of FXa, thrombin and the deposition of fibrin to limit hemorrhage. In contrast to its beneficial role in hemostasis, TF initiates life-threatening intravascular thrombosis in sepsis, atherosclerosis and cancer. More recently, TF has been proposed to play a role in other biological processes, including tumor-associated angiogenesis, metastasis and inflammation. Indeed, gene targeting of TF resulted in embryonic lethality, which appeared to be due to a defect in the yolk sac vasculature.

Topics: Animals; Blood Coagulation; Blood Coagulation Disorders; Gene Silencing; Gene Targeting; Hemostasis; Humans; Mice; Neoplasm Metastasis; Neovascularization, Physiologic; Signal Transduction; Thrombin; Thromboplastin; Yolk Sac

Cancer patients are prone to venous thromboembolism (VTE), and this hypercoagulability favors tumor growth and metastasis. After a brief review of the clinical aspects of VTE and cancer, we discuss the pathogenesis of hypercoagulability with an emphasis on the role of tissue factor (TF). The discovery that, in addition to tumor cells, TF is expressed by tumor-associated macrophages and tumor-associated endothelial cells led to studies of the role of TF in the regulation of tumor angiogenesis. In human lung cancer, melanoma, and breast cancer, TF and vascular endothelial growth factor (VEGF) co-localize in tumor cells; a close correlation exists between TF and VEGF synthesis (P = .001) in tumor cell lines and with angiogenesis in vivo in a severe, combined immunodeficient mouse model. Transfection of a TF/VEGF low-producing human tumor cell line with full length TF complementary DNA (cDNA) results in conversion to a high producer of TF and VEGF; transfection of a deletion-mutant TF cDNA lacking cytoplasmic serine residues restores full TF procoagulant activity but not VEGF synthesis to the cells. These results suggest that the cytoplasmic tail of TF is necessary for tumor cell VEGF synthesis. Targeting of TF in tumors and tumor-associated blood vessels is discussed as a strategy for drug delivery and rational anti-cancer and anti-angiogenesis drug design.

Topics: Cell Division; Fibrin; Hemostatics; Humans; Neoplasm Metastasis; Neoplasms; Neovascularization, Pathologic; Thromboplastin

Topics: Animals; Binding Sites; Blood Coagulation; Cardiovascular Diseases; Catalysis; Clinical Trials as Topic; Dogs; Endothelial Growth Factors; Enzyme Activation; Factor VIIa; Gene Expression Regulation; Granulocytes; Humans; Kinetics; Lymphokines; Macromolecular Substances; MAP Kinase Signaling System; Models, Molecular; Neoplasm Metastasis; Neoplasm Proteins; Protein Binding; Protein Conformation; Protein Structure, Tertiary; Signal Transduction; Structure-Activity Relationship; Substrate Specificity; Thermodynamics; Thromboplastin; Thrombosis; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors

Human tissue factor pathway inhibitor (TFPI) is a modular protein comprised of three Kunitz type domains flanked by peptide segments that are less structured. The sequential order of the elements are: an N-terminal acidic region followed by the first Kunitz domain (K1), a linker region, a second Kunitz domain (K2), a second linker region, the third Kunitz domain (K3), and the C-terminal basic region. The K1 domain inhibits factor VIIa complexed to tissue factor (TF) while the K2 domain inhibits factor Xa. No direct protease inhibiting functions have been demonstrated for the K3 domain. Importantly, the Xa-TFPI complex is a much more potent inhibitor of the VIIa-TF than TFPI by itself. Furthermore, the C-terminal basic region of TFPI is required for rapid physiologic inhibition of coagulation and is needed for the inhibition of smooth muscle cell proliferation. Although a number of additional targets for attachment have been reported, the C-terminal basic region appears to play an important role in binding of TFPI to cell surfaces. A primary site of TFPI synthesis is endothelium and the endothelium-bound TFPI contributes to the antithrombotic potential of the vascular endothelium. Further, increased levels of plasma TFPI under septic conditions may represent endothelial dysfunction. We have proposed that the extravascular cells that synthesize TF also synthesize TFPI providing dual components necessary for the regulation of clotting in their microenvironment. Like the TF synthesis in these cells is augmented by serum, so is the case with the TFPI gene expression. TFPI gene knock out mice reveal embryonic lethality suggesting a possible role of this protein in early development. Since TF-induced coagulation is thought to play a significant role in many disease states, including disseminated intravascular clotting, sepsis, acute lung injury and cancer, recombinant TFPI may be a beneficial therapeutic agent in these disease states to attenuate pathologic clotting. The purpose of this review is to outline recent developments in the field related to the structural specificity and biology of TFPI.

Topics: Acute Disease; Amino Acid Sequence; Amino Acids; Antiphospholipid Syndrome; Blood Coagulation; Cardiovascular Diseases; Endothelium, Vascular; Humans; Lipoproteins; Lung Diseases; Models, Biological; Models, Molecular; Molecular Sequence Data; Neoplasm Metastasis; Neoplasms; Protein Conformation; Protein Structure, Tertiary; Sepsis; Sequence Alignment; Sequence Homology, Amino Acid; Structure-Activity Relationship; Thrombophilia; Thromboplastin

Tissue factor, a 47 kDa membrane glycoprotein, lies at the basis of the extrinsic pathway of the coagulation cascade. Interaction of TF with factor VIIa results in the formation of fibrin from fibrinogen, thereby inducing the formation of a blood clot. In addition to this well-established role in blood coagulation, TF is associated with various other physiological processes such as sepsis, inflammation, angiogenesis, metastasis and atherosclerosis. The molecular basis of the latter events is slowly emerging. It has become clear that TF-FVIIa interaction elicits a variety of intracellular signalling events that may be implicated in these actions. These events include the sequential activation of Src-like kinases, MAP kinases, small GTPases and calcium signalling. How this progress in the understanding of TF associated signal transduction may generate answers as to the mechanism through which TF exerts it pleiotropic effects will be focus of this review.

Topics: Animals; Arteriosclerosis; Blood Coagulation; Calcium Signaling; Cytokines; Disseminated Intravascular Coagulation; Factor VIIa; Fibroblasts; Gene Expression Regulation; GTP Phosphohydrolases; Humans; Inflammation; Lipoproteins; Lipoproteins, LDL; MAP Kinase Signaling System; Mice; Mice, Knockout; Muscle, Smooth, Vascular; Neoplasm Metastasis; Neoplasms; Neovascularization, Physiologic; Papio; Phagocytes; Sepsis; Signal Transduction; src-Family Kinases; Thromboplastin

Accumulating evidence suggests that protease receptors and their cognate protease ligands play important roles in cell-signaling events that regulate cell adhesion and migration in inflammation as well as tumor invasion and metastasis. Tissue factor (TF), the cell surface receptor for the serine protease VIIa and the initiator of the coagulation pathways, supports metastatic implantation by activating extracellular, protease-dependent signaling pathways and by intracellular links to the actin cytoskeleton. The adhesion of TF-expressing tumor cells can be mediated by interactions of the receptor-protease complex with specific matrix-associated inhibitors, suggesting a novel bridging mechanism by which proteases participate in migratory functions of cells.

Topics: Humans; Inflammation; Neoplasm Metastasis; Serine Endopeptidases; Signal Transduction; Thromboplastin

Topics: Animals; Blood Coagulation; Cell Adhesion; Cell Movement; Contractile Proteins; Factor VIIa; Filamins; Humans; Ligands; Microfilament Proteins; Neoplasm Metastasis; Phosphorylation; Signal Transduction; Thromboplastin; Tumor Cells, Cultured

There is considerable evidence that the hemostatic system is involved in the growth and spread of malignant disease. There is an increased incidence of thromboembolic disease in patients with cancers and hemostatic abnormalities are extremely common in such patients. Antihemostatic agents have been successfully used to treat a variety of experimental tumors, and several clinical trials in humans have been initiated. Although metastasis is undoubtedly multifactorial, intravascular coagulation activation and peritumor fibrin deposition seem to be important. The mechanisms by which hemostatic activation facilitates the malignant process remain to be completely elucidated. Of central importance may be the presence on malignant cells of tissue factor and urokinase receptor. Recent studies have suggested that these proteins, and others, may be involved at several stages of metastasis, including the key event of neovascularization. Tissue factor, the principal initiator of coagulation, may have additional roles, outside of fibrin formation, that are central to the biology of some solid tumors.

Topics: Animals; Anticoagulants; Antineoplastic Agents; Biomarkers; Blood Coagulation Tests; Cell Adhesion; Cysteine Endopeptidases; Disseminated Intravascular Coagulation; Factor Xa; Fibrin; Fibrinolysis; Hemostasis; Heparin; Humans; Monocytes; Neoplasm Metastasis; Neoplasm Proteins; Neoplasms; Neoplasms, Experimental; Neoplastic Cells, Circulating; Neovascularization, Pathologic; Platelet Activation; Platelet Aggregation Inhibitors; Receptors, Thrombin; Thrombophilia; Thrombophlebitis; Thromboplastin; Vitamin K

Tumor cells frequently express tissue factor, a transmembrane glycoprotein that functions as the cellular receptor and catalytic cofactor for the serine protease factor VIIa. In human tumors, tissue factor expression correlates spatially with neovascularization, indicating that tissue factor function may be linked to angiogenic properties of malignant tumors. Tissue factor also supports hematogenous tumor dissemination. The role of tissue factor in metastasis appears to involve both the coagulation pathway triggered by interactions of the tissue factor extracellular domain as well as cellular events dependent on the short cytoplasmic domain that may participate in tissue factor-mediated outside-in signaling.

Topics: Humans; Neoplasm Metastasis; Neoplasms; Neovascularization, Pathologic; Thromboplastin

Cancer cells may promote fibrin formation in the tumor microenvironment through availability of procoagulant activities which are mainly of two types: tissue thromboplastin-like or direct activator of coagulation factor X. The pharmacological modulation of these activities could be potentially important in the control of metastasis growth. However, very limited information is available so far on this issue; it has recently been shown that the direct activator of coagulation factor X is a vitamin K-dependent activity which is depressed by warfarin treatment, not by anticoagulation with heparin or defibrinating enzymes. Whether the inhibition of this peculiar cancer procoagulant is involved in the antimetastatic activity of warfarin is a stimulating hypothesis which needs to be further substantiated.

Topics: Adenocarcinoma; Animals; Antineoplastic Agents; Blood Coagulation Factors; Cell Membrane; Cysteine Endopeptidases; Endopeptidases; Humans; Leukemia, Myeloid, Acute; Mice; Mucins; Neoplasm Metastasis; Neoplasm Proteins; Thromboplastin; Warfarin

Topics: Blood Coagulation; Fibrin; Fibrinogen; Fibrinolysis; Hemorrhage; Heparin; Humans; Neoplasm Metastasis; Neoplasms; Thromboplastin; Thrombosis

Topics: Animals; Aorta; Autoradiography; Basement Membrane; Blood Vessels; Embolism; Endothelium; Fibrin; Fibrinolysis; HeLa Cells; Humans; Microscopy, Electron; Mitosis; Neoplasm Metastasis; Neoplasms; Neoplasms, Experimental; Neoplastic Cells, Circulating; Plasminogen; Platelet Adhesiveness; Saphenous Vein; Thromboplastin; Thymidine; Tritium

Few effective second-line treatments exist for women with recurrent or metastatic cervical cancer. Accordingly, we aimed to evaluate the efficacy and safety of tisotumab vedotin, a tissue factor-directed antibody-drug conjugate, in this patient population.. This multicentre, open-label, single-arm, phase 2 study was done across 35 academic centres, hospitals, and community practices in Europe and the USA. The study included patients aged 18 years or older who had recurrent or metastatic squamous cell, adenocarcinoma, or adenosquamous cervical cancer; disease progression on or after doublet chemotherapy with bevacizumab (if eligible by local standards); who had received two or fewer previous systemic regimens for recurrent or metastatic disease; had measurable disease based on Response Evaluation Criteria in Solid Tumors (RECIST; version 1.1); and had an Eastern Cooperative Oncology Group performance status of 0 or 1. Patients received 2·0 mg/kg (up to a maximum of 200 mg) tisotumab vedotin intravenously once every 3 weeks until disease progression (determined by the independent review committee) or unacceptable toxicity. The primary endpoint was confirmed objective response rate based on RECIST (version 1.1), as assessed by the independent review committee. Activity and safety analyses were done in patients who received at least one dose of the drug. This study is ongoing with recruitment completed and is registered with ClinicalTrials.gov, NCT03438396.. 102 patients were enrolled between June 12, 2018, and April 11, 2019; 101 patients received at least one dose of tisotumab vedotin. Median follow-up at the time of analysis was 10·0 months (IQR 6·1-13·0). The confirmed objective response rate was 24% (95% CI 16-33), with seven (7%) complete responses and 17 (17%) partial responses. The most common treatment-related adverse events included alopecia (38 [38%] of 101 patients), epistaxis (30 [30%]), nausea (27 [27%]), conjunctivitis (26 [26%]), fatigue (26 [26%]), and dry eye (23 [23%]). Grade 3 or worse treatment-related adverse events were reported in 28 (28%) patients and included neutropenia (three [3%] patients), fatigue (two [2%]), ulcerative keratitis (two [2%]), and peripheral neuropathies (two [2%] each with sensory, motor, sensorimotor, and neuropathy peripheral). Serious treatment-related adverse events occurred in 13 (13%) patients, the most common of which included peripheral sensorimotor neuropathy (two [2%] patients) and pyrexia (two [2%]). One death due to septic shock was considered by the investigator to be related to therapy. Three deaths unrelated to treatment were reported, including one case of ileus and two unknown causes.. Tisotumab vedotin showed clinically meaningful and durable antitumour activity with a manageable and tolerable safety profile in women with previously treated recurrent or metastatic cervical cancer. Given the poor prognosis for this patient population and the low activity of current therapies in this setting, tisotumab vedotin, if approved, would represent a new treatment for women with recurrent or metastatic cervical cancer.. Genmab, Seagen, Gynaecologic Oncology Group, and European Network of Gynaecological Oncological Trial Groups.

Topics: Adult; Antibodies, Monoclonal, Humanized; Female; Humans; Middle Aged; Neoplasm Metastasis; Neoplasm Recurrence, Local; Oligopeptides; Thromboplastin; Uterine Cervical Neoplasms

Epithelial-mesenchymal transitions (EMTs) are high-profile in the field of circulating tumor cells (CTCs). EMT-shifted CTCs are considered to encompass pre-metastatic subpopulations though underlying molecular mechanisms remain elusive. Our previous work identified tissue factor (TF) as an EMT-induced gene providing tumor cells with coagulant properties and supporting metastatic colonization by CTCs. We here report that vimentin, the type III intermediate filament considered a canonical EMT marker, contributes to TF regulation and positively supports coagulant properties and early metastasis. Different evidence further pointed to a new post-transcriptional regulatory mechanism of TF mRNA by vimentin: (1) vimentin silencing accelerated TF mRNA decay after actinomycin D treatment, reflecting TF mRNA stabilization, (2) RNA immunoprecipitation revealed enriched levels of TF mRNA in vimentin immunoprecipitate, (3) TF 3'-UTR-luciferase reporter vector assays implicated the 3'-UTR of TF mRNA in vimentin-dependent TF regulation, and (4) using different TF 3'UTR-luciferase reporter vectors mutated for potential miR binding sites and specific Target Site Blockers identified a key miR binding site in vimentin-dependent TF mRNA regulation. All together, these data support a novel mechanism by which vimentin interferes with a miR-dependent negative regulation of TF mRNA, thereby promoting coagulant activity and early metastasis of vimentin-expressing CTCs.

Topics: 3' Untranslated Regions; Breast Neoplasms; Cell Line, Tumor; Dactinomycin; Epithelial-Mesenchymal Transition; Female; Gene Expression Regulation, Neoplastic; Humans; MicroRNAs; Neoplasm Metastasis; Neoplastic Cells, Circulating; RNA Stability; RNA, Messenger; Thromboplastin; Vimentin

CD142 promotes cell mobility, which contributes to carcinogenesis. However, the role of CD142 on colorectal cancer (CRC) mobility is unclear. This study showed that CD142 expression increased in CRC tissues, especially in those with invasion or metastasis. The positive sorting or overexpression of CD142 promoted the invasion and migration of CRC cells. Overall, CD142 may be responsible for CRC mobility.

