A RECOMBINANT FACTOR VIIA ANALOGUE WITH ENHANCED POTENCY, EXPRESSED BY THROMBIN GENERATION, AND PHOSPHOLIPIDS DEPENDENCY: AN IN VITRO STUDY

Recombinant factor VIIa (rFVIIa) is a novel prohemostatic drug for patients with hemophilia who have developed inhibitory antibodies. The postulation has been made that hemophilia is not only a disorder of coagulation, but that hyperfibrinolysis due to a defective activation of thrombin activatable fibrinolysis inhibitor (TAFI) might also play a role. In this in vitro study, the potential of rFVIIa to down-regulate fibrinolysis via activation of TAFI was investigated. rFVIIa was able to prolong clot lysis time in plasmas from 17 patients with severe hemophilia A. The prolongation of clot lysis time by rFVIIa was completely abolished by addition of an inhibitor of activated TAFI. The concentration of rFVIIa required for half maximal prolongation of clot lysis time (Clys½-VIIa) varied widely between patients (median, 73.0 U/mL; range, 10.8-250 U/mL). The concentration of rFVIIa required for half maximal reduction of clotting time (Cclot½-VIIa) was approximately 10-fold lower than the Clys½-VIIa value (median, 8.4 U/mL; range, 1.7-22.5 U/mL). Inhibition of TFPI with a polyclonal antibody significantly decreased Clys½-VIIa values (median, 2.6 U/mL; range, 0-86.9 U/mL), whereas Cclot½-VIIa values did not change (median, 7.2 U/mL; range, 2.2-22.5 U/mL). On addition of 100 ng/mL recombinant full-length TFPI, a nonsignificant increase of Clys½-VIIa values was observed (median, 119.2 U/mL; range, 12.3-375.0 U/mL), whereas Cclot½-VIIa values did not change (median, 8.8 U/mL; range, 2.6-34.6 U/mL). In conclusion, this study shows that rFVIIa both accelerates clot formation and inhibits fibrinolysis by activation of TAFI in factor VIII-deficient plasma. However, a large variability in antifibrinolytic potential of rFVIIa exists between patients.

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Recombinant factor VIIa reverses the inhibitory effect of aspirin or aspirin plus clopidogrel on in vitro thrombin generation

Journal of Thrombosis and Haemostasis ◽

10.1111/j.1538-7836.2006.02088.x ◽

2006 ◽

Vol 4 (9) ◽

pp. 2022-2027 ◽

Cited By ~ 56

Author(s):

R. ALTMAN ◽

A. SCAZZIOTA ◽

M. DE LOURDES HERRERA ◽

C. GONZALEZ

Keyword(s):

Thrombin Generation ◽

Recombinant Factor Viia ◽

Factor Viia ◽

Inhibitory Effect

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Recombinant Factor VIIa Does not Induce Hypercoagulability In Vitro

Thrombosis and Haemostasis ◽

10.1055/s-0037-1614451 ◽

1999 ◽

Vol 81 (02) ◽

pp. 245-249 ◽

Cited By ~ 44

Author(s):

Gerhard Cvirn ◽

Wolfgang Muntean ◽

Siegfried Gallistl

Keyword(s):

Factor Viii ◽

Thrombin Generation ◽

Recombinant Factor Viia ◽

Factor Viia ◽

Factor Ix ◽

Control Factor ◽

Prothrombin Complex Concentrates ◽

Thrombotic Complications ◽

Activated Prothrombin Complex Concentrates

SummaryRecombinant factor VIIa (rVIIa) has been reported to be clinically effective and safe in haemophilic patients with inhibitor antibodies. Compared to activated prothrombin complex concentrates the risk of thrombotic complications seems to be very low after rVIIa administration. Determination of free thrombin generation has been shown to identify hypercoagulability. Therefore, free thrombin and prothrombinase activity (Xa generation) were assessed after extrinsic activation of rVIIa supplemented factor VIII and factor IX deficient plasma. Free thrombin generation was also determined after supplementation of (activated) prothrombin complex concentrates. Addition of 150 U rVIIa/ml shortened the clotting times markedly in control, factor VIII, and factor IX deficient plasma. In contrast, free thrombin and Xa generation were not different in the absence or presence of 150 U rVIIa/ml. Addition of (activated) prothrombin complex concentrates resulted in a marked increase of free thrombin generation in all investigated plasmas. Although in vitro studies cannot reflect specific clinical circumstances our results support the notion that rVIIa does not induce a hypercoagulable state as sporadically observed after administration of (activated) prothrombin complex concentrates.

