Lien entre l'hémostase et le développement néoplasique : rôle spécifique du facteur tissulaire et de l'inhibiteur du facteur tissulaire

For more then a century now, haemostatic disorders, also refered as coagulopathies, are recognized to be associated to neoplasia development. At first, these coagulopathies were often seen as the first clinical symptom of an occult malignancy and as such they were foreseen as merely consequence of cancer. However recent studies allowed to determine that a molecular link between haemostasis and cancer exists. Haemostatic protein were found to interact with the tumor environment or the tumor itself. In this thesis, we look at two important haemostatic proteins involved in anti- or pro-tumoral activities. These proteins we looked at were, tissue factor (TF), which as been identified as the main trigger of blood coagulation an its natural inhibitor which is named tissue factor pathway inhibitor (TFPI). In our investigations we focus on the non-haemostatic function of these proteins. First of all, we studied the effect of TFPI on angiogenesis. Angiogenesis, the formation of new blood vessels from pre-existing ones, is a critical component of cancer. In this first paper we demonstrate that, in addition to its anticoagulant properties, emerging data show that TFPI may also regulate endothelial cell functions via a non-haemostatic pathway. At physiological concentration, TFPI inhibits both endothelial cell migration and their differentiation into capillary-like structures in vitro. These effects were specific to endothelial cells since no inhibitory effect was observed on the migration of tumor (glioblastoma) cells. Inhibition of endothelial cell migration was correlated with a concomitant loss in cell adhesion, suggesting an alteration of focal adhesion complex integrity. Accordingly, we observed that TFPI inhibited the phosphorylation of focal adhesion kinase and paxillin, two key proteins involved in the scaffolding of these complexes and that this effect was specific to endothelial cells. These results suggest that TFPI influences the angiogenic process via a non-haemostatic pathway, by downregulating the migratory mechanisms of endothelial cells.   Met, the only known receptor for hepatocyte growth factor (HGF) is a member of the receptor tyrosine kinases (RTKs) family and recently this RTK emerged as one important contributor to human neoplasia. In physiologic and pathologic conditions, Met trigger various cellular functions related to cell proliferation and migration, inhibition of apoptosis and also regulate a genetic program leading to coagulation. Since medulloblastoma (MB) express high level of tissue factor (TF), the main initiator of blood coagulation, we therefore wanted to know if there was a link between Met and TF expression in these pediatric tumors. In this work we demonstrate that stimulation of MB cells in vitro with HGF lead to overt expression of TF. These data correlate with analysis of clinical MB tumors specimens, were tumors expressing high level of Met also express high level of TF. This HGF/Met/TF pathway seems specific to MB only since no effect was observed on glioblastoma cells. TF expression in MB leads to a pro-coagulation state which could initiate thrombosis and the formation of a provisional fibrin matrix. Furthermore, TF can also mediate non-haemostatic function such as cell migration, thus helping cancer cells to form distant metastasis.  Taken together these results suggest that HGF/Met pathway is responsible for TF expression in MB and explained how TF participate to tumor aggressiveness.   In the third part of this thesis we look at the effect of TF on cell survival as a huge body of evidence suggest a role for TF and its natural ligand FVIIa on cells death (apoptosis) protection. In this study, we show that HGF-induce anti-apoptotic effect on DAOY MB cells is correlated to TF expression. Using TF siRNA we completely reversed this anti-apoptosis activity. TF is responsible of p53 inactivation, thus allowing DAOY to resist to very high concentrations of the chemotherapeutic drug Etoposide. These finding highlights the importance of understanding the molecular biology of MB, to achieve a better clinical management and also find new therapeutic targets.   Overall, this thesis demonstrates that normal haemostatic protein such as TF and TFPI are also involved in tumor development. The different activities of TF and TFPI on cancer reflected the fact that every physiological or pathological system is dependent of a balance between activator and inhibitor. Furthermore the molecular link described in this thesis between haemostasis and cancer constitutes an argument against the fact that the haemostatic system is simply a consequence of cancer. Instead we show that TF is part of a strategy designed to improve tumor expansion while TFPI try to restrain the tumor growth by limiting tumor angiogenesis.

Keyphrases: Coagulopathies, TF, TFPI, Angiogenesis, TF, TFPI, Apoptosis, Coagulopathies