Topics: Cell Line, Tumor; Cell Movement; Colorectal Neoplasms; Gene Expression Regulation, Neoplastic; Humans; Neoplasm Invasiveness; Neoplasm Metastasis; Thromboplastin

Within tumors, the coagulation-inducing protein tissue factor (TF), a major initiator of blood coagulation, has been shown to play a critical role in the hematogenous metastasis of tumors, due to its effects on tumor hypercoagulability and on the mediation of interactions between platelets and tumor cells. Targeting tumor-associated TF has therefore great therapeutic potential for antimetastasis therapy and preventing thrombotic complication in cancer patients. Herein, we reported a novel peptide-based nanoparticle that targets delivery and release of small interfering RNA (siRNA) into the tumor site to silence the expression of tumor-associated TF. We showed that suppression of TF expression in tumor cells blocks platelet adhesion surrounding tumor cells

Topics: Animals; Cell Line, Tumor; Female; Gene Expression Regulation, Neoplastic; Gene Silencing; Humans; Lung Neoplasms; Mice, Nude; Nanoparticles; Neoplasm Metastasis; Neoplasm Proteins; Neoplasms, Experimental; Neoplastic Cells, Circulating; RNA, Small Interfering; Thrombophilia; Thromboplastin; Thrombosis

Metastasis results from a complex set of traits acquired by tumor cells, distinct from those necessary for tumorigenesis. Here, we investigate the contribution of enhancer elements to the metastatic phenotype of osteosarcoma. Through epigenomic profiling, we identify substantial differences in enhancer activity between primary and metastatic human tumors and between near isogenic pairs of highly lung metastatic and nonmetastatic osteosarcoma cell lines. We term these regions metastatic variant enhancer loci (Met-VELs). Met-VELs drive coordinated waves of gene expression during metastatic colonization of the lung. Met-VELs cluster nonrandomly in the genome, indicating that activity of these enhancers and expression of their associated gene targets are positively selected. As evidence of this causal association, osteosarcoma lung metastasis is inhibited by global interruptions of Met-VEL-associated gene expression via pharmacologic BET inhibition, by knockdown of AP-1 transcription factors that occupy Met-VELs, and by knockdown or functional inhibition of individual genes activated by Met-VELs, such as that encoding coagulation factor III/tissue factor (F3). We further show that genetic deletion of a single Met-VEL at the F3 locus blocks metastatic cell outgrowth in the lung. These findings indicate that Met-VELs and the genes they regulate play a functional role in metastasis and may be suitable targets for antimetastatic therapies.

Topics: Carcinogenesis; Cell Line, Tumor; Enhancer Elements, Genetic; Epigenomics; Gene Expression Regulation, Neoplastic; Genome, Human; Humans; Lung Neoplasms; Neoplasm Metastasis; Osteosarcoma; Proteins; Selection, Genetic; Thromboplastin; Transcription Factor AP-1; Tumor Microenvironment

Thrombosis is one of the leading causes of mortality in cancer patients. The aim of the study was to evaluate the concentrations and activities of selected haemostatic parameters in the plasma of patients diagnosed with breast cancer (BrCa) and to make an attempt at finding associations with their levels and selected clinicopathological factors; clinical classification, histological grading, and molecular subtype of BrCa. The study involved 145 Caucasian ethnicity women. Eighty-five women aged 45-66 with primary BrCa without distant metastases (M0). Inclusion criteria were as follows: histopathological examination confirming the diagnosis of primary BrCa, without previous radiotherapy and chemotherapy. The control group consisted of 60, post-menopausal women, aged 45-68. Haemostatic profile expressed by concentrations and activities of tissue factor (TF) and its inhibitor (TFPI) as well as concentrations of tissue plasminogen activator (t-PA) and plasminogen activator inhibitor type 1 (PAI-1) were measured applying immunoassay techniques. A significantly higher concentration of PAI-1 was noted in patients with BrCa localized in the left breast. We observed significantly lower activity of TFPI and significantly higher concentration of PAI-1 in the group of patients with invasive ductal carcinoma as compared with invasive lobular carcinoma. A significantly higher concentration of t-PA in patients with pT2 BrCa in relation to pT1 cases was noted. Based on comprehensive analysis of haemostatic profile depending on clinicopathological features, we suggest that haemostatic parameters play crucial roles in invasion and metastases of malignant tumours.

Topics: Aged; Breast Neoplasms; Female; Humans; Lipoproteins; Middle Aged; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasm Proteins; Plasminogen Activator Inhibitor 1; Thromboplastin; Tissue Plasminogen Activator

Prostate cancer (PC) patients in advanced stages of the disease have high risk of blood coagulation complications. The procoagulant molecule Tissue factor (TF), and the fibrinolysis inhibitor plasminogen activator inhibitor-1 PAI-1 play important role in this complication. Extracellular vesicles (EV) shed from cancer cells may contribute to the regulation of TF and PAI-1. The procoagulant activity of EV can be associated with the oncogenic and metastatic characteristics of their cells.. We have expressed EGFRvIII in DU145 cells to assess the role of this oncogene in the procoagulant activity of EV. The intercellular exchange of TF via EV was assessed by downregulating its expression in DU145 cells using shRNA vector, and determining the transfer of TF via EV enriched with the protein. Two PC cell lines with different metastatic potential were used to assess the correlation between the procoagulant activity of EV and the metastatic potential of PC cells. Photometric assays were used to determine FXa-activity and thrombin generation as indicators for the procoagulant activity of EV. Double-tagged proteinase-activated receptor 1(PAR-1) expressed in CHO cells to assess its activation by EV.. The expression of EGFRvIII in DU145 cells led to increased mRNA levels for TF and PAI-1, but the increase in these proteins expression was detected mostly in the EV. EV with enhanced levels of TF protein conferred higher TF procoagulant activity on the acceptor cells by intercellular exchange of this protein. Procoagulant activity of EV, assessed by FXa activity, and thrombin generation, was correlated with the oncogenic and metastatic potential of PC cells. The ability of EV to generate thrombin led to the activation of PAR-1, which was evident by the truncation of tagged-PAR-1.. The active oncogene EGFRvIII increases the concentration of TF and PAI-1 in EV. The procoagulant activity of EV is associated with the oncogenic and metastatic characteristics of their PC cells. Also, EV may contribute to the high procoagulant activity in the tumour microenvironment by the intercellular exchange of TF. Finally, through the generation of thrombin, EV can activate PAR-1, which evidently contributes to cancer progression, linking the coagulation system to tumor progression.

Topics: Cell Line, Tumor; Extracellular Vesicles; Humans; Male; Neoplasm Metastasis; Plasminogen Activator Inhibitor 1; Prostatic Neoplasms; Thrombin; Thromboplastin

Platelets are thought to play an important role in metastasis formation, although the mechanisms involved remain incompletely understood. Here we studied the influence of platelet numbers on organ-specific metastasis to the lungs and lymph nodes using Tpo deficient mice that have low platelet counts. After tail vein injection of 4T1 breast cancer cells, the number of lung metastases was significantly lower in Tpo

Topics: Animals; Blood Platelets; Cell Line, Tumor; Female; Lung Neoplasms; Lymphatic Metastasis; Mammary Neoplasms, Experimental; Mice; Mice, Inbred BALB C; Neoplasm Metastasis; Thromboplastin

Apart from blood coagulation, coagulation proteases are involved inextricably in cancer progression/propagation via intra/inter-cellular signaling, mediated predominantly by protease-activated receptors (PARs). Microvesicles (MVs), a plasma membrane shredded component, has recently been identified as an important contributor to human breast cancer metastasis. However, the role of PAR2 in promoting MVs generation from breast cancer cells remains largely unexplored. The objective of this study is to investigate whether coagulation protease-mediated human breast cancer propagation commences via MVs and also to decipher the underlying signaling mechanism. Here, we elicited that coagulation factor-FVIIa and Trypsin activates PAR2, which governs MVs shedding from MDAMB231 cells by altering actomyosin dynamics. Treatment of cells with PAR2 activators facilitate MVs generation by activating three independent (MAPK, P38, and Rho) signaling cascades. MAPK, signals through activating MLCK followed by MLC phosphorylation to alter myosin organization whereas, P38 reorganizes actin dynamics by the sequential activation of MK2 and HSP27. RhoA-dependent ROCK-II activation again contributes to remodeling myosin II activity. Further, both our in vitro and in vivo analyses showed that these MVs potentiate invasive and migratory property to the recipient cells. Breast cancer patients blood show an elevation of TF-bearing, pro-metastatic MVs than normal. These findings give an insight into the detailed signaling mechanism involved in the production of MVs with transforming ability from PAR2-activated human breast cancer cells. Understanding these mechanistic details will certainly help to identify crucial targets for therapeutic interventions in MVs-associated human breast cancer metastasis.

Topics: Actomyosin; Animals; Breast Neoplasms; Cell Line; Cell Line, Tumor; Cell-Derived Microparticles; Factor VIIa; Female; Humans; MCF-7 Cells; Mice; Neoplasm Metastasis; Neoplasm Transplantation; Phosphorylation; Receptor, PAR-2; Receptors, G-Protein-Coupled; Signal Transduction; Thromboplastin; Trypsin

Essentials Heparanase forms a complex with tissue factor and enhances the generation of factor Xa. The present study was aimed to identify the procoagulant domain of heparanase. Procoagulant peptides significantly shortened bleeding time and enhanced wound healing. Tissue factor pathway inhibitor (TFPI)-2 derived peptides inhibited the procoagulant peptides.. Background Heparanase, which is known to be involved in angiogenesis and metastasis, was shown to form a complex with tissue factor (TF) and to enhance the generation of activated factor X (FXa). Our study demonstrated that peptides derived from TF pathway inhibitor (TFPI)-2 impeded the procoagulant effect of heparanase, and attenuated inflammation, tumor growth, and vascularization. Aims To identify the procoagulant domain in the heparanase molecule, and to evaluate its effects in a model of wound healing that involves inflammation and angiogenesis. Methods Twenty-four potential peptides derived from heparanase were generated, and their effect was studied in an assay of FXa generation. Peptides 14 and 16, which showed the best procoagulant effect, were studied in a bleeding mouse model and in a wound-healing mouse model. Results Peptides 14 and 16 increased FXa levels by two-fold to three-fold, and, at high levels, caused consumption coagulopathy. The TFPI-2-derived peptides explored in our previous study were found to inhibit the procoagulant effect induced by peptides 14 and 16. In the bleeding model, time to clot formation was shortened by 50% when peptide 14 or peptide 16 was topically applied or injected subcutaneously. In the wound-healing model, the wound became more vascular, and its size was reduced to one-fifth as compared with controls, upon 1 week of exposure to peptide 14 or peptide 16 applied topically or injected subcutaneously. Conclusions The putative heparanase procoagulant domain was identified. Peptides derived from this domain significantly shortened bleeding time and enhanced wound healing.

Topics: Animals; Blood Coagulation; Coagulants; Factor Xa; Fibrin Fibrinogen Degradation Products; Fibrinogen; Glucuronidase; Glycoproteins; Hematologic Agents; Hemorrhage; Humans; Inflammation; Male; Mice; Neoplasm Metastasis; Neovascularization, Pathologic; Partial Thromboplastin Time; Peptides; Protein Domains; Prothrombin Time; Thrombelastography; Thromboplastin; Thrombosis; Wound Healing

To investigate whether cancer stem\ cells (CSCs) more efficiently activating platelets and\ evading immune surveillance than non-CSCs thus\ promoting metastasis.. We enriched and identified sphere-forming\ cells (SFCs) and coincubated washed platelets\ with several platelet activators including collagen,\ 4T1 and SFCs. Platelet-coating tumor cells,\ platelet activation and TGF-β1 release were analyzed.\ Then natural kell cells (NK) were incubated\ with supernatants of different activated platelet\ samples what we called sample release (SR). The degranulation\ assay and NKG2D expression on NK\ cells were conducted by flow cytometry. Finally tissue\ factor (TF) expression of SFCs or 4T1 were evaluated\ by western blot.. Breast cancer cell line 4T1 could form\ spheres in serum-free medium at low adherence.\ Sphere-forming cells expressed high levels of the\ CD24-/lowCD44 + stem cell phenotype. Both\ sphere-forming cells or 4T1 were coated with abundant\ platelets while sphere-forming cells induced\ significantly higher expression of platelet activating\ receptor CD62p than 4T1 did (P < 0.01). And\ sphere-forming cells induced platelets to produce\ more TGF-β1 than 4T1 did (P < 0.01). Furthermore,\ sample releases induced by sphere-forming cells\ caused more vigorous inhibition of NK cells antitumor\ reactivity (P < 0.05) and reduced NKG2D expression\ (P < 0.01). The final results showed that\ sphere-forming cells expressed higher levels of TF than 4T1 (P < 0.05).. Our findings indicate that CSCs\ could efficiently activate platelets, induce platelets\ to secrete more TGF-β1, decrease NKG2D expression\ and inhibit antitumor activity of NK cell, compared\ with 4T1. And higher levels of TF expression\ of CSCs may account for this correlation of CSCs and platelets.

Topics: Animals; Blood Platelets; Breast Neoplasms; Cell Line, Tumor; Female; Humans; Immune Evasion; Killer Cells, Natural; Mice; Mice, Inbred BALB C; Neoplasm Metastasis; Neoplastic Stem Cells; Thromboplastin

Tumor invasiveness depends on the ability of tumor cells to breach endothelial barriers. In this study, we investigated the mechanism by which the adhesion of melanoma cells to endothelium regulates adherens junction integrity and modulates tumor transendothelial migration (TEM) by initiating thrombin generation. We found that the B-Raf(V600E) mutation in metastatic melanoma cells up-regulated tissue factor (TF) expression on cell membranes and promoted thrombin production. Co-culture of endothelial monolayers with metastatic melanoma cells mediated the opening of inter-endothelial spaces near melanoma cell contact sites in the presence of platelet-free plasma (PFP). By using small interfering RNA (siRNA), we demonstrated that B-Raf(V600E) and TF silencing attenuated the focal disassembly of adherens junction induced by tumor contact. Vascular endothelial-cadherin (VE-cadherin) disassembly was dependent on phosphorylation of p120-catenin on Ser-879 and VE-cadherin on Tyr-658, Tyr-685, and Tyr-731, which can be prevented by treatment with the thrombin inhibitor, hirudin, or by silencing the thrombin receptor, protease-activated receptor-1, in endothelial cells. We also provided strong evidence that tumor-derived thrombin enhanced melanoma TEM by inducing ubiquitination-coupled VE-cadherin internalization, focal adhesion formation, and actin assembly in endothelium. Confocal microscopic analysis of tumor TEM revealed that junctions transiently opened and resealed as tumor cells accomplished TEM. In addition, in the presence of PFP, tumor cells preferentially transmigrated via paracellular routes. PFP supported melanoma transmigration under shear conditions via a B-Raf(V600E)-thrombin-dependent mechanism. We concluded that the activation of thrombin generation by cancer cells in plasma is an important process regulating melanoma extravasation by disrupting endothelial junction integrity.

Topics: Antigens, CD; Cadherins; Cell Adhesion; Cell Line, Tumor; Cell Membrane; Cell Movement; Electric Impedance; Endothelium, Vascular; Gene Silencing; Human Umbilical Vein Endothelial Cells; Humans; Melanoma; Mutation; Neoplasm Metastasis; Permeability; Phosphorylation; Plasma; Proto-Oncogene Proteins B-raf; RNA, Small Interfering; Skin Neoplasms; Thrombin; Thromboplastin; Ubiquitination

Venous thromboembolism constitutes one of the main causes of death during the progression of a cancer. We previously demonstrated that tissue factor (TF)-bearing cancer cell-derived microparticles accumulate at the site of injury in mice developing a pancreatic cancer. The presence of these microparticles at the site of thrombosis correlates with the size of the platelet-rich thrombus. The objective of this study was to determine the involvement of TF expressed by cancer cell-derived microparticles on thrombosis associated with cancer. We observed that pancreatic cancer cell derived microparticles expressed TF, its inhibitor tissue factor pathway inhibitor (TFPI) as well as the integrins αvβ1 and αvβ3. In mice bearing a tumor under-expressing TF, a significant decrease in circulating TF activity associated with an increase bleeding time and a 100-fold diminished fibrin generation and platelet accumulation at the site of injury were observed. This was mainly due to the interaction of circulating cancer cell-derived microparticles expressing TFPI with activated platelets and fibrinogen. In an ectopic model of cancer, treatment of mice with Clopidogrel, an anti-platelet drug, decreased the size of the tumors and restored hemostasis by preventing the accumulation of cancer cell-derived microparticles at the site of thrombosis. In a syngeneic orthotopic model of pancreatic cancer Clopidogrel also significantly inhibited the development of metastases. Together, these results indicate that an anti-platelet strategy may efficiently treat thrombosis associated with cancer and reduce the progression of pancreatic cancer in mice.

