A Novel Specific Immunoassay for Plasma Two-Chain Factor VIIa: Investigation of FVIIa Levels in Normal Individuals and in Patients With Acute Coronary Syndromes

Philippou, Helen; Adami, Antonella; Amersey, Rajiv; Stubbs, P J; Lane, David A.

doi:10.1182/blood.v89.3.767

Cited by 43 publications

(29 citation statements)

References 20 publications

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“…The levels of FVII:C and FVII:Ag, as well as free TFPI, were within normal ranges as compared with the control group. This finding is in accordance with the results published by other groups (Morrissey et al, 1993;Philippou et al, 1997;Giannitsis et al, 2000). The present study showed for the first time that the treatment of unstable angina using a LMWH (enoxaparin) inhibits in vivo the generation and the activity of factor VIIa.…”

Section: Discussionsupporting

confidence: 94%

Inhibition of factor VIIa generation and prothrombin activation by treatment with enoxaparin in patients with unstable angina*

Gerotziafas

Zafiropoulos²,

Dreden³

et al. 2003

Br J Haematol

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Summary. Factor VIIa (FVIIa) and thrombin generation occur in patients suffering an acute coronary event. We studied the effect of treatment with enoxaparin on FVIIa and prothrombin activation in patients with unstable angina. Anti-Xa activity, FVIIa, FVII coagulant activity (FVII:C) and FVII antigen (FVII:Ag), free tissue factor pathway inhibitor (TFPI), and prothrombin fragments 1 + 2 (F 1+2 ) were measured in patients' plasma, over a 24-h treatment period with enoxaparin. All 14 patients recruited in the study (mean age 68 years) were treated with a subcutaneous injection of enoxaparin, 1 mg/kg twice daily. Blood was drawn just before, and at different time intervals after, the first injection. Before enoxaparin administration, the levels of FVIIa (4AE02 ± 0AE8 ng/ml) and F 1+2 (2AE68 ± 0AE2 nmol/l) were significantly increased as compared with control subjects (2AE3 ± 0AE3 ng/ml and 0AE9 ± 0AE1 nmol/l respectively, P < 0AE05). Free TFPI, FVII:C and FVII:Ag were within normal ranges. One hour after the first injection of enoxaparin, FVIIa and F 1+2 levels decreased by 65% and 50%, respectively, and no significant fluctuations were noted throughout the observation period. The concentrations of FVII:C and FVII:Ag were not modified as compared with baseline values. After each injection, the peak concentrations of free TFPI and anti-Xa activity were observed at 2 and 4 h respectively. The kinetics of FVIIa and F 1+2 inhibition did not follow those of anti-Xa activity and TFPI release.

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Section: Discussionsupporting

confidence: 94%

Inhibition of factor VIIa generation and prothrombin activation by treatment with enoxaparin in patients with unstable angina*

Gerotziafas

Zafiropoulos²,

Dreden³

et al. 2003

Br J Haematol

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“…However, caution should be used in these interpretations. First, the validity of the FVIIa used in this study has been questioned recently and it may be that the levels we obtained are overestimations due to a possible in vitro activation of factor VII (Philippou et al, 1997). Alternatively, it is possible that the antibody used by Philippou et al (1997) cannot recognize the FVIIa bound to lipid or tissue factor.…”

Section: Discussionmentioning

confidence: 93%

“…First, the validity of the FVIIa used in this study has been questioned recently and it may be that the levels we obtained are overestimations due to a possible in vitro activation of factor VII (Philippou et al, 1997). Alternatively, it is possible that the antibody used by Philippou et al (1997) cannot recognize the FVIIa bound to lipid or tissue factor. In addition, the FVIIa clotting assay which uses a soluble mutant of tissue factor may measure partially activated FVII forms which are not recognized by immunoassay.…”

Section: Discussionmentioning

confidence: 93%

“…Until recently, levels of FVIIa and FXIIa were estimated by indirect methods, but new assays have recently been developed. For FVIIa a clotting assay using a soluble mutant of tissue factor (Morrisey et al, 1993;Ford et al, 1996) and an ELISA are now available (Philippou et al, 1997). These assays, which enable direct measurement of FVIIa in plasma, are apparently free of interference from the FVII zymogen (Wildgoose et al, 1992;Morrisey et al, 1993;Eichinger et al, 1997;Philippou et al, 1997).…”