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Monitoring low dose recombinant factor VIIa therapy in patients with severe factor XI deficiency undergoing surgery

Thrombosis and Haemostasis ◽

10.1160/th10-12-0816 ◽

2011 ◽

Vol 106 (09) ◽

pp. 521-527 ◽

Cited By ~ 29

Author(s):

Anne Riddell ◽

Rezan Abdul-Kadir ◽

Debra Pollard ◽

Edward Tuddenham ◽

Keith Gomez

Keyword(s):

Thrombin Generation ◽

Low Dose ◽

Recombinant Factor Viia ◽

Factor Viia ◽

Venous Blood ◽

Optimal Dose ◽

Factor Xi ◽

Factor Xi Deficiency

SummaryAlthough factor XI (FXI) concentrate is an effective replacement therapy in severe FXI deficiency without inhibitors, some patients are unwilling to receive it because it is plasma-derived. We report on the use and monitoring of low dose, recombinant factor VIIa (rFVIIa, NovoSeven®), to cover surgery (caesarean section, cholecystectomy and abdominoplasty) in four female patients (FXI:C 2–4 IU/dl, aged 32–51 years) who wished to avoid exposure to plasma. None of our patients had inhibitors to FXI. Our aim was to find the optimal dose of rFVIIa by in vitro spiking of patient samples and to correlate this with the response to rFVIIa in vivo. Prior to surgery, venous blood was collected into sodium citrate with corn trypsin inhibitor and spiked with 0.25–1.0 μg/ml rFVIIa in vitro, equivalent to a 15–70 μg/kg dose of rFVIIa in vivo. Analysis using thromboelastometry and thrombin generation assays, triggered with tissue factor, showed that the thrombin generation assay was insufficiently sensitive to the haemostatic defect in these patients. A concentration of 0.5 μg/ml was as effective as 1.0 μg/ml FVIIa in normalising thromboelastometry in vitro in all four patients. Therefore, patients received 15–30 μg/kg rFVIIa at 2–4 hourly intervals with tranexamic acid 1g every six hours. Post treatment samples were taken at 10–240 minutes and showed initial normalisation of thromboelastometry with gradual return to baseline after 2–4 hours. In conclusion, low-dose rFVIIa therapy was successfully used in four patients with severe FXI deficiency undergoing surgery to prevent bleeding and can be monitored using thromboelastometry.

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In vitro study of the anticoagulant effects of edoxaban and its effect on thrombin generation in comparison to fondaparinux

Thrombosis Research ◽

10.1016/j.thromres.2011.07.026 ◽

2012 ◽

Vol 129 (4) ◽

pp. e77-e82 ◽

Cited By ~ 49

Author(s):

Meyer Michel Samama ◽

Jeanne Mendell ◽

Céline Guinet ◽

Léna Le Flem ◽

Satoshi Kunitada

Keyword(s):

Thrombin Generation ◽

In Vitro Study ◽

Vitro Study

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Thrombin Generation by Fetoscopic Trauma to the Fetal Membranes: An in vivo and in vitro Study

Fetal Diagnosis and Therapy ◽

10.1159/000439304 ◽

2015 ◽

Vol 39 (4) ◽

pp. 261-268 ◽

Cited By ~ 2

Author(s):

Alexander C. Engels ◽

Dries Bauters ◽

Rita Rynkevic ◽

Savitree Pranpanus ◽

Jute Richter ◽

...

Keyword(s):