Topics: Animals; Blood Coagulation; Blotting, Western; Cell-Derived Microparticles; Clopidogrel; Disease Models, Animal; Flow Cytometry; Fluorescent Antibody Technique; Integrins; Mice; Mice, Inbred C57BL; Neoplasm Metastasis; P-Selectin; Pancreatic Neoplasms; Platelet Activation; Platelet Aggregation Inhibitors; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Thromboplastin; Thrombosis; Ticlopidine; Tumor Cells, Cultured

Hypercoagulability in malignancy increases the risk of thrombosis, but is also involved in cancer progression. Experimental studies suggest that tissue factor (TF) and tissue factor pathway inhibitor (TFPI) are involved in cancer biology as a tumor- promoter and suppressor, respectively, but the clinical significance is less clear. Here, we aimed to investigate the clinical relevance of TF and TFPI genetic and phenotypic diversity in breast cancer.. The relationship between tumor messenger RNA (mRNA) expression and plasma levels of TF and TFPI (α and β), tagging single nucleotide polymorphisms (tagSNPs) in F3 (TF) (n=6) and TFPI (n=18), and clinicopathological characteristics and molecular tumor subtypes were explored in 152 treatment naive breast cancer patients. The effect of tumor expressed TF and TFPIα and TFPIβ on survival was investigated in a merged breast cancer dataset of 1881 patients.. Progesterone receptor negative patients had higher mRNA expression of total TFPI (α+β) (P=0.021) and TFPIβ (P=0.014) in tumors. TF mRNA expression was decreased in grade 3 tumors (P=0.003). In plasma, total TFPI levels were decreased in patients with larger tumors (P=0.013). SNP haplotypes of TFPI, but not TF, were associated with specific clinicopathological characteristics like tumor size (odds ratio (OR) 3.14, P=0.004), triple negativity (OR 2.4, P=0.004), lymph node spread (OR 3.34, P=0.006), and basal-like (OR 2.3, P=0.011) and luminal B (OR 3.5, P=0.005) molecular tumor subtypes. Increased expression levels of TFPIα and TFPIβ in breast tumors were associated with better outcome in all tumor subtypes combined (P=0.007 and P=0.005) and in multiple subgroups, including lymph node positive subjects (P=0.006 and P=0.034).. This study indicates that genetic and phenotypic variation of both TFPIα and TFPIβ, more than TF, are markers of cancer progression. Together with the previously demonstrated tumor suppressor effects of TFPI, the beneficial effect of tumor expressed TFPI on survival, renders TFPI as a potential anticancer agent, and the clinical significance of TFPI in cancer deserves further investigation.

Topics: Adult; Aged; Alleles; Biomarkers, Tumor; Breast Neoplasms; Female; Gene Expression; Genetic Association Studies; Genotype; Haplotypes; Humans; Lipoproteins; Middle Aged; Neoplasm Grading; Neoplasm Metastasis; Neoplasm Staging; Phenotype; Polymorphism, Single Nucleotide; Prognosis; RNA, Messenger; Thromboplastin; Tumor Burden

Cancer is often associated with an increased risk of thrombotic events which are exacerbated by treatment with chemotherapeutics such as cyclosphosphamide (CP). Evidence suggests that thrombin can stimulate tumor progression via formation of fibrin and activation of protease-activated receptors (PARs) and platelets. We examined the effect of co-treatment with CP and dabigatran etexilate, a direct inhibitor of thrombin, using the murine orthotopic 4T1 tumor model. Mice receiving co-treatment with both low dose CP and dabigatran etexilate had significantly smaller mammary tumors and fewer lung metastases than mice treated with CP or dabigratran etexilate alone. Co-treatment with dabigatran etexilate and low dose CP also significantly decreased the number of arginase(+)Gr-1(+)CD11b(+) myeloid derived suppressor cells as well as levels of TGF-β in spleens from tumor bearing mice. 4T1 tumors express procoagulant tissue factor (TF) and spontaneously release TF(+) microparticles which are potent procoagulant factors that promote thrombin generation. Treatment with dabigatran etexilate alone prevented tumor-induced increases in circulating TF(+) microparticles and also decreased the numbers of tumor-induced activated platelets by 40%. These results show that co-treatment with dabigatran etexilate and CP synergistically inhibits growth and metastasis of mammary tumors, suggesting that oral administration of the thrombin inhibitor dabigatran etexilate may be beneficial in not only preventing thrombotic events in cancer patients but also in treating malignant tumors themselves.

Topics: Animals; Antineoplastic Agents, Alkylating; Antithrombins; Cell Line, Tumor; Cell-Derived Microparticles; Cyclophosphamide; Dabigatran; Disease Models, Animal; Disease Progression; Drug Synergism; Female; Mice; Myeloid Cells; Neoplasm Metastasis; Neoplasms; Platelet Activation; Thrombin; Thromboplastin

Highly elevated microparticle (MP)-associated tissue factor (TF) activity was found in patients with pancreatic cancer, one of the most prothrombotic malignancies. It remains to be elucidated whether MP-TF activity reflects the prothrombotic state in these patients. MP-TF activity levels and the TF-dependent and -independent effect of MPs on fibrin clot formation were determined in patients with metastatic pancreatic cancer (n = 27), in healthy individuals (n = 10) and in plasma samples from lipopolysaccharide (LPS)-stimulated blood (LPS-plasma), which is rich in monocyte-derived TF-bearing MPs. The median MP-TF activity was 1.06 pg/mL (range, from 0.19 to 10.34 pg/mL) in patients with pancreatic cancer, 0.61 pg/mL (range, from 0.36 to 0.79 pg/mL) in LPS-plasma, and 0.18 pg/mL (range, from 0.04 to 0.39 pg/mL) in healthy individuals. MPs derived from LPS-plasma had the strongest impact on fibrin clot formation time (median, 157.6 seconds; range, from 149.5 to 170.4 seconds). Fibrin clot formation occurred significantly later in MPs derived from patients with pancreatic cancer (median, 273.4 seconds; range, from 146.6 to 354.4 seconds; P < 0.001) and in healthy individuals (median, 299.0 seconds; range, from 261.1 to 417.9 seconds; P < 0.001). Only in MPs derived from LPS-plasma the fibrin clot formation time dependent strongly on TF (median prolongation after TF blockade: 68% in LPS-plasma, 10% in patients with pancreatic cancer, and 4% in healthy individuals). In conclusion, highly elevated MP-TF activity was found in patients with metastatic pancreatic cancer, but TF-bearing MPs had a small effect on fibrin clot formation. TF-bearing MPs might not be the main mediators of the prothrombotic state associated with pancreatic cancer. However, the small but significant increase in coagulation potential by TF-bearing MPs might contribute to the multifactorial pathogenesis of venous thromboembolism in pancreatic cancer.

Topics: Adenocarcinoma; Adult; Aged; Aged, 80 and over; Case-Control Studies; Female; Fibrin; Humans; Male; Middle Aged; Neoplasm Metastasis; Pancreatic Neoplasms; Thromboplastin; Thrombosis

Topics: Adenocarcinoma; Female; Fibrin; Humans; Male; Neoplasm Metastasis; Pancreatic Neoplasms; Thromboplastin; Thrombosis

Microvesicles (MV) in the blood stream are associated with distant metastasis in cancer. Platelet or endothelial cell-related MV actively participate in thrombogenesis, which is an important step in cancer metastasis. This study investigated the correlations between MV levels of platelet-poor plasma and distant metastasis in lung cancer.. Platelet-poor plasma from 44 treatment-naive lung cancer (23 with distant metastasis) and 19 normal subjects was used to determine the levels of glycoprotein Iβ (CD42) + platelet MV (PMV), P-selectin (CD62P) + PMV, VE-cadherin (CD144) + endothelial MV (EMV), tissue factor (CD142) + MV, thrombin-antithrombin complex and vascular endothelial growth factor (VEGF).. The level of CD142 + MV was significant (odds ratio 5.86, 95 % confidence interval 1.31-38.3) in predicting distant metastasis in lung cancer, and a cutoff value of 2.668 (after logarithm transformation) in the ROC curve had a specificity of 90 % and a sensitivity of 59 %. The presence of distant metastasis showed a significant correlation between CD144 + EMV and VEGF, but not between CD144 + EMV and CD42 + PMV or CD62P + PMV in lung cancer subjects.. The finding of CD142 + MV in platelet-poor plasma may be useful for suggesting distant metastasis in lung cancer. In addition to thrombogenesis, interaction between VE-cadherin and VEGF may be needed for successful metastasis in lung cancer.

Topics: Aged; Case-Control Studies; Cytoplasmic Vesicles; Female; Flow Cytometry; Humans; Lung Neoplasms; Male; Neoplasm Metastasis; ROC Curve; Thromboplastin

Tumor cell tissue factor (TF)-initiated coagulation supports hematogenous metastasis by fibrin formation, platelet activation and monocyte/macrophage recruitment. Recent studies identified host anticoagulant mechanisms as a major impediment to successful hematogenous tumor cell metastasis.. Here we address mechanisms that contribute to enhanced metastasis in hyperthrombotic mice with functional thrombomodulin deficiency (TM(Pro) mice).. Pharmacological and genetic approaches were combined to characterize relevant thrombin targets in a mouse model of experimental hematogenous metastasis.. TF-dependent, but contact pathway-independent, syngeneic breast cancer metastasis was associated with marked platelet hyperreactivity and formation of leukocyte-platelet aggregates in immune-competent TM(Pro) mice. Blockade of CD11b or genetic deletion of platelet glycoprotein Ibα excluded contributions of these receptors to enhanced platelet-dependent metastasis in hyperthrombotic mice. Mice with very low levels of the endothelial protein C receptor (EPCR) did not phenocopy the enhanced metastasis seen in TM(Pro) mice. Genetic deletion of the thrombin receptor PAR1 or endothelial thrombin signaling targets alone did not diminish enhanced metastasis in TM(Pro) mice. Combined deficiency of PAR1 on tumor cells and the host reduced metastasis in TM(Pro) mice.. Metastasis in the hyperthrombotic TM(Pro) mouse model is mediated by platelet hyperreactivity and contributions of PAR1 signaling on tumor and host cells.

Topics: Animals; Base Sequence; Cell Line, Tumor; DNA Primers; Mice; Mice, Inbred C57BL; Neoplasm Metastasis; Platelet Activation; Polymerase Chain Reaction; Thrombin; Thromboplastin; Thrombosis

The metastasis-related molecules CD133, CD44v6 and human tissue factor (TF) have been shown to be associated with tumor invasion and metastasis. This study aimed to determine whether co-expression of these three molecules was associated with metastasis and overall prognosis in pancreatic carcinoma. We analyzed the expression profiles of these three molecules by immunohistochemistry and evaluated the relationship of their expression profiles with metastasis and prognosis in 109 pancreatic carcinomas. The results showed that the expression levels of CD133, CD44v6 and TF were increased in pancreatic carcinoma. Co-expression of CD133, CD44v6 and TF (tri-expression) was also detected in pancreatic carcinoma. Clinical analysis showed that individual expression of CD133, CD44v6 or TF was associated with vessel invasion, lymph node metastasis and liver metastasis, while tri-expression was associated with lymph node metastasis. Survival analysis showed that patients with co-expression of CD133 and TF or tri-expression had lower and the lowest overall survival rates, respectively. Univariate analysis showed that T-factor, lymph node metastasis, TNM stage, and individual levels or tri-expression of CD133, CD44v6 and TF were survival risk factors. Multivariate analysis showed that tri-expression of CD133, CD44v6 and TF was an independent predictor of survival. These results suggest that overexpression of CD133, CD44v6 and TF is associated with pancreatic carcinoma metastasis. Tri-expression of these three molecules may be a useful predictor for pancreatic carcinoma prognosis.

Topics: AC133 Antigen; Adult; Aged; Aged, 80 and over; Antigens, CD; Female; Gene Expression Regulation, Neoplastic; Glycoproteins; Humans; Hyaluronan Receptors; Male; Middle Aged; Neoplasm Grading; Neoplasm Metastasis; Pancreatic Neoplasms; Peptides; Survival Analysis; Survival Rate; Thromboplastin

Hypoxia up-regulated expression of tissue factor (TF) may facilitate tumor cell metastasis, but transcriptional mechanisms remain undefined.. To verify the role of Egr-1 in hypoxia-induced TF expression in breast cancer cells, quantitative PCR and Western blot analysis were performed. The secretion of VEGF under hypoxia was detected by enzyme-linked immunosorbent assay (ELISA). Egr-1 and HIF-1α siRNA were transiently transfected into breast cancer cells to evaluate their specific roles.. The increased Egr-1 expression occurring in hypoxic breast cancer cells can up-regulate TF expression and stimulating protein 1(Sp1) was not responsible for the hypoxia-induced expression of TF. HIF-1α mediated the hypoxia-induced up-regulation of TF expression through vascular endothelial growth factor (VEGF). The regulatory effects of Egr-1 on TF under hypoxia were independent of HIF-1α. Either Egr-1 or HIF-1α was responsible for hypoxic induction of tumor cells adhesion. HIF-1α, but not Egr-1, had a pivotal role in human breast cancer cells invasion. Both Egr-1 and HIF-1α were critical to angiogenesis induced by hypoxic conditions in MDA-MB-231 and HUVEC co-cultures. In nude mice, both Egr-1 and HIF-1α small interfering RNA (siRNA) could decrease extravasation of MDA-MB-435 cells in the lung after tail vein injection.. Hypoxia-induced expression of TF in human breast cancer cells depends on Egr-1 and HIF-1α, and both of these proteins may play an important role in breast cancer metastasis, either directly or indirectly through the TF pathway.

Topics: Animals; Breast Neoplasms; Carcinoma; Cell Hypoxia; Cells, Cultured; Coculture Techniques; Early Growth Response Protein 1; Female; Gene Expression Regulation, Neoplastic; Human Umbilical Vein Endothelial Cells; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Mice; Mice, Nude; Neoplasm Metastasis; Thromboplastin

Holothurian glycosaminoglycan (hGAG) is a high-molecular-weight form of fucosylated chondroitin sulfate and has an antithrombotic effect. Our previous studies demonstrated that hGAG efficiently inhibited tumor cell metastasis. The interplays between thrombosis and tumor progression may have a major impact on hematogenous metastasis. In this study, we demonstrated that the mouse melanoma B16F10 cells treated with hGAG displayed a significant reduction of metastasis and coagulation capacity in vitro and in vivo. Mechanistic studies revealed that hGAG treatment in B16F10 cells remarkably inhibited the formation of fibrin through attenuating the generation of activated Factor Xa (FXa), without affecting the expression of urokinase (uPA) and plasminogen activator inhibitor 1 (PAI-1) that involved in fibrinolysis. Moreover, hGAG treatment downregulated the transcription and protein expression of tissue factor (TF). Promoter deletions, site mutations and functional studies identified that the nuclear transcription factor NF-κB binding region is responsible for hGAG-induced inhibition of TF expression. While the hGAG treatment of B16F10 cells was unable to inhibit NF-κB expression and phosphorylation, hGAG significantly prevented nuclear translocation of NF-κB from the cytosol, a potential mechanism underlying the transcriptional suppression of TF. Moreover, hGAG markedly suppressed the activation of p38MAPK and ERK1/2 signaling pathways, the central regulators for the expression of metastasis-related matrix metalloproteinases (MMPs). Consequently, hGAG exerts a dual function in the inhibition of metastasis and coagulation activity in mouse melanoma B16F10 cells. Our studies suggest hGAG to be a promising therapeutic agent for metastatic cancer treatment.

