Factor VIIa bound to endothelial cell protein C receptor activates protease activated receptor-1 and mediates cell signaling and barrier protection.

Center for Biomedical Research, The University of Texas Health Science Center at Tyler, Tyler, TX, USA.
Blood (Impact Factor: 9.78). 01/2011; 117(11):3199-208. DOI: 10.1182/blood-2010-09-310706
Source: PubMed

ABSTRACT Recent studies have shown that factor VIIa (FVIIa) binds to the endothelial cell protein C receptor (EPCR), a cellular receptor for protein C and activated protein C, but the physiologic significance of this interaction is unclear. In the present study, we show that FVIIa, upon binding to EPCR on endothelial cells, activates endogenous protease activated receptor-1 (PAR1) and induces PAR1-mediated p44/42 mitogen-activated protein kinase (MAPK) activation. Pretreatment of endothelial cells with FVIIa protected against thrombin-induced barrier disruption. This FVIIa-induced, barrier-protective effect was EPCR dependent and did not involve PAR2. Pretreatment of confluent endothelial monolayers with FVIIa before thrombin reduced the development of thrombin-induced transcellular actin stress fibers, cellular contractions, and paracellular gap formation. FVIIa-induced p44/42 MAPK activation and the barrier-protective effect are mediated via Rac1 activation. Consistent with in vitro findings, in vivo studies using mice showed that administration of FVIIa before lipopolysaccharide (LPS) treatment attenuated LPS-induced vascular leakage in the lung and kidney. Overall, our present data provide evidence that FVIIa bound to EPCR on endothelial cells activates PAR1-mediated cell signaling and provides a barrier-protective effect. These findings are novel and of great clinical significance, because FVIIa is used clinically for the prevention of bleeding in hemophilia and other bleeding disorders.

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    ABSTRACT: Recombinant activated human Factor VII (rhFVIIa) is an established hemostatic agent in hemophilia but its mechanism of action remains unclear. Although tissue factor (TF) is its natural receptor, rhFVIIa also interacts with the endothelial protein C receptor (EPCR) through its γ-carboxyglutamic acid (Gla) domain with unknown hemostatic consequences in vivo. Here, we study whether EPCR facilitates rhFVIIa hemostasis in hemophilia using a mouse model system. Mouse activated FVII (mFVIIa) is functionally homologous to rhFVIIa, but binds poorly to mouse EPCR (mEPCR). We modified mFVIIa to gain mEPCR binding using 3 amino acid changes in its Gla-domain. The resulting molecule mFVIIa-FMR specifically bound mEPCR in vitro and in vivo and was identical to mFVIIa with respect to TF affinity and procoagulant functions. In macrovascular injury models in hemophilic mice, administered mFVIIa-FMR exhibited superior hemostatic activity compared to mFVIIa. This was abolished by blocking mEPCR and was absent in ex vivo whole blood coagulation assays, implicating a specific mFVIIa-FMR and endothelial mEPCR interaction. Since mFVIIa-FMR models the TF-dependent as well as EPCR binding properties of rhFVIIa, our data unmask a novel contribution of EPCR on the action of rhFVIIa administration in hemophilia, prompting the rational design of improved and safer rhFVIIa therapeutics.
    Blood 06/2014; · 9.78 Impact Factor
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    ABSTRACT: Background. Because of the very short half-life of factor VII, prophylaxis is considered a difficult endeavor in FVII deficiency. Design and Methods. Clinical efficacy and safety of prophylaxis regimens, and indications for their use, were evaluated in factor VII-deficient patients in the Seven Treatment Evaluation Registry. Results. Prophylaxis data (38 courses) were analyzed from 34 severe factor VII-deficient patients (<1-45 years, 21 female). Severest phenotypes (central nervous system, gastrointestinal, joint bleeding episodes) were highly prevalent. Twenty-one patients received recombinant activated factor VII (24 courses), four received plasma-derived factor VII, and ten fresh frozen plasma. Prophylaxis schedules clustered into 'frequent' (3 times weekly [n=23]) and 'infrequent' (≤2 times weekly [n=15]) courses. Excluding courses for menorrhagia, 'frequent' and 'infrequent' courses produced 18/23 (78%) and 5/12 (41%) 'excellent' outcomes, respectively; relative risk, 1.88; 95% confidence interval, 0.93-3.79; p=0.079. Long-term prophylaxis lasted from 1 to >10 years. No thrombosis or new inhibitors occurred. Conclusions. A subset of patients with factor VII deficiency needed prophylaxis because of severe bleeding. Recombinant activated factor VII schedules based on 'frequent' administrations (3 times weekly) and a 90 μg/kg total weekly dose were effective. These data provide a rationale for long-term, safe prophylaxis in factor VII deficiency ( NCT01269138).
    Haematologica 02/2013; · 5.94 Impact Factor
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    ABSTRACT: The protein C pathway provides multiple important functions to maintain a regulated balance between hemostasis and host defense systems in response to vascular and inflammatory injury. The anticoagulant protein C pathway is designed to regulate coagulation, maintain the fluidity of blood within the vasculature, and prevent thrombosis, whereas the cytoprotective protein C pathway prevents vascular damage and stress. The cytoprotective activities of activated protein C (APC) include anti-apoptotic activity, anti-inflammatory activity, beneficial alterations of gene expression profiles, and endothelial barrier stabilization. These cytoprotective activities of APC, which require the endothelial protein C receptor (EPCR) and protease-activated receptor-1 (PAR1), have been a major research focus. Recent insights, such as non-canonical activation of PAR1 at Arg46 by APC and biased PAR1 signaling, provided better understanding of the molecular mechanisms by which APC elicits cytoprotective signaling through cleavage of PAR1. The discovery and development of anticoagulant-selective and cytoprotective-selective APC mutants provided unique opportunities for preclinical research that has been and may continue to be translated to clinical research. New mechanisms for the regulation of EPCR functionality, such as modulation of EPCR-bound lipids that affect APC's cytoprotective activities, may provide new research directions to improve the efficacy of APC to convey its cytoprotective activities to cells. Moreover, emerging novel functions for EPCR expand the roles of EPCR beyond mediating protein C activation and APC-induced PAR1 cleavage. These discoveries increasingly develop our understanding of the protein C pathway, which will conceivably expand its physiological implications to many areas in the future.
    Journal of Thrombosis and Haemostasis 06/2013; 11(s1). · 6.08 Impact Factor


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May 22, 2014