Amniotic Fluid ◽

Thrombin Generation ◽

In Vitro Study ◽

Vitro Study ◽

Thrombin Inhibitor ◽

Thrombin Receptor ◽

Fetal Membranes

Objective: We first aimed to investigate in vivo thrombin generation induced by fetoscopy, and second we used term membrane explants for measurement of thrombin generation, thrombin receptor location and induction of selected matrix metalloproteinases (MMPs) in tissue culture. Materials and Methods: In vivo study (37 cases): samples of amniotic fluid were taken at the beginning and end of fetoscopy (mean gestational age 26.7 weeks) and analyzed by ELISA for thrombin-antithrombin complexes. In vitro study: fetal membranes were put in culture and punctured for measurement of thrombin generation by calibrated automated thrombography and ELISA. Induction of MMP-9 and MMP-2 was analyzed by zymography. PAR-1 was localized by immunohistochemistry. Results: No significant increase in thrombin-antithrombin was measured in amniotic fluid obtained during fetoscopy. In vitro, thrombin generation induced by needle trauma of membrane cultures is correlated to the amount of plasma. Activity of MMP-9 but not MMP-2 was elevated in cultured membranes but could not be inhibited by a thrombin inhibitor. On histology, the thrombin receptor PAR-1 was located in the chorion and decidua, but not in the amnion. Discussion: Despite the influence of thrombin on punctured fetal membranes in vitro, the role of thrombin in iatrogenic preterm premature rupture of membranes is questionable.

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Recombinant Factor VIIa Partially Reverses the Effects of Anticoagulants and Antiplatelet Agents on Thrombin Generation and Thromboelastography.

Blood ◽

10.1182/blood.v106.11.4143.4143 ◽

2005 ◽

Vol 106 (11) ◽

pp. 4143-4143

Author(s):

Mariam L. Abdullatif ◽

Elizabeth Donnachie ◽

Miguel Escobar

Keyword(s):

Whole Blood ◽

Thrombin Generation ◽

Recombinant Factor Viia ◽

Factor Viia ◽

Antiplatelet Agents ◽

Adult Patients ◽

Bleeding Complications ◽

Time To Peak ◽

Patient Groups

Abstract Commonly used anticoagulants and anti-platelet agents have the potential drawback of bleeding complications and some lack an antidote for their quick reversal. Recombinant factor VIIa (rFVIIa) has recently been suggested as a general hemostatic agent but its use has only been approved for hemophiliacs with inhibitors. The purpose of this study was to evaluate the in-vitro ability of rFVIIa to reverse the effects of the following anticoagulant and/or anti-platelet agents: aspirin/plavix (n=12), heparin/aspirin/plavix (n=10), and coumadin (n=10). The criteria for patient selection were as follows: adult patients receiving coumadin; INR≥2, adult patients receiving heparin; aPTT≥ 50 seconds. After IRB approval and informed consent, blood was collected from patients receiving these medications and the effects of rFVIIa were studied using the thrombin generation assay and rotational thromboelastography (RoTEG®). Primary hemostasis in whole blood was assessed using the Multiplate® platelet analyzer following stimulation with collagen (final concentration was 3.2 μg/ml). Patients in the aspirin/plavix and heparin/aspirin/plavix groups, but not the coumadin group, had significantly decreased platelet activation compared to the control group. In platelet-poor plasma, the lagtime for thrombin generation was increased in all patient groups receiving anticoagulants and/or antiplatelet agents but the time-to-peak thrombin concentration was increased only in the heparin/ASA/plavix and coumadin goups. The addition of rFVIIa (2 and 6 μg/ml) shortened the lagtime in all patient groups so that it was not significantly different from controls. In the coumadin group, the time-to-peak thrombin concentration was reversed with rFVIIa at 2 μg/ml but was not reversed in the heparin/aspirin/plavix group even at 6 μg/ml. The endogenous thrombin potential (ETP) was decreased only in patients receiving coumadin and could not be reversed at the highest concentration of rFVIIa (6 μg/ml). In whole-blood thromboelastography, the clotting time (CT) in the heparin/aspirin/plavix and coumadin groups were increased compared to controls. Addition of rFVIIa (6 ug/ml) reversed this anticoagulant effect in the coumadin group but not the heparin/aspirin/plavix group. The mean clot firmness (MCF) was decreased only in the heparin/aspirin/plavix group and addition of rFVIIa (6 ug/ml) reversed this effect. Although the number of patients collected in the heparin/aspirin /plavix group was 10, only 4 could be used in the final analysis for thrombin generation and thromboelastography since those with aPTTs>85 seconds did not achieve a CT nor did they generate sufficient thrombin to be evaluated in the allotted time (30 minutes for thromboelastography and 90 minutes for thrombin generation) The in vitro addition of rFVIIa (2 and 6 μg/ml) did not reverse this effect. This study shows that rFVIIa accelerates thrombin generation and CT, but does not completely reverse the effects of these agents on thrombin generation and thromboelastography.

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