Topics: Animals; Calcium; Factor Xa; Factor Xa Inhibitors; Gene Expression Regulation, Neoplastic; Glycosaminoglycans; Holothurin; Humans; Melanoma; Melanoma, Experimental; Mice; Neoplasm Metastasis; NF-kappa B; Phosphorylation; Signal Transduction; Thromboplastin; Thrombosis

Malignant pleural effusion (MPE) is a poor prognostic sign for patients with lung cancer. Tissue factor (TF) is a coagulation factor that participates in angiogenesis and vascular permeability and is abundant in MPE. We previously demonstrated that autocrine IL-6-activated Stat3 contributes to tumor metastasis and upregulation of VEGF, resulting in the generation of MPE in lung adenocarcinoma. In this study, we found IL-6-triggered Stat3 activation also induces TF expression. By using pharmacologic inhibitors, it was shown that JAK2 kinase, but not Src kinase, contributed to autocrine IL-6-induced TF expression. Inhibition of Stat3 activation by dominant negative Stat3 (S3D) in lung adenocarcinoma suppressed TF-induced coagulation, anchorage-independent growth in vitro, and tumor growth in vivo. Consistently, knockdown of TF expression by siRNA resulted in a reduction of anchorage-independent growth of lung adenocarcinoma cells. Inhibition of TF expression also decreased the adhesion ability of cancer cells in normal lung tissues. In the nude mouse model, both lung metastasis and MPE generation were decreased when PC14PE6/AS2-siTF cells (TF expression was silenced) were intravenously injected. PC14PE6/AS2-siTF cells also produced less malignant ascites through inhibition of vascular permeability. In summary, we showed that TF expression plays a pivotal role in the pathogenesis of MPE generation via regulating of tumor metastasis and vascular permeability in lung adenocarcinoma bearing activated Stat3.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Animals; Blotting, Western; Capillary Permeability; DNA Primers; Flow Cytometry; Fluorescent Antibody Technique; Gene Knockdown Techniques; Interleukin-6; Janus Kinase 2; Luciferases; Lung Neoplasms; Mice; Mice, Nude; Neoplasm Metastasis; Pleural Effusion, Malignant; Reverse Transcriptase Polymerase Chain Reaction; RNA, Small Interfering; STAT3 Transcription Factor; Thromboplastin; Up-Regulation

Topics: Antineoplastic Combined Chemotherapy Protocols; Bleomycin; Cell-Derived Microparticles; Cisplatin; Etoposide; Humans; Male; Neoplasm Metastasis; Testicular Neoplasms; Thromboplastin

Tissue factor (TF), a 47-kDa transmembrane glycoprotein that initiates blood coagulation when complexed with factor VIIa (FVIIa), is expressed in several tumor types. TF has been shown to play a role in cell signaling, inflammation, angiogenesis, as well as tumor growth and metastasis. Activation of the TF signaling pathway has been implicated in mediating the function of many tumor cell types and has led to TF as a potential target in the treatment of several malignancies. Formation of the TF-FVIIa complex in breast cancer cells has been shown to exert an antiapoptotic effect and play a key role in tumor growth and metastasis. Breast cancer growth is suppressed by inhibition of TF-mediated PAR2 signaling, and deficiency in PAR2 delays spontaneous breast cancer development in mice. TF is expressed in triple-negative breast cancer (TNBC), an aggressive type of breast cancer in which there is currently a paucity of available targets. Various methods of targeting TF have been investigated and include immunoconjugates or icons, anti-TF antibodies, TF pathway inhibitors, targeted photodynamic therapy, and microRNAs. These investigations may give way to promising clinical therapies for breast cancer, especially in TNBC, for which there are relatively few effective treatment options.

Topics: Animals; Breast Neoplasms; Factor VIIa; Female; Humans; Mice; Molecular Targeted Therapy; Neoplasm Metastasis; Neovascularization, Pathologic; Receptor, PAR-2; Signal Transduction; Thromboplastin

Tissue factor (TF) expression by tumor cells correlates with metastasis clinically and supports metastasis in experimental settings. However, the precise pathways coupling TF to malignancy remain incompletely defined. Here, we show that clot formation by TF indirectly enhances tumor cell survival after arrest in the lung, during experimental lung metastasis, by recruiting macrophages characterized by CD11b, CD68, F4/80, and CX(3)CR1 (but not CD11c) expression. Genetic or pharmacologic inhibition of coagulation, by either induction of TF pathway inhibitor ex-pression or by treatment with hirudin, respectively, abrogated macrophage recruitment and tumor cell survival. Furthermore, impairment of macrophage function, in either Mac1-deficient mice or in CD11b-diphtheria toxin receptor mice in which CD11b-positive cells were ablated, decreased tumor cell survival without altering clot formation, demonstrating that the recruitment of functional macrophages was essential for tumor cell survival. This effect was independent of NK cells. Moreover, a similar population of macrophages was also recruited to the lung during the formation of a premetastatic niche. Anticoagulation inhibited their accumulation and prevented the enhanced metastasis associated with the formation of the niche. Our study, for the first time, links TF induced coagulation to macrophage recruitment in the metastatic process.

Topics: Animals; Blood Coagulation; Cell Movement; Cell Survival; Cells, Cultured; Humans; Macrophages; Melanoma, Experimental; Mice; Mice, Inbred C57BL; Mice, SCID; Mice, Transgenic; Monocytes; Neoplasm Metastasis; Neoplasms; Neoplastic Stem Cells; Stem Cell Niche; Thromboplastin

Osteopontin (OPN) is a secreted phosphoprotein often overexpressed at high levels in the blood and primary tumors of breast cancer patients. OPN contains two integrin-binding sites and a thrombin cleavage domain located in close proximity to each other.. To study the role of the thrombin cleavage site of OPN, MDA-MB-468 human breast cancer cells were stably transfected with either wildtype OPN (468-OPN), mutant OPN lacking the thrombin cleavage domain (468-ΔTC) or an empty vector (468-CON) and assessed for in vitro and in vivo functional differences in malignant/metastatic behavior.. All three cell lines were found to equivalently express thrombin, tissue factor, CD44, αvβ5 integrin and β1 integrin. Relative to 468-OPN and 468-CON cells, 468-ΔTC cells expressing OPN with a deleted thrombin cleavage domain demonstrated decreased cell adhesion (p < 0.001), decreased mRNA expression of MCAM, maspin and TRAIL (p < 0.01), and increased uPA expression and activity (p < 0.01) in vitro. Furthermore, injection of 468-ΔTC cells into the mammary fat pad of nude mice resulted in decreased primary tumor latency time (p < 0.01) and increased primary tumor growth and lymph node metastatic burden (p < 0.001) compared to 468-OPN and 468-CON cells.. The results presented here suggest that expression of thrombin-uncleavable OPN imparts an early tumor formation advantage as well as a metastatic advantage for breast cancer cells, possibly due to increased proteolytic activity and decreased adhesion and apoptosis. Clarification of the mechanisms responsible for these observations and the translation of this knowledge into the clinic could ultimately provide new therapeutic opportunities for combating breast cancer.

Topics: Animals; Binding Sites; Blotting, Western; Breast Neoplasms; Cell Adhesion; Cell Line, Tumor; Cell Movement; Cell Proliferation; Female; Gene Expression Regulation, Neoplastic; Humans; Integrins; Mammary Neoplasms, Experimental; Mice; Mice, Nude; Neoplasm Metastasis; Osteopontin; Reverse Transcriptase Polymerase Chain Reaction; Sequence Deletion; Serpins; Thrombin; Thromboplastin; TNF-Related Apoptosis-Inducing Ligand; Transfection; Transplantation, Heterologous

To evaluate the effect of tissue factor (TF) in extravascular migration of fibrosarcoma cells and hematogenous metastasis.. The expression of tissue factor in fibrosarcoma HT1080 cells was analyzed by flow cytometry. The extravascular migration of fibrosarcoma cells was observed in a constructed monolayer vascular endothelial cells and extra-cellular matrix model.. Tissue factor was highly expressed in HT1080 cells. HT1080 migrated and passed through the monolayer vascular endothelial cells to the collagen gel in a time-dependent manner. Anti-TF antibody inhibited extravascular migration of fibrosarcoma cells and the inhibition was concentration-dependent (P<0.05).. Tissue factor may enhance hematogenous metastasis through extravascular migration of fibrosarcoma cells.

Topics: Cell Movement; Cells, Cultured; Endothelial Cells; Fibrosarcoma; Humans; Neoplasm Metastasis; Thromboplastin

The coagulation function in carcinoma patients is abnormal, but in renal cell carcinoma the extent and relationships of coagulation function remain unclear. This study retrospectively investigated the relationships between coagulation function, clinical stage and metastasis in patients with renal cell carcinoma.. A total of 350 consecutive patients admitted to our Urology Department from 2004 to 2010 were diagnosed with renal cell carcinoma by histopathologic examination and were included in this study. A total of 231 cases of renal benign tumors were considered as the control group. Fibrinogen, prothrombin time, activated partial thromboplastin time and international normalized ratio were evaluated in all subjects. Tumor size, clinical stage, lymph node metastasis, and distant metastasis were evaluated using radiologic imaging, intraoperative findings, and histological studies.. The preoperative plasma fibrinogen levels of patients with renal cell carcinoma ((383.9 ± 146.7) mg/dl) were significantly higher than those of the control group ((316.7 ± 62.0) mg/dl) (P < 0.01). We divided the renal cell carcinoma group into stages Ia, Ib, II, III, and IV. The fibrinogen values were (315.6 ± 64.6) mg/dl, (358.3 ± 91.1) mg/dl, (465.6 ± 164.7) mg/dl, (500.0 ± 202.1) mg/dl, and (585.8 ± 179.7) mg/dl, respectively. There were no significant differences in fibrinogen values between stage Ia and control groups. However, results of other stages showed significant differences when compared to control group values (P < 0.01). Using the cutoff value of 440 mg/dl, which defines hyperfibrinogenemia, plasma fibrinogen levels had a positive predictive value of 39.8% and a negative predictive value of 93.3% for predicting distant metastasis, with a sensitivity of 64.7% and specificity of 83.3%.. Preoperative plasma fibrinogen levels are elevated in patients with renal cell carcinoma with distant metastasis or lymph node metastasis. Potential metastasis is more likely if the tumor size larger than 4 cm. Increased preoperative plasma fibrinogen levels, especially hyperfibrinogenemia, may be an indicator of metastasis.

Topics: Adult; Aged; Blood Coagulation; Carcinoma, Renal Cell; Female; Fibrinogen; Humans; Kidney Neoplasms; Lymphatic Metastasis; Male; Middle Aged; Neoplasm Metastasis; Neoplasm Staging; Retrospective Studies; Thromboplastin

Tumor-associated macrophage infiltration and up-regulation of tissue factor-factor VII (TF-FVIIa) complex have been observed in the peritoneum and stroma of epithelial ovarian cancer (EOC). However, it is not clear how tumor-associated macrophage and TF-FVIIa complex promotes EOC invasion. In the present study, we aimed to determine the mechanism by which interaction of TF-FVIIa and monocytes (MOs) promotes EOC metastasis.. Matrigel invasion assay was used to analyze the potential of EOC metastasis. Enzyme-linked immunosorbent assay and real-time polymerase chain reaction were used to detect expressions of cytokines and chemokines. Fluorescence-activated cell sorting was used to count the percentage of CD14, CD68, and CD163 of MOs.. We found that the TF-FVIIa complex caused dynamic changes in MOs cytokine and chemokine expression. CD14 and CD163 were also upregulated on MOs by TF-FVIIa. Epithelial ovarian cancer cells were cocultured with TF-FVIIa-stimulated MOs, demonstrating increased invasion potential. Interleukin 8 (IL-8) was proposed as the major chemoattractant mediating EOC invasion based on MOs messenger RNA and protein expression profiles. Anti-IL-8 monoclonal neutralizing antibody attenuated EOC cell invasion in a concentration-dependent manner, and tumor necrosis factor α from TF-FVIIa-stimulated MOs was observed to amplify IL-8 production. The following transcription factors in MOs were activated by TF-FVIIa and inhibited by the tissue factor pathway inhibitor: oncogenes HIF-1α, HIF-1β, Oct I, Oct II, and Egr-1; inflammatory mediators c-Fos and c-Rel; and STAT family members STAT5A and STAT5B.. Our study suggested that the interaction between the TF-FVIIa complex might play a role in mediating EOC invasion and metastasis depending on MOs mechanism.

Topics: Carcinoma, Ovarian Epithelial; Cell Differentiation; Cell Line, Tumor; Cells, Cultured; Factor VIIa; Female; Gene Expression Regulation, Neoplastic; Humans; Monocytes; Multiprotein Complexes; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasms, Glandular and Epithelial; Ovarian Neoplasms; Signal Transduction; Thromboplastin; Up-Regulation

Tissue factor (TF), the initiating cell surface receptor for the blood coagulation cascade, plays an important role in malignant transformation of the pancreas, although the precise mechanism remains unresolved. Here, we report that the TF - factor VIIa complex in human pancreatic cancer cells produced a significant amount of MMP-9 and promoted invasion ability in vitro and invasion and metastasis in vivo. For treatment, we successfully developed an anti-human TF monoclonal antibody that inhibits both cellular signalling and blood coagulation cascade via TF. Invasive capability and MMP-9 expression were significantly reduced by the antibody. The antibody inhibited not only tumour invasion in the orthotopic model, but also haematogenous metastasis in the portal-injection liver metastasis model. In conclusion, the TF-VIIa complex plays an important role in invasion-metastasis by enhancing tumour cell infiltration ability and forming microthrombi. The newly established anti-human TF neutralisation antibody may be useful for the treatment of pancreatic and other invasive cancers.

Topics: Antibodies, Monoclonal; Blood Coagulation; Cell Transformation, Neoplastic; Factor VIIa; Humans; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasm Proteins; Pancreatic Neoplasms; Second Messenger Systems; Signal Transduction; Thromboplastin; Tumor Cells, Cultured

Numerous studies indicate that tissue factor (TF), namely tissue thromboplastin, has a close relationship with malignant tumor genesis and progress. It contributes to blood coagulation as well as the regulation of cellular differentiation, the formation of blood vessels, and also tumor recurrence and metastasis. The present study aimed to detect TF expression in hepatocellular carcinoma (HCC) patients and to elucidate its association with prognosis and clinical features of the disease.. The plasma TF levels of 50 HCC patients and 30 controls were assayed by ELISA. The expressions of TF mRNA and protein in HCC tissues, adjacent tissues and normal tissues were detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting. The acquired data were analyzed with related clinic-pathological documents. The patients were followed up for five years, and the relationship between TF and prognosis was analyzed.. The plasma TF levels were significantly increased in HCC compared to the controls (P < 0.05), presenting a close relationship with differentiation level, tumor size and hepatocirrhosis occurrence (P < 0.05). There were remarkably higher values in cases of lymphatic metastasis, extrahepatic metastasis and portal tumor thrombus (PTT) (P < 0.05) compared to non-metastasis or non-tumor thrombus, but no significant difference with different focus number or envelope (P > 0.05). The positive rates and the relative expression of TF mRNA in HCC tissue were 63.0% (17/27) and 0.567 ± 0.268, respectively, significantly higher than that in adjacent tissues or normal tissues (P < 0.05). In the patients with positive results, the relative expression intensity varied significantly with different tumor size and index of local invasion and metastasis (P < 0.05). The positive rates and the relative expression intensities of TF protein in HCC tissue were 74.1% (20/27) and 4.093 ± 1.256, respectively, significantly higher than those in adjacent tissue or normal tissue (P < 0.05). In the patients with positive results, the relative expression intensity showed significant difference in different tumor size, differentiation level, and index of local invasion and metastasis (P < 0.05).. The TF levels were significantly higher in plasma and tissues of HCC patients, presenting a close relationship with the index of invasion and metastasis. It indicated that TF might be related to differentiation and metastasis of HCC.

Topics: Adult; Aged; Blotting, Western; Carcinoma, Hepatocellular; Enzyme-Linked Immunosorbent Assay; Female; Humans; Liver Neoplasms; Male; Middle Aged; Neoplasm Metastasis; Reverse Transcriptase Polymerase Chain Reaction; Thromboplastin

It is well established that the blood coagulation system is activated in cancer. In addition, there is considerable evidence to suggest that clotting activation plays an important role in the biology of malignant tumors, including the process of blood-borne metastasis. For many years our laboratory has used experimental models of lung metastasis to study the events that follow the introduction of procoagulant-bearing tumor cells into circulating blood. This chapter focuses on the basic methods involved in assessing the anti-metastatic effects of anticoagulants and anti-platelet agents using rodent models of experimental metastasis. In addition, it summarizes our experience with these models, which collectively suggests that intravascular coagulation and platelet activation are a necessary prelude to lung tumor formation and that interruption of coagulation pathways or platelet aggregation may be an effective anti-metastatic strategy.

Topics: Animals; Anticoagulants; Antineoplastic Agents; Blood Coagulation; Cell Line, Tumor; Enoxaparin; Humans; Lung Neoplasms; Mice; Neoplasm Metastasis; Neoplasms, Experimental; Platelet Aggregation Inhibitors; Thromboplastin; Warfarin

Tongue cancer metastasis is mainly through blood stream and possibly associated with tumor cell-induced platelet aggregation (TCIPA).. Platelet aggregation was induced by different amounts of SAS tongue cancer cells with/without inhibitors and the latent period for induction of platelet aggregation was recorded. Gene expression was analyzed by reverse transcriptase-polymerase chain reaction.. SAS cells (4 x 10(4) to 1 x 10(6) cells/ml) induced platelet aggregation in a cell density-dependent manner. The latent period for induction of platelet aggregation reduced from 11.3 min (2 x 10(5) cells/ml) to 0.9 min (5 x 10(5) cells/ml). The extent of platelet aggregation increased from 39% to 76% by 2 x 10(5) and 5 x 10(5) SAS cells. Pre-treatment of SAS cells with aspirin showed little effect on its induction of platelet aggregation. SAS cells expressed tissue factor (TF) mRNA and the SAS cells-induced TCIPA was inhibited by TF neutralization antibody (5-20 microg/ml), heparin (5-10 U/ml), Hirudin fragment 54-65 (50 microg/ml) and D-Phenylalanyl-L-prolyl-L-arginine chloromethyl ketone. But areca nut (AN, a betel quid component known to generate reactive oxygen species (ROS)) extract showed little effect on TF expression in SAS cells. Pre-treatment with U73122 and 2-aminoethoxydiphenylborate inhibited SAS-induced TCIPA. Interestingly, catalase suppressed SAS cells-induced TCIPA, whereas AN extract enhanced this event.. These results suggest that tongue cancer cells may induce TCIPA and enhance tumor metastasis. SAS-induced TCIPA is related to TF secretion, thrombin generation and associated with Phospholipase C-Inositol triphosphate signaling and ROS production. Betel quid chewing may potentially promote tongue cancer metastasis.

Topics: Areca; Coculture Techniques; Epithelial Cells; Gene Expression Profiling; Gingiva; Humans; Mouth Mucosa; Neoplasm Metastasis; Neoplasm Proteins; Plant Extracts; Platelet Aggregation; RNA, Messenger; Second Messenger Systems; Signal Transduction; Statistics, Nonparametric; Thromboplastin; Time Factors; Tongue Neoplasms; Tumor Cells, Cultured

Expression of the principal initiator of coagulation, tissue factor (TF), by colorectal cancer (CRC) cells is involved in tumoral angiogenesis and metastasis progression, after binding of factor VIIa (FVIIa) to TF and generation of TF-FVIIa activity. We thus hypothesized that inhibition of the TF pathway by active site-blocked FVIIa (FFR-FVIIa) may prevent the development of hepatic metastasis in CRC.. Rat tumoral cells (DHDK12 proB cells) expressing high levels of TF were injected in the portal vein in syngenic BDIX rats. Rats received intraperitoneal injection of either FFR-FVIIa, from d 3 to d 7 (adjuvant treatment) (n = 19), or solvent buffer (n = 18) (control group). Additionally, cancer cells were infused subcutaneously in 20 other rats, which were assigned to FFR-FVIIa adjuvant treatment (n = 10), or buffer treatment (n = 10). Macroscopic and histological analysis was performed at d 14.. In the control group, infusion of cancer cells resulted in development of macroscopic hepatic tumors in 17/18 rats. In the adjuvant FFR-FVIIa group, macroscopic hepatic tumors were visible on the liver surface in 3/19 rats (P = 0.002 versus control). All rats with subcutaneous injection of proB cells exhibited macroscopic tumors, with no significant difference between the control and the treated ones.. Inhibition of the proteolytic activity of TF-FVIIa complex blunted hematogenous hepatic metastasis, suggesting that TF-FVIIa is a relevant target for the prevention of hepatic metastasis in CRC. TF-blocking agents should be investigated as adjuvant treatment in this setting.

Topics: Animals; Carcinoma; Cell Line, Tumor; Colorectal Neoplasms; Factor VIIa; Liver Neoplasms; Male; Neoplasm Metastasis; Neoplasms, Experimental; Rats; Thromboplastin

To study the expression of coagulation factor VII(FVII)/tissue factor(TF)complex in colorectal carcinoma (CRC)and its correlation with clinicopathologic factor.. The expression of coagulation factor VII protein was studied by immunohistochemistry and Western blot.The expression of tissue factor and coagulation factor VII at the mRNA levels were evaluated by quantitative realtime RT-PCR in 45 cases of CRC.. (1) FVII overexpression was ectopicly detected in CRC specimens at protein level by immunohistochemistry and Western blot, but not in adjacent non-cancerous mucosa of colorectum;(2)FVII protein mainly localized in the cytoplasm of colon cancer cells.The positive ratios of FVII protein expression in stages I, II, III and IV by immunohistochemistry assay were 33.3%, 40.0%, 64.7% and 80.0% respectively(P=0.001); (3)The expression of FVII mRNA in CRC with hepatic metastasis was significantly higher than that in CRC without hepatic metastasis.The relative expression was 5.33+/-2.88 and 1.47+/-0.51 respectively(P=0.03). Overexpression FVII gene was unrelated with tumor size, differentiation, depth of invasion, lymph node metastasis and TNM staging.There existed some relation between the gene and protein level by Spearman correlation, r=0.58, P=0.003;(4)The expression of TF mRNA in CRC significantly correlated with lymph node metastasis, hepatic metastasis and TNM staging.The expression of tissue factor was a critical factor to predict liver metastasis by logistic regression analysis(P=0.001).. Colorectal cancer can ectopicly synthesize coagulation factor VII.Tissue factor expression may play a role in the process of developing hepatic metastasis.The microenvironment of high dose FVII protein may promote tumor metastasis.

Topics: Adult; Aged; Colorectal Neoplasms; Factor VII; Female; Humans; Immunohistochemistry; Liver Neoplasms; Logistic Models; Male; Middle Aged; Neoplasm Metastasis; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Thromboplastin

Non-small cell lung cancer (NSCLC) comprises of 75% of all lung cancers. Human full length tissue factor (flHTF), the physiological initiator of blood coagulation, is aberrantly expressed in certain solid tumors. FlHTF and its soluble isoform, alternatively spliced human tissue factor (asHTF), have been shown to contribute to thrombogenicity of the blood of healthy individuals. The aim of this study was to quantify flHTF and asHTF on mRNA and protein levels (using immunohistochemistry, immunoblotting, and ELISA) on a panel of human NSCLC tissue and plasma specimens. The tissue factor (TF) expression of 21 pulmonary adenomatous (AC) and 12 normal healthy tissues was assessed by real-time qRT-PCR. The TF protein concentration was quantified by ELISA in a subset of 11 AC and 9 normal tissue specimens as well as in the plasma of 13 lung cancer patients and 15 healthy controls. We found a significant increase in the ratio of flHTF/HGAPDH mRNA in AC (0.24+/-0.06 vs. 0.07+/-0.01; p=0.02 vs. controls) and in asHTF/HGAPDH mRNA (0.027+/-0.01 vs. 0.004+/-0.001; p=0.03 AC vs. controls). AsHTF mRNA expression was significantly lower in patients with stage IA disease compared to patients with higher grade stages, pointing to TF as being a marker of malignancy and metastases. TF protein of lung tumors was significantly increased in AC (p=0.004 vs. controls). TF in plasma was up-regulated in lung cancer patients (334.9+/-95.4 vs. 124.1+/-14.8 pg/ml; p=0.02 vs. controls). Immunohistochemical and immunoblotting data are in line with the increased TF expression, showing elevated blood thrombogenicity of NSCLC patients. The up-regulation of flHTF and, especially, asHTF in AC suggests not only a raised risk of thrombosis, but also of tumor progression, thereby, indicating a poor prognosis in these patients.

Topics: Adenocarcinoma; Blotting, Western; Carcinoma, Non-Small-Cell Lung; Humans; Immunohistochemistry; Lung Neoplasms; Neoplasm Metastasis; RNA, Messenger; Thromboplastin; Thrombosis

Increased tissue factor (TF) expression is observed in many types of cancer, associated with more aggressive disease, and in thrombosis. The mechanism by which TF promotes tumor growth remains unclear. Anticoagulation has been shown to result in a trend toward improved survival; no direct antitumor effect has been shown in cancer patients. Alternatively spliced tissue factor (asTF) was recently described, in which exon 5 is deleted. Because of a frame-shift in exon 6, the transmembrane and cytoplasmic domains are replaced with a unique COOH-terminal domain, making asTF soluble. Both alternatively spliced human tissue factor (asHTF) and full-length tissue factor (flTF) are expressed in human pancreatic cancer lines and in pancreatic cancer specimens. We studied the role of asHTF and flTF in a mouse model of pancreatic cancer. Although lacking procoagulant activity, asTF promotes primary growth of human pancreatic cancer cells in mice and augments tumor-associated angiogenesis. This body of work suggests a new paradigm for the role of TF in pancreatic cancer: that asHTF contributes to cancer growth, independent of procoagulant activity.

Topics: Alternative Splicing; Animals; Anticoagulants; Cell Line, Tumor; Humans; Mice; Neoplasm Metastasis; Neovascularization, Pathologic; Pancreatic Neoplasms; Thromboplastin; Transfection

The role of host-derived tissue factor (TF) in tumor growth, angiogenesis, and metastasis has hitherto been unclear and was investigated in this study.. We compared tumor growth, vascularity, and responses to cyclophosphamide (CTX) of tumors in wild-type (wt) mice, or in animals with TF levels reduced by 99% (low-TF mice). Global growth rate of 3 different types of transplantable tumors (LLC, B16F1, and ES teratoma) or metastasis were unchanged in low-TF mice. However, several unexpected tumor/context-specific alterations were observed in these mice, including: (1) reduced tumor blood vessel size in B16F1 tumors; (2) larger spleen size and greater tolerance to CTX toxicity in the LLC model; (3) aborted tumor growth after inoculation of TF-deficient tumor cells (ES TF(-/-)) in low-TF mice. TF-deficient tumor cells grew readily in mice with normal TF levels and attracted exclusively host-related blood vessels (without vasculogenic mimicry). We postulate that this complementarity may result from tumor-vascular transfer of TF-containing microvesicles, as we observed such transfer using human cancer cells (A431) and mouse endothelial cells, both in vitro and in vivo.. Our study points to an important but context-dependent role of host TF in tumor formation, angiogenesis and therapy.

Topics: Animals; Antineoplastic Agents, Alkylating; Carcinoma, Lewis Lung; Cell Line, Tumor; Cell Survival; Cyclophosphamide; Embryonic Stem Cells; Endothelial Cells; Humans; Melanoma, Experimental; Mice; Mice, Inbred C57BL; Mice, SCID; Neoplasm Metastasis; Neoplastic Stem Cells; Neovascularization, Pathologic; Secretory Vesicles; Teratoma; Thromboplastin; Time Factors

Topics: Animals; Antineoplastic Agents; Cell Survival; Embryonic Stem Cells; Endothelial Cells; Humans; Neoplasm Metastasis; Neoplasms, Experimental; Neoplastic Stem Cells; Neovascularization, Pathologic; Secretory Vesicles; Thromboplastin

ErbB oncogenes drive the progression of several human cancers. Our study shows that in human carcinoma (A431) and glioma (U373) cells, the oncogenic forms of epidermal growth factor receptor (EGFR; including EGFRvIII) trigger the up-regulation of tissue factor (TF), the transmembrane protein responsible for initiating blood coagulation and signaling through interaction with coagulation factor VIIa. We show that A431 cancer cells in culture exhibit a uniform TF expression profile; however, these same cells in vivo exhibit a heterogeneous TF expression and show signs of E-cadherin inactivation, which is coupled with multilineage (epithelial and mesenchymal) differentiation. Blockade of E-cadherin in vitro, leads to the acquisition of spindle morphology and de novo expression of vimentin, features consistent with epithelial-to-mesenchymal transition. These changes were associated with an increase in EGFR-dependent TF expression, and with enhanced stimulation of vascular endothelial growth factor production, particularly following cancer cell treatment with coagulation factor VIIa. In vivo, cells undergoing epithelial-to-mesenchymal transition exhibited an increased metastatic potential. Furthermore, injections of the TF-blocking antibody (CNTO 859) delayed the initiation of A431 tumors in immunodeficient mice, and reduced tumor growth, vascularization, and vascular endothelial growth factor expression. Collectively, our data suggest that TF is regulated by both oncogenic and differentiation pathways, and that it functions in tumor initiation, tumor growth, angiogenesis, and metastasis. Thus, TF could serve as a therapeutic target in EGFR-dependent malignancies.

Topics: Animals; Cadherins; Carcinoma, Squamous Cell; Cell Differentiation; Cell Line, Tumor; Epithelial Cells; ErbB Receptors; Flow Cytometry; Glioma; Humans; Mesoderm; Mice; Mice, SCID; Neoplasm Metastasis; Neovascularization, Pathologic; Thromboplastin; Up-Regulation; Vascular Endothelial Growth Factor A; Vimentin

We previously reported that overexpression of tissue factor (TF) protected HT29 tumour cells from cellular cytotoxicity through a mechanism requiring the presence of the cytoplasmic domain of TF. In this investigation the mechanism of TF-mediated immune evasion has been examined.. The influence of alanine-substitution at Ser253 and Ser258 of TF (TF(Ala253) and TF( Ala258)) on the induction of cytotoxic evasion, as well as expression of vascular cell adhesion molecule-1 and intra-cellular adhesion molecule-1 (VCAM-1 and ICAM-1) was investigated. Moreover, we examined the effect of transfection of four 20-mer peptides, corresponding to the C-terminal residues of TF, with different phosphorylation states, on promotion of evasion from cell cytotoxicity.. Cells overexpressing TF(Ala258) and to a lesser extent overexpressing TF(Ala253,) exhibited a reduced ability to evade cellular cytotoxicity compared to cells overexpressing the wild-type TF. Furthermore, the increase in protection acquired was greatest on transfection of Ser258-phosphsorylated form of the cytoplasmic peptide, lower in double-phosphorylated and Ser253-phosphorylated peptides respectively, and lowest in the unphosphorylated form. Finally, the expression of VCAM-1 mRNA as well as surface antigen was reduced on overexpression of TF(wt) but was partially reverted in the cells transfected to overexpress TF(Ala253) or TF(Ala258).. These data show that the phosphorylation of TF at Ser258 and to a lesser extent Ser253, plays an essential role in the protective influence of TF on immune evasion by tumour cells, and that the mechanism could involve the downregulation of key surface antigens, such as adhesion proteins, involved in cell:cell interaction.

Topics: Alanine; Cell Line, Tumor; Gene Expression Regulation, Neoplastic; Humans; Immune System; Models, Biological; Mutagenesis, Site-Directed; Neoplasm Metastasis; Peptides; Phosphorylation; Protein Structure, Tertiary; Serine; Thromboplastin; Transfection; Vascular Cell Adhesion Molecule-1

Tumor cell-associated tissue factor (TF) is a powerful determinant of metastatic potential. TF may increase metastasis by supporting thrombin-mediated proteolysis, through intracellular signaling events mediated by the TF cytoplasmic domain, through TF/fVIIa/fXa-mediated activation of protease-activated receptors, or through a combination of these processes. To better define the relationship between tumor cell-associated TF and circulating hemostatic factors in malignancy, we generated a set of C57Bl/6-derived tumor lines genetically lacking TF, expressing wild-type murine TF, or expressing a mutant TF lacking the cytoplasmic domain. Comparison of the metastatic potential of these cells in immunocompetent mice with genetic deficits in prothrombin, platelet function, or fibrinogen revealed that TF supports metastasis through mechanisms independent of the cytoplasmic domain, but dependent on each of these distal hemostatic factors. TF was neither required for primary tumor growth nor necessary for initial localization of embolized tumor cells within the lungs. Rather, tumor cell fate studies indicated TF supports metastasis by increasing the survival of micrometastases. One mechanism linking TF to metastasis is through a fibrin(ogen)-dependent and platelet-dependent restriction in natural killer cell-mediated clearance of micrometastases. However, TF also supported the early success of micrometastases through an additional mechanism independent of natural killer cells, but coupled to circulating prothrombin.

Topics: Animals; Blood Coagulation Factors; Blood Platelets; Cell Line, Tumor; Fibrin; Killer Cells, Natural; Mice; Neoplasm Metastasis; Neoplastic Cells, Circulating; Prothrombin; Thromboplastin

Interaction between platelet and carcinoma cell contributes to pathogenesis of cancer-related thrombosis and metastasis. This study investigated whether physical exercise affects platelet-nasopharyngeal carcinoma cell (NPC) interaction and platelet-promoted tissue factor (TF) and matrix metalloproteinase (MMP) activities of NPC. Thirty sedentary men performed on three occasions moderate-intensity exercise [MIE, 60% maximal oxygen consumption (V(.)o(2max)) for 40 min] and high-intensity exercise (HIE, up to V(.)o(2max)), with and without warm-up exercise (WUE, 60% V(.)o(2max) for 20 min) on a bicycle ergometer. Before and immediately after exercise, platelet-NPC aggregation, the TF, MMP-2 and MMP-9 expressions and activities, and TF pathway inhibitor (TFPI) and tissue inhibitor of MMP-1 levels of NPC and platelet were measured. The results of this study demonstrated that HIE enhanced platelet-NPC aggregation in the presence of fibrinogen and was accompanied by increased platelet-promoted TF activity, expression of NPC, decreased platelet-promoted MMP-2 and MMP-9 activities, and TFPI release of NPC, whereas these alterations to HIE on platelet-NPC interactions were ameliorated by WUE pretreatment. Conversely, MIE reduced the formation of platelet-NPC aggregates, but did not change the TF, TFPI, MMP-2, MMP-9, tissue inhibitor of MMP activities, and/or levels of NPC mediated by platelet. It is concluded that HIE may enhance aggregation and coagulation and reduce MMP bioactivity related to platelet-NPC interactions, by raising the binding affinity to fibrinogen and TF activity and expression and lowering TFPI release and MMP-2 and -9 activities. These effects on HIE diminish after WUE. However, MIE minimizes the risk of thrombosis induced by platelet-NPC interactions.

Topics: Adult; Blood Platelets; Carcinoma; Cell Aggregation; Exercise; Humans; Lipoproteins; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Nasopharyngeal Neoplasms; Neoplasm Metastasis; Thromboplastin; Thrombosis; Tissue Inhibitor of Metalloproteinase-1

We have shown that the thrombin G-protein coupled receptors (GPCR) designated as protease-activated receptors (PAR-1) are expressed in primary cancer cells isolated from peritoneal and pleural malignant effusions. Here, our main goal was to evaluate several coagulation and thrombin activation effectors and markers in a series of 136 malignant effusions from cancer patients with gastrointestinal, lung and mammary carcinomas. All these patients present a highly activated coagulation system in blood and their malignant effusions, as indicated by high levels of prothrombin F1.2 fragments and D-dimers. Notably, we detected in the effusions all the coagulation factors of the tissue factor pathway inducing thrombin activation, namely factors VII, V, X and II, as well as high VEGF levels and IGF-II in mature and precursor forms. Fibrin clot formation also correlated with higher levels of free ionized calcium (iCa), suggesting that iCa and its binding protein albumin are regulatory factors for fibrinogenesis in effusions. Consequently, thrombin, VEGF and IGFII appear to converge in the promotion of survival and invasivity of the metastatic cancer cells from blood to the malignant effusions. Thus, we add new insights on the interconnections between blood coagulation disorders in cancer patients and thrombin activation in malignant effusions, including their functional interaction with PAR in metastatic cancer cells. Based on these data we propose to counteract the metastatic cascades by targeted invalidation of key effectors of the coagulation system. Therefore, potential therapeutic approaches include the application of thrombin protease inhibitors, VEGF-blocking antibodies, and drugs targeting the VEGF and thrombin signaling pathways, such as tyrosine kinase or GPCR inhibitors.

Topics: Aged; Antineoplastic Agents; Antithrombins; Ascitic Fluid; Blood Coagulation; Blood Coagulation Factors; Breast Neoplasms; Calcium; Case-Control Studies; Factor V; Factor VII; Factor X; Female; Fibrin; Fibrin Fibrinogen Degradation Products; Gastrointestinal Neoplasms; Humans; Insulin-Like Growth Factor II; Lung Neoplasms; Male; Middle Aged; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasms; Peptide Fragments; Pericardial Effusion; Pleural Effusion, Malignant; Proteins; Prothrombin; Serum Albumin; Thrombin; Thromboplastin; Vascular Endothelial Growth Factor A

Topics: Animals; Blood Coagulation; Hemostasis; Humans; Immunity, Innate; Mice; Neoplasm Metastasis; Thromboplastin

Tumor development depends on multiple reciprocal interactions of tumor cells with the host cell compartment. Tumor cells initiate TF-dependent crosstalks with the tumor microenvironment by releasing procoagulant microparticles, soluble cytokines and angiogenic growth factors. Conversely, the hemostatic system in the host compartment provides multiple circuits that regulate tumor growth and sustain angiogenesis. A combination of experimental models of spontaneous and transplanted tumor development and metastasis start to delineate the role of TF in tumor progression and identified potential therapeutic approaches to target the TF pathway.

Topics: Animals; Disease Progression; Humans; Mice; Neoplasm Metastasis; Neoplasms; Neovascularization, Pathologic; Receptors, Proteinase-Activated; Signal Transduction; Thromboplastin

The association between tissue factor (TF) expression and increased rate of tumour metastasis is well established. In this study, we have examined the hypothesis that the expression of TF by disseminated tumour cells confers protection against immune recognition and cytotoxicity.. A hybrid EGFP-TF protein was expressed in HT29 colon carcinoma and K562 lymphoblast cell lines. To assess the cytotoxic activity against tumour cells over-expressing TF, a novel method was used, based on the direct measurement of fluorescently labelled HT29 or K562 target cells.. Upon challenge with peripheral blood mononuclear cells (PBMC), tumour cells expressing TF partially evaded cellular cytotoxicity (Delta=15-40% reduction in cytotoxicity). Moreover, the influence of TF was not primarily dependent on its procoagulant function, although the inclusion of 20% (v/v) plasma did lower the rate of cytotoxicity against untransfected cells. However, expression of a truncated form of TF, devoid of the cytoplasmic domain, did not mediate any degree of inhibition of cytotoxicity, suggesting that the protective function of TF is principally due to this domain.. We conclude that TF can promote immune evasion in tumour cells expressing this protein leading to increased survival and therefore metastatic rate in such cells.

Topics: Cell Line, Tumor; Colorectal Neoplasms; Cytoplasm; Gene Expression Regulation, Neoplastic; Humans; K562 Cells; Leukocytes, Mononuclear; Lymphocytes; Neoplasm Metastasis; Protein Structure, Tertiary; Prothrombin Time; Thromboplastin

Topics: Animals; Antibodies, Monoclonal; Baculoviridae; Blood Coagulation; Enzyme-Linked Immunosorbent Assay; Hybridomas; Immunohistochemistry; Lung Neoplasms; Melanoma, Experimental; Mice; Mice, Inbred C57BL; Neoplasm Metastasis; Neoplasm Transplantation; Rats; Recombinant Proteins; Thromboplastin

To explore the role of tissue factor/activated factor VII (TF/FVIIa) complex in human ovarian cancer invasion and metastasis.. (1) Constructed an expression vector of TF, pcDNA3-TF and established a human ovarian cell line A2780/TF expressing high level TF by using molecular cloning and gene transfection techniques. (2) By Boyden chamber assay to count the numbers of A2780 and A2780/TF cells that penetrated the matrigel to the back of PVPF membrane after FVIIa stimulation. (3) BALB/c nude mice were used to establish experimental model of metastasis with A2780 or A2780/TF and the lung tissue sections were examined by microscopy for cancer metastasis.. (1) Compared with their parental A2780 cells, A2780/TF cells expressed high level of TF mRNA (3.99 +/- 0.15 vs 0.97 +/- 0.23, P < 0.01) and TF antigen on cell surface \\[(48.56 +/- 9.53)% vs (2.73 +/- 1.15)%, P < 0.01\\]. (2) After stimulation, the A2780/TF cell number on the back of PVPF membrane increased from basal level 157.3 +/- 19.2 to 447.7 +/- 39.4 (P < 0.01), which could decreased to basal level when coincubated with anti-TF antibody. (3) Cancer metastasis was found in 22.2% of nude mice transplanted with A2780 cells, while in 88.9% of those transplanted with A2780/TF cells.. TF could promote the invasion and metastasis of human ovarian cancer cells through TF/FVIIa pathway.

Topics: Animals; Cell Line, Tumor; Cell Movement; Cloning, Molecular; Factor VIIa; Female; Genetic Vectors; Humans; Mice; Mice, Inbred BALB C; Mice, Nude; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasm Transplantation; Ovarian Neoplasms; Thromboplastin; Transfection; Transplantation, Heterologous

To study the changes of hemostatic molecular markers in patients with gastric or intestinal cancer for elucidating their clinical significance.. The plasma levels of tissue factor (TF), thrombin antithrombin complex (TAT), tissue plasminogen activator (t-PA), urokinase plasminogen activator (u-PA), urokinase plasminogen activator receptor (u-PAR) and plasmin antiplasmin complex (PAP) were measured by ELISA. Gene transcription of TF, t-PA, u-PA mRNA were detected by real-time RT-PCR.. The plasma levels of TF, TAT, u-PA, u-PAR and PAP were elevated in gastric or intestinal cancer patients (P < 0.05), while u-PA, u-PAR remarkably increased in patients with local infiltration, lymph node involvement or distal metastasis (P < 0.01). Plasma level of TF, TAT, PAP were remained higher than control even after surgery. TF, u-PA mRNA were higher (P < 0.01) and t-PA was lower (P > 0.05) in gastric or intestinal cancer compared to normal tissue.. Their existed over expression of TF and u-PA, increasing formation of thrombin and plasminogen in gastric or intestinal cancer patients. Hypercoagulability and hyperfibrinolysis were important factors related with metastasis potential of gastric or intestinal cancer. t-PA may be a character of well differentiated tissue. Quantitative detection of TF and u-PA gene expression by the method of real-time RT-PCR is feasible for them as observation markers in gastric or intestinal cancer patients.

Topics: Adult; Blood Coagulation; Female; Humans; Intestinal Neoplasms; Male; Middle Aged; Neoplasm Invasiveness; Neoplasm Metastasis; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Stomach Neoplasms; Thromboplastin; Tissue Plasminogen Activator; Urokinase-Type Plasminogen Activator

Topics: Blood Coagulation Factors; Hemostasis; Humans; Neoplasm Metastasis; Neoplasms; Thromboplastin

To evaluate variations in the plasma tissue factor (TF) and urokinase type plasminogen activator (u-PA) system and their relationship with clinical cancer type, pathological classification and metastatic status in cancer patients.. Plasma levels of TF and its inhibitor (TFPI), as well as u-PA and its receptor (u-PAR) were measured using ELISA in 76 patients with malignant tumors and 24 patients with benign tumors.. Plasma levels of TF and u-PAR in the malignant tumor group were significantly higher than those of the benign tumor group and the normal control. U-PA and u-PAR increased significantly in patients with esophageal and gastric cancer. However, most of these parameters except TFPI did not vary according to pathological classification. A significant elevation was evident in patients with local infiltration, lymph node involvement and distal metastasis, while u-PAR only increased in the latter two categories.. Both the TF and u-PA systems are activated in cancer patients. U-PA and its receptor might prove to be a clinically useful marker for disease progression.

Topics: Adult; Aged; Aged, 80 and over; Female; Humans; Lipoproteins; Male; Middle Aged; Neoplasm Metastasis; Neoplasms; Receptors, Cell Surface; Receptors, Urokinase Plasminogen Activator; Thromboplastin; Urokinase-Type Plasminogen Activator

Tissue factor, the cellular initiator of blood coagulation, has been implicated as a determinant of metastatic potential in human melanoma cells. Here, we report that differential expression of tissue factor in murine melanoma cell lines of known metastatic behavior is mediated by AP-1-dependent and 12S E1A oncoprotein-repressible gene transcription. When compared to weakly metastatic C10 cells, highly metastatic M4 cells possessed elevated levels of tissue factor cofactor activity, transfected promoter activity, and heterodimeric AP-1 DNA-binding complexes containing Fra-1. Transient co-expression of the adenovirus E1A 12S oncoprotein strongly repressed transcription of an AP-1-driven tissue factor reporter gene indicating the additional requirement of N-terminal E1A-interacting coactivators. Stable expression of E1A mutants defective in CBP/p300-binding failed to suppress tissue factor expression and experimental metastasis by M4 cells while clones expressing wild type E1A exhibited greatly reduced tissue factor cofactor activity and metastatic potential in vivo. Overexpression of functional tissue factor in cells containing wild type E1A failed to restore the highly metastatic M4 phenotype suggesting that additional E1A-responsive and CBP/p300-dependent genes are required to facilitate metastasis of murine melanoma cells demonstrating high tissue factor expression and cofactor activity.

Topics: Adenovirus E1A Proteins; Animals; E1A-Associated p300 Protein; Gene Expression Regulation, Neoplastic; Hematologic Neoplasms; Lung Neoplasms; Melanoma; Mice; Mice, SCID; Models, Theoretical; Neoplasm Metastasis; Nuclear Proteins; Promoter Regions, Genetic; Protein Structure, Tertiary; Proto-Oncogene Proteins c-fos; Thromboplastin; Trans-Activators; Transcription Factor AP-1; Transcription, Genetic; Tumor Cells, Cultured

Infection of a human melanoma cell line by a retroviral vector resulted in transmission of a mouse VL30 (mVL30-1) retroelement RNA to some of the cells infected by the retrovirus, followed by synthesis, integration, and expression of the mVL30-1 cDNA. One vector carried a tissue factor (TF) transgene that generated high TF melanoma clones, and another vector was a control without the TF transgene that generated low TF clones. Some high TF melanoma clones contained the mVL30-1 retroelement and others did not, and some low TF melanoma clones contained the mVL30-1 retroelement and others did not. Each type of melanoma clone was tested for its metastatic potential in severe combined immunodeficient (SCID) mice, by i.v. injection of the cells to generate lung tumors. None of the low TF clones that either contained or lacked the mVL30-1 retroelement generated lung tumors, consistent with earlier results showing that high TF expression promoted metastasis. The high TF clones containing the mVL30-1 retroelement were strongly metastatic, in contrast to the high TF clones lacking the mVL30-1 retroelement, which were weakly metastatic. Southern hybridization analyses showed that the mVL30-1 cDNA integrated into different genomic sites in different melanoma clones, suggesting that the effect of the mVL30-1 retroelement on metastasis depends not on integration per se but instead on expression of the mVL30-1 RNA. A role for the mVL30-1 RNA in metastasis and possibly other cell functions is an unexpected finding, because the RNA appears to lack significant coding potential for a functional protein. The metastatic effect might be mediated directly by a noncoding mVL30-1 RNA or by a peptide or small protein encoded by one of the short ORFs in the mVL30-1 RNA.

Topics: Animals; Base Sequence; Blotting, Southern; DNA, Complementary; Humans; Melanoma; Mice; Mice, SCID; Molecular Sequence Data; Neoplasm Metastasis; Polymerase Chain Reaction; Retroviridae; Reverse Transcriptase Polymerase Chain Reaction; RNA; Thromboplastin; Transfection; Transgenes; Tumor Cells, Cultured

Cerivastatin is used in the treatment of hypercholesterolemia to inhibit 3-hydroxy 3-methylglutaryl coenzyme A reductase and thus prevent the synthesis of cholesterol precursors, such as farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP), responsible, respectively, for translocation of Ras and Rho to the cell membrane, a step required for their cell signaling, leading to cell proliferation and migration. Recently, it has been suggested that non lipid-related effects of statins could play a beneficial role in cancer therapy. In this study, we have investigated the mechanisms by which statins inhibit cancer and the types of cancers which could benefit from this therapy. In MDA-MB-231 cells, an aggressive breast cancer cell line with spontaneous activation of Ras and NFkappaB and overexpression of RhoA, cerivastatin induced inhibition of both cell proliferation and invasion through Matrigel. This anti-proliferative effect was related to G(1)/S arrest due to an increase in p21(Waf1/Cip1). The anti-invasive effect was observed from 18 h and could be explained by RhoA delocalization from the cell membrane, resulting in disorganization of the actin fibers and disappearance of focal adhesion sites. The importance of RhoA inactivation in both these inhibitory effects was proved by their reversion by GGPP but not by FPP. Moreover, cerivastatin was also shown to induce inactivation of NFkappaB, in a RhoA inhibition-dependent manner, resulting in a decrease in urokinase and metalloproteinase-9 expression, two proteases involved in cell migration. The participation of Ras inactivation is considered a subsidiary mechanism for the effects of cerivastatin, as they were not rescued by FPP. Prolonged treatment of MDA-MB-231 cells with high doses of cerivastatin induced a loss of cell attachment. Interestingly, the effect of cerivastatin was considerably lower on poorly invasive MCF-7 cells. In conclusion, our results suggest that cerivastatin inhibits cell signaling pathways involved in the invasiveness and metastatic properties of highly invasive cancers.

Topics: Apoptosis; Base Sequence; Breast Neoplasms; Cell Cycle; Cell Division; Cyclin-Dependent Kinase Inhibitor p21; Cyclins; DNA Primers; Gene Expression Regulation, Neoplastic; Humans; Hydroxymethylglutaryl-CoA Reductase Inhibitors; I-kappa B Proteins; In Vitro Techniques; Matrix Metalloproteinase 9; Neoplasm Invasiveness; Neoplasm Metastasis; NF-kappa B; Pyridines; RNA, Messenger; Signal Transduction; Thromboplastin; Tumor Cells, Cultured; Urokinase-Type Plasminogen Activator

Tissue factor (TF) is a transmembrane glycoprotein that complexes with factor VIIa to initiate blood coagulation. We previously reported that expression of high levels of TF in a human melanoma cell line promotes metastasis. Both the cytoplasmic domain of TF and its extracellular domain complexed with factor VIIa are required for the metastatic effect. To further explore the mechanism of TF-mediated metastasis, we investigated the possibility that a protease-activated receptor (PAR) might play a role. For this purpose, we first determined the expression levels of the known PARs (PAR1-4) in a human melanoma cell line, SIT1, that has low endogenous levels of TF and low metastatic potential. We found negligible levels of all of the known PARs and transfection of this cell line with human TF cDNA did not alter expression of the known PARs. To study the possible role of PAR1 in TF-mediated metastasis, we prepared a panel of transfected cell lines with varying levels of TF and PAR1. Our studies show that TF promotes metastasis by a pathway that does not involve high expression of known PARs by tumor cells. In addition, while overexpression of PAR1 is insufficient to induce metastasis in cells with low TF expression, it enhances the metastatic potential of cells with high TF expression, indicating a possible synergy between TF and PAR1 in promoting metastasis.

Topics: Animals; Disease Models, Animal; Drug Synergism; Female; Humans; Lung Neoplasms; Melanoma; Mice; Mice, SCID; Neoplasm Metastasis; Receptor, PAR-1; Receptors, Thrombin; Thromboplastin; Transfection; Tumor Cells, Cultured

TF expression is a hallmark of cancer progression. The procoagulant functions of TF that lead to thrombin generation are critically important to support metastasis, in part through the generation of fibrin that assures prolonged arrest of tumor cells in target organs. In addition, the coagulation initiation complex, i.e. TF-VIIa-Xa, generates autocrine cell signaling though protease activated receptors. A cooperation of the TF cytoplasmic domain with protease signaling may explain the diverse contributions of TF to metastasis and angiogenesis.

Topics: Blood Coagulation; Cysteine Endopeptidases; Factor X; Humans; Neoplasm Metastasis; Neoplasm Proteins; Neoplasms; Receptor, PAR-1; Receptor, PAR-2; Receptors, Thrombin; Thromboplastin; Up-Regulation

Tissue factor (TF) is an initiator of the extrinsic cascade of blood coagulation. Although recent studies have revealed a relationship between metastatic properties and TF expression in some neoplastic cells, the significance of TF in lung cancer, especially in non-small-cell lung cancer (NSCLC), is still unclear. In this study, TF was detected in NSCLC cell lines by functional study, Western blot analysis and immunocytochemical staining. TF levels in eight NSCLC cell lines were also quantitated by enzyme-linked immunosorbent assay (ELISA), and TF expression was evaluated in 55 specimens of surgically resected NSCLCs. NSCLC cell lines derived from metastatic lesions produced high levels of TF (48.3+/-23.5 ng 10(-6) cells, mean +/- s.e.m.), whereas those derived from primary lesions produced low levels of TF (0.2+/-0.1 ng 10(-6) cells). Immunohistochemical studies disclosed significantly stronger staining for TF in cells from NSCLC patients with metastasis than in those without metastasis. Among the 28 patients with metastasis, ten were strongly positive, 16 were moderately positive and two were negative for TF. In contrast, among the 27 patients without metastasis, only two were strongly positive, 18 were moderately positive and seven were negative for TF. Therefore, malignant cells from patients with lung cancer produce various levels of TF, and TF may play an important role in the metastatic process.

Topics: Biomarkers, Tumor; Blotting, Western; Carcinoma, Non-Small-Cell Lung; Enzyme-Linked Immunosorbent Assay; Humans; Immunohistochemistry; Lung Neoplasms; Neoplasm Metastasis; Thromboplastin; Tumor Cells, Cultured

Tissue factor (TF) is a transmembrane glycoprotein that complexes with factor VIIa to initiate blood coagulation. It was reported in an earlier study that expression of high levels of TF in a human melanoma cell line promotes metastasis, and that the cytoplasmic domain of TF is required for this metastatic effect. To analyze the functions of the cytoplasmic and extracellular domains of TF in metastasis, two TF mutants were constructed; in one mutant alanine was substituted for each of the three serine residues in the cytoplasmic domain, preventing phosphorylation; in the other mutant alanine was substituted for four key residues in the extracellular domain, preventing binding of factor VIIa and consequently eliminating the initiation of blood coagulation by the TF-VIIa complex. Melanoma lines expressing high levels of either mutant form of TF were weakly metastatic in SCID mice, indicating that phosphorylation of the cytoplasmic domain and formation of a complex with VIIa by the extracellular domain are required for the full metastatic effect of TF. It was also found that increasing TF expression in human melanoma cells does not increase expression of vascular endothelial growth factor or promote growth and vascularization of tumors derived from the melanoma cells, suggesting that TF acts by a mechanism other than angiogenesis to promote metastasis.

Topics: Animals; Gene Expression Regulation, Neoplastic; Humans; Melanoma; Mice; Mice, SCID; Mutation; Neoplasm Metastasis; Rabbits; Thromboplastin; Tumor Cells, Cultured

Tissue factor (TF), the initiating cell surface receptor of the coagulation cascade, plays important roles in embryogenesis, angiogenesis, and tumor cell metastasis. It is controversial whether proteolytic function of TF complexed with its serine protease ligand VIIa is required for metastatic tumor dissemination. We show here in a model for TF-dependent experimental hematogenous metastasis, that TF supports metastasis by both proteolytic activity of the TF-VIIa complex and currently undefined functions of the cytoplasmic domain. We demonstrate that ligand binding of VIIa to TF is required for metastasis. Antimetastatic properties of covalently inactivated VIIa provide evidence that ligand binding is insufficient per se to support metastasis, emphasizing that proteolytic activity is necessary for the metastatic process. Ala or Asp mutations of cytoplasmic serine residues were introduced to preclude or mimic phosphorylation. In vivo analysis of these mutants suggests that local protease generation on the tumor cell surface does not serve simply to activate the cytoplasmic domain of TF by serine phosphorylation. Thus, extracellular functions of the catalytically active TF-VIIa complex cooperate with specific functions of the TF cytoplasmic domain to support the complex process of hematogenous tumor cell dissemination.

Topics: Animals; CHO Cells; Cricetinae; Cytoplasm; Factor VIIa; Female; Lung Neoplasms; Mice; Mice, SCID; Neoplasm Metastasis; Phosphorylation; Structure-Activity Relationship; Thromboplastin; Transfection

Tissue factor (TF) is a cell-surface glycoprotein responsible for initiating the extrinsic pathway of coagulation. The overexpression of TF in human malignancy has been correlated with the angiogenic phenotype, poor prognosis, and thromboembolic complications. The mechanisms underlying constitutive expression of TF in cancer cells are poorly defined. We cloned TF cDNA on the basis of its strong expression in metastatic MDA-MB-231 breast carcinoma cells in contrast to its weak expression in non-metastatic MCF-7 cells. Transient transfection analysis showed that TF promoter activity in MCF-7 cells could be stimulated by expression of a membrane-targeted raf kinase (raf-CAAX). raf-induced activity was dependent on the presence of an AP-1/NF-kappaB motif in the TF promoter and was inhibited by dominant-negative mutants of jun and by I-kappaB alpha. MDA-MB-231 cells were found to contain higher levels of ERK1/2 kinase activity than did MCF-7 cells. Electrophoretic mobility shift assays showed that MDA-MB-231 nuclear proteins bound strongly to an oligonucleotide corresponding to the AP-1/NF-kappaB sequence, whereas MCF-7 nuclear extracts showed weak binding to this element. Finally, we showed that TF mRNA levels in MDA-MB-231 cells declined after addition of the mitogen-activated protein kinase kinase inhibitor PD98059. Our data showed that activation of the raf-ERK pathway led to activation of TF expression in breast carcinoma cells and suggested that constitutive activation of this pathway leads to high TF expression in MDA-MB-231 cells.

Topics: Base Sequence; Benzoquinones; Breast Neoplasms; Calcium-Calmodulin-Dependent Protein Kinases; Dactinomycin; DNA, Complementary; Enzyme Activation; Enzyme Induction; Enzyme Inhibitors; Female; Flavonoids; Gene Expression Regulation, Neoplastic; Genistein; Humans; Hydroquinones; Lactams, Macrocyclic; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinases; Molecular Sequence Data; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasm Proteins; Neovascularization, Pathologic; NF-kappa B; Okadaic Acid; Phenols; Proto-Oncogene Proteins c-raf; Quinones; Rifabutin; Signal Transduction; Tetradecanoylphorbol Acetate; Thromboplastin; Transcription Factor AP-1; Transcription, Genetic; Tretinoin; Tumor Cells, Cultured

Tissue factor (TF), the membrane glycoprotein that initiates blood coagulation, is constitutively expressed by many tumor cells and is implicated in peri-tumor fibrin deposition and hypercoagulability in cancer. Upregulation of tumor TF correlates with enhanced metastatic potential. Furthermore, TF has been colocalized with VEGF in breast cancer, specially at sites of early angiogenesis. There are no data on the effect of hypoxia on tumor cell TF expression. Since hypoxia is known to stimulate VEGF production, we studied whether this also induces tumor cell TF expression. Confluent monolayers of A375 melanoma, MCF-7 breast carcinoma and A549 lung carcinoma were cultured in either 95% air, 5% CO2 (normoxic) or 95% N2, 5% CO2 (hypoxic; 25-30 mmHg) for 24 h. Procoagulant activity (PCA) was measured by amidolytic and clotting assays, surface TF antigen by flow cytometry, early apoptosis by annexin V binding and VEGF levels in culture supernatants by ELISA. Hypoxia significantly increased tumor cell PCA in all three cell lines tested and TF antigen on A375 cells was increased four-fold (P <0.05). Pentoxifylline (PTX), a methylxanthine derivative, significantly inhibited the hypoxia-induced increase in PCA as well as VEGF release in all three cell lines tested. In A375 cells, PTX significantly inhibited TF antigen expression by both normoxic and hypoxic cells. Hypoxia induced a slight (5%) but not significant, increase in early apoptosis. Intravenous injection of hypoxic A375 cells into nude rats produced more pronounced thrombocytopenia (n = 5, P <0.01) and more lung metastases (n = 3, P <0.05) compared to normoxic cells. We conclude that hypoxia increases TF expression by malignant cells which enhances tumor cell-platelet binding and hematogenous metastasis. Hypoxia-induced upregulation of TF appears to parallel that of VEGF, although the mechanism remains unclear.

Topics: Animals; Cell Hypoxia; Endothelial Growth Factors; Free Radical Scavengers; Humans; Lymphokines; Neoplasm Metastasis; Neoplasms, Experimental; Pentoxifylline; Rats; Thromboplastin; Tumor Cells, Cultured; Up-Regulation; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors

Several studies have previously demonstrated tissue factor (TF) expression in solid tumors. In our study, we evaluated by immunohistochemical staining TF expression in 79 cases of colorectal cancer and 17 cases of metastatic cancer of the liver from colorectal cancer, and investigated the relationship between the clinicopathological features and TF expression. TF was detected in the tumor of 57% of colorectal cancer patients, and its expression was significantly increased (p=0.01) in metastatic tumors (88%). TF expression was more commonly observed in metastatic tumors than in any Dukes' stage of primary cancer. In primary cancer, the detection of TF was more frequent in cases with lymph node metastasis (Dukes' C, 63%) or with hematogenous metastasis (Dukes' D, 82%) than in tumors without lymph node or hematogenous metastasis (Dukes' A and B, 46%, p=0.03). These results suggest that the expression of TF is related with the metastatic potential of colorectal cancer.

Topics: Adenocarcinoma; Adult; Aged; Biomarkers, Tumor; Cell Differentiation; Colorectal Neoplasms; Disease Progression; Female; Humans; Liver Neoplasms; Lymphatic Metastasis; Male; Middle Aged; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasm Proteins; Neoplastic Cells, Circulating; Prognosis; Thromboplastin

Topics: Animals; Antineoplastic Agents; Blood Coagulation; Clinical Trials as Topic; Endothelium, Vascular; Humans; Neoplasm Metastasis; Neoplasms; Neovascularization, Pathologic; Thromboplastin

Procoagulant activity of pairs of cell lines, which were derived from the same original cell type but which possess different growth characteristics and metastatic properties, was examined. The following characteristics were considered suggestive of a greater likelihood of metastatic potential: high histological grade; establishment of the line from a metastatic rather than a nonmetastatic cancer; increased tumorigenicity in nude mice; and/or estrogen receptor-negative mammary cancer. Procoagulant activity was evaluated by a two stage clotting assay. Procoagulant activity was highly variable, with up to a 1,300-fold difference, among the cancer cell lines examined. The rate of clot formation was factor VII dependent and was totally inhibited by an anti tissue factor monoclonal antibody, indicating that tissue factor was the only significant procoagulant present in these cancer cells. Tissue factor antigen expression, evaluated by ELISA, correlated with procoagulant activity. In all pairs of cancer cell lines, those with characteristics of increased proliferative potential had increased tissue factor levels compared to cell lines that originated from the same cell type, but which possess less aggressive characteristics. Tissue factor activity in these cancer cells was increased by cell lysis or by exposure of intact cells to a calcium ionophore, similar to results previously obtained in fibroblasts. Tissue factor mRNA was evaluated by northern blot analysis using a specific probe complementary to tissue factor mRNA. The previously described predominant tissue factor mRNA species of 2.2 kb was identified in the majority of cancer cell lines examined, but tissue factor mRNA species of 3.2 to 3.4 kb were also identified.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenocarcinoma; Blood Coagulation; Blotting, Northern; Breast Neoplasms; Carcinoma, Renal Cell; Carcinoma, Transitional Cell; Cell Line; Gastrointestinal Neoplasms; Gene Expression; Humans; Kidney Neoplasms; Neoplasm Metastasis; RNA, Messenger; Thromboplastin; Tumor Cells, Cultured

The procoagulant activity observed in many types of tissue and cultured cells is due to tissue factor, a 30 kd transmembrane protein. The mRNA for tissue factor is a 2.2-kb species, which in some non-cancer cells can be up-regulated or induced by cytokines or by serum stimulation. In this study, induction of procoagulant activity in cancer cells was evaluated using the breast cancer cell line, MCF-7, and an adriamycin resistant subline, AdrRMCF-7, which has increased tumorigenicity in nude mice compared to the parental cell line. Procoagulant activity was factor VIIa dependent and was inhibited by an anti-tissue factor antibody. MCF-7 cells had minimal tissue factor activity, while AdrRMCF-7 cells had an 10-fold increase compared to the parental line. This increase was not observed in MCF-7 cells transfected with the multi-drug resistant gene, which is associated with adriamycin resistance. Serum stimulation of quiescent MCF-7 cells increased tissue factor activity 5-fold over baseline level, but did not increase activity in cells grown in serum-replete medium. Tissue factor activity of AdrRMCF-7 quiescent cells and AdrMCF-7 cells grown in serum-replete medium was enhanced 2-fold by serum stimulation. The predominant tissue factor mRNA species in MCF-7 cells was a 3.2 to 3.4-kb band, which increased in response to serum stimulation of cells grown in serum-replete medium. The mature 2.2-kb tissue factor mRNA band was detected in quiescent MCF-7 cells within six hours of serum stimulation and remained present 24 hours after stimulation. Synthesis of the 2.2-kb tissue factor mRNA species in MCF-7 and AdrRMCF-7 cells correlated with appearance of procoagulant activity. Thus, while procoagulant activity correlates with the level of the 2.2-kb tissue factor mRNA species in these cancer cells, there are inherent differences in tissue factor activity, antigen, and mRNA levels, as well as in regulation of its synthesis between these cells.

Topics: Animals; Antigens, Neoplasm; Blood Coagulation Factors; Blood Physiological Phenomena; Breast Neoplasms; Female; Humans; Mice; Mice, Nude; Neoplasm Metastasis; Neoplasm Transplantation; RNA, Messenger; Thromboplastin; Tumor Cells, Cultured

Metastasis is a multistep process which requires highly adapted interactions of tumor cells with host target organs. Compared with nonmetastatic cells, metastatic human melanoma cells express 1000-fold higher level of tissue factor (TF), the major cellular initiator of the plasma coagulation protease cascades. To explore whether TF may contribute to metastatic tumor dissemination, we analyzed the effect of specific inhibition of TF function on human melanoma metastasis in severe combined immunodeficient (SCID) mice. Using species-specific antibodies to TF, we demonstrate that initial adherence in insufficient for successful tumor cell implantation in a target organ. Rapid arrest of human tumor cells in the lungs of mice was not diminished by inhibition of TF. However, inhibition of TF receptor function and consequent reduction in local protease generation abolished prolonged adherence of tumor cells, resulting in significantly reduced numbers of tumor cells retained in the vasculature of the lungs. The growth of pulmonary metastases was also significantly inhibited by a blocking anti-TF monoclonal antibody and Fab fragments thereof, whereas a noninhibitory antibody lacked antimetastatic effects. Cell surface expression of functional TF thus contributes to melanoma progression by allowing metastatic cells to provide requisite signals for prolonged adhesive interactions and/or transmigration of tumor cells across the endothelium, resulting in successful metastatic tumor implantation.

Topics: Animals; Cell Adhesion; Female; Gene Expression; Humans; Lung Neoplasms; Melanoma; Mice; Mice, SCID; Neoplasm Metastasis; Neoplasm Transplantation; RNA, Messenger; RNA, Neoplasm; Thromboplastin

Topics: Aged; Endothelium, Vascular; Factor VII; Female; Heparin; Humans; Lipoproteins; Male; Middle Aged; Neoplasm Metastasis; Neoplasms; Thromboplastin

Activation of blood coagulation and local fibrin deposition may contribute to tumor metastasis. We have examined the ability of four human tumor cell lines (COLO 205, HepG2, J82 and CAPAN-2) to augment the conversion of prothrombin to thrombin by factor Xa and calcium in the presence and absence of exogenous factor Va. Using a chromogenic substrate assay to assess thrombin formation, we observed that all the above cell lines accelerated prothrombin activation in the absence of exogenous factor Va. The order of effectiveness was COLO 205 greater than HepG2 greater than J82 greater than CAPAN-2. In the absence of cells, no detectable thrombin formation occurred. Pretreatment of COLO 205 and HepG2 cells with anti-human factor V IgG inhibited prothrombin activation in a dose-dependent manner, but was without effect in J82 and CAPAN-2 incubation mixtures. Factor V coagulant activity was observed in COLO 205 and HepG2 cells as well as their culture media, but was not detected in J82 and CAPAN-2 cells or their culture media. Biosynthetic labeling and immunoprecipitation studies revealed that COLO 205 and HepG2 cells constitutively synthesized factor V or a factor-V-like molecule that comigrated with human factor V/Va on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. All four tumor cell lines exhibited saturation binding of exogenous human factor Va resulting in a dose-dependent enhancement of their ability to augment prothrombin activation. Our results indicate that these tumor cells can readily assemble a functional cell surface prothrombinase complex that may be important in fibrin deposition associated with the growth and metastatic progression of these, and perhaps, other tumors.

Topics: Enzyme Activation; Factor V; Factor Xa; Fibrin; Humans; Membrane Proteins; Neoplasm Metastasis; Neoplasm Proteins; Prothrombin; Thrombin; Thromboplastin; Tumor Cells, Cultured

No previous studies on the possible contribution of cancer-cell procoagulants to metastasis have fulfilled all the criteria for attaining biologically relevant and readily interpretable data (Grimstad et al., 1986), viz: (1) Spontaneous metastasis from primary tumors should be assessed in syngeneic animals; (2) cloned cell lines should be used to correlate cell properties, because heterogeneity within the cell lines employed is a source of serious error; (3) enough clones, derived from the same original tumor, should be used to identify only nonrandom correlations. Observing these criteria, we examined the procoagulant activities of 19 murine fibrosarcoma cell clones and 4 uncloned cell lines with high to moderate or low potential for lung metastases formation. The procoagulant activity found was exclusively of the thromboplastin (tissue factor, factor III) type. It occurred in all cell homogenates, but the quantities did not correlate with metastatic potential. In contrast, all highly to moderately metastatic cell clones and lines from 2 different fibrosarcomas shed thromboplastin activity into the culture medium, whereas no weakly metastatic cells did. Histological examination further supported these indications that release of thromboplastin from cancer cells can promote metastasis by initiating blood clotting and thereby facilitating arrest of the cancer cells in target organ vessels. Examination of a third fibrosarcoma showed that release of thromboplastin activity is not necessary for metastasis in all tumors.

Topics: Animals; Clone Cells; Female; Fibrosarcoma; Mice; Neoplasm Metastasis; Thromboplastin

Vitamin K deficiency, either dietary or pharmacologically induced by warfarin, was unable to affect the metastatic capacity of cells from a benzopyrene-induced fibrosarcoma in C57BL/6J mice. The same cells had a procoagulant activity, of tissue thromboplastin type, which was also completely unaffected by vitamin K antagonism or deficiency. In another murine model of spontaneous metastasis we previously suggested that depression of a particular procoagulant such as a direct factor X activator might contribute to the antimetastatic activity of warfarin. The failure of vitamin K deficiency to affect both the procoagulant and the metastatic capacity of the model reported here offers strong negative support to the same concept.

Topics: Animals; Fibrosarcoma; Male; Mice; Mice, Inbred C57BL; Neoplasm Metastasis; Thromboplastin; Vitamin K Deficiency; Warfarin

The mechanisms by which B16, 3LL and MH134 tumor cells induce platelet aggregation were studied. The B16 and 3LL tumors, which have high or moderate procoagulant activities, aggregated platelets only in the presence of Ca2+ and plasma factors. MH134, which had much lower procoagulant activity, aggregated platelets even in the absence of these factors. The induction of aggregation by B16 and 3LL could be prevented by thrombin inhibitors but not by the ADP scavenger, suggesting that thrombin, generated by procoagulant activities of tumor cells themselves, might play a major role in initiating aggregation. MH134-induced aggregation was not affected by any of the inhibitors, indicating that the mechanisms by which MH134 initiate platelet aggregation are independent of both thrombin and ADP.

Topics: Adenosine Diphosphate; Animals; Blood Coagulation; Calcium; Liver Neoplasms, Experimental; Lung Neoplasms; Male; Melanoma; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Neoplasm Metastasis; Platelet Aggregation; Thrombin; Thromboplastin

Topics: Animals; Anticoagulants; Blood Coagulation; Blood Coagulation Disorders; Blood Coagulation Factors; Cysteine Endopeptidases; Disseminated Intravascular Coagulation; Endopeptidases; Factor X; Factor Xa; Female; Fibrin Fibrinogen Degradation Products; Fibrinolysis; Hemorrhage; Humans; Male; Neoplasm Metastasis; Neoplasm Proteins; Neoplasms; Neoplasms, Experimental; Thromboembolism; Thromboplastin

Experiments were made to evaluate the potential role played by thrombogenic factors on the hematogenous arrest of circulating tumor cells in mice with demonstrable coagulopathies associated with the presence of a primary tumor, by administration of "therapeutic" doses of anticoagulants. The effects of warfarin, aspirin and heparin administration on the early arrest patterns of 125IdUrd-labelled TA3 carcinoma and Gardner lymphosarcoma cells injected intravenously into tumor-bearing mice were examined. Several hematologic parameters of carcinoma- and lymphosarcoma-bearing animals were measured prior to anticoagulation experiments and the results indicated that mice had coagulopathies similar to those found in cancer patients with disseminated intravascular coagulation syndrome, i.e., thrombocytopenia and elevated fibrinogen levels. Despite the presence of coagulation abnormalities and effective anticoagulation in recipient animals, all three agents were without effect on localization patterns of both tumor types. It was concluded that the proposed involvement of thrombogenesis in metastasis was probably not due to any role played by those clotting factors inhibited by aspirin, warfarin and heparin in early intravascular tumor cell arrest.

Topics: Animals; Anticoagulants; Aspirin; Blood Coagulation; Carcinoma; Heparin; Lymphoma, Non-Hodgkin; Mice; Neoplasm Metastasis; Neoplasm Transplantation; Neoplasms, Experimental; Neoplastic Cells, Circulating; Thromboplastin; Warfarin

Thromboplastic and fibrinolytic activities of rat ascites tumor cells avoiding any stromal elements were examined and the role of these activities in the blood-borne metastasis was discussed. Ten lines of tumor cells showed varied thromboplastic and fibrinolytic activities. Tumor cell lines examined were classified into four groups; (1) lines AH-130, AH-62, and AH-7974 with high thromboplastic and high fibrinolytic activities, (2) lines AH-130F(N), AH-66F, and AH-7974F with low thromboplastic and low fibrinolytic activities, (3) line SLC with high thromboplastic and low fibrinolytic activities, and (4) lines AH-109A, AH-41A, and AH-41C with moderate thromboplastic and low fibrinolytic activities. The cell lines AH-130 and AH-130F(N), as well as AH-7974 and AH-7974F, have the same origin and showed different enzymic activities. AH-130 caused more prominent thrombus formation in the pulmonary vessels of rats in the early stage of intravenous inoculation and induced more prominent decrease in the number of platelets and fibrinogen levels in peripheral blood than AH-130F(N). Also, AH-130 developed more abundant metastatic foci in the lung 72 hr and 7 days after intravenous inoculation than AH-130F(N).

Topics: Animals; Blood Coagulation; Cell Line; Fibrinolysis; Liver Neoplasms, Experimental; Lung Neoplasms; Neoplasm Metastasis; Neoplasms, Experimental; Plasminogen Activators; Pulmonary Artery; Pulmonary Embolism; Rats; Thromboplastin

Topics: Adult; Aged; Animals; Blood Coagulation Disorders; Carcinoma, Bronchogenic; Carcinoma, Ehrlich Tumor; Disseminated Intravascular Coagulation; Female; Fibrin Fibrinogen Degradation Products; Fibrinogen; Fibrinolysin; Fibrinolysis; Heparin; Humans; Lung Neoplasms; Male; Mice; Middle Aged; Neoplasm Metastasis; Neoplasms; Neoplastic Cells, Circulating; Thromboembolism; Thromboplastin

Topics: Blood Coagulation Tests; Female; Humans; Laparotomy; Neoplasm Metastasis; Ovarian Neoplasms; Thromboplastin

Topics: Blood Cell Count; Blood Coagulation; Blood Platelets; Blood Protein Disorders; Cholestasis; Diagnosis, Differential; Duodenal Neoplasms; Factor VII; Hemorrhage; Heparin; Humans; Liver Neoplasms; Neoplasm Metastasis; Pancreatic Neoplasms; Syndrome; Thromboplastin

Experiments were undertaken to test a new hypothesis for the mechanism underlying the Révész effect. The hypothesis proposes that lethally irradiated (LI) tumour cells enhance the take probability of a small number of transplanted viable (V) tumour cells mixed with them by exerting a thromboplastic effect at the site of injection; local fibrin formation prevents emigration of V cells from the site or secures their survival there. The evidence presented to support this hypothesis is as follows: in the case of 3 isogeneically transplanted tumours, admixed particulate brain extract simulated the effect of LI cells in increasing the take probability of V cells; brain extract simulated the effect of LI cells in greatly delaying the disappearance of (125)IUdR-labelled viable carcinoma cells from the injection site; V cells acquired a raised take probability by their incorporation in fibrin clots; it was confirmed that admixed erythrocytes increased the take probability of V cells; using a newly devised microscopical test for detection of the thromboplastic activity of individual cells, it was found that cell death was almost always required for the display of such activity; lymphocytes and bone marrow cells, ineffective in enhancing the take of V cells, were almost totally devoid of thromboplastic activity. Possible explanations are given for failure of a fibrinogen depleting agent, ancrod (Arvin) to inhibit the Révész effect when administered to recipients. It is concluded that the evidence strongly supports the hypothesis presented whilst seriously weakening the long-standing theories that admixed LI cells act by provision of nutrients or by local quenching of postulated immune reactivity.

Topics: Animals; Brain; Cell Line; Cell Survival; Fibrin; Leukemia, Experimental; Mice; Mice, Inbred Strains; Neoplasm Metastasis; Neoplasm Transplantation; Neoplasms; Neoplasms, Experimental; Peptide Hydrolases; Radiation Effects; Thromboplastin; Tissue Extracts; Venoms

Topics: Aged; Blood Coagulation Tests; Blood Platelet Disorders; Chronic Disease; Disseminated Intravascular Coagulation; Fibrin; Gastrointestinal Hemorrhage; Heparin; Humans; Male; Middle Aged; Neoplasm Metastasis; Prostatic Neoplasms; Prothrombin Time; Thromboplastin

Topics: Aged; Anticoagulants; Autopsy; Blood Cell Count; Blood Coagulation Disorders; Blood Platelets; Disseminated Intravascular Coagulation; Female; Humans; Kidney Glomerulus; Melanoma; Methods; Myocardium; Neoplasm Metastasis; Proteus; Prothrombin Time; Thromboplastin

Topics: Breast Neoplasms; Female; Humans; Lung Neoplasms; Middle Aged; Necrosis; Neoplasm Metastasis; Neoplasm Seeding; Neoplasms, Radiation-Induced; Radiotherapy; Skin Neoplasms; Thromboplastin