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confidence: 99%

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Longitudinal study on activated factors XII and VII levels during normal pregnancy

Amiral²,

Vissac³

et al. 1998

Br J Haematol

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Summary. Levels of activated factor XII (FXIIa) and VII (FVIIa) were determined in 100 women with uneventful pregnancies. Samples were divided into five study intervals: three during pregnancy, one at delivery and one 3 d postpartum.The median (range) for FXIIa levels were 3·4 ng/ml (1·2-9·1) from 11 to 20 weeks, 4·6 ng/ml (1·4-15·2) from 21 to 30 weeks, 5·4 ng/ml (1·9-14·3) from 31st week to delivery, 5·2 (1·3-11·4) at delivery and 4·3 (1·8-8·5) ng/ ml in the postpartum sample. For FVIIa the median and range levels for the five periods were 4·9 (1·7-77·3), 7·2 (2·5-80·4), 11·1 (2·9-90·6), 12·0 (3·1-64·1) and 8·2 (4·0-23·5) ng/ml. Although the increase of FVIIa was higher than that of FXIIa during pregnancy, the overall changes of FXIIa and FVIIa were highly correlated (P < 0·0001). At each time period the changes of FVIIa correlated with FVII:C which was not the case with FVII:Ag. These data indicate that during pregnancy both the contact phase and extrinsic pathway are activated.Keywords: activated factor VII, factor VII antigen, factor VII activity, activated factor XII, pregnancy.Activation of coagulation can be mediated by the extrinsic or intrinsic pathways. The first enzyme of the extrinsic pathway is factor VIIa (FVIIa), which is the enzymatically active form of the zymogen factor VII. Activated factor XII (FXIIa), on the other hand, is the central enzyme of the contact phase. Until recently, levels of FVIIa and FXIIa were estimated by indirect methods, but new assays have recently been developed. For FVIIa a clotting assay using a soluble mutant of tissue factor (Morrisey et al, 1993;Ford et al, 1996) and an ELISA are now available (Philippou et al, 1997). These assays, which enable direct measurement of FVIIa in plasma, are apparently free of interference from the FVII zymogen (Wildgoose et al, 1992; Morrisey et al, 1993; Eichinger et al, 1997;Philippou et al, 1997). For FXIIa an immunoassay is now also available (Boisclair et al, 1993); it measures FXIIa directly in plasma with minimal crossreaction with the zymogen factor XII.Due to the many changes observed in haemostatic parameters, pregnancy is a particular subject of interest in understanding the activation of coagulation in a physiological state. Two preliminary reports have indicated either an increase of FVIIa (Morrissey et al, 1993) or of FXIIa (Coppolla et al, 1996) in normal pregnancies. Since no longitudinal studies on a large group of normal pregnant women have been performed, the aim of our study was to extend these preliminary observations by measuring systematically during pregnancy, at delivery and in the postpartum both FVIIa and FXIIa, and by comparing their levels with those of FVII clotting activity, FVII antigen and fibrinogen levels. PATIENTS AND METHODSThe study protocol was approved by the Ethical Committee of the University of Geneva Hospital. 100 women with uneventful pregnancies were included in the final analysis; 21 women were not included due to gestational hypertension, pre-eclampsia, diabetes mellitus and intrauterine...

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“…Philippou et al generated a polyclonal antibody directed against the C-terminal part of the FVIIa light chain, which displayed >3000-fold greater reactivity to FVIIa than FVII. 19 Andersen et al 18 reported on antibodies that show preferential binding to FVIIa and stimulated FVIIa activity. Most recently, a llama-derived single-domain antibody (sdAb) that binds to FVIIa but not to FVII has been described.…”

Section: Introductionmentioning

confidence: 99%

A single‐domain antibody that blocks factor VIIa activity in the absence but not presence of tissue factor

Ferrière

Kawecki

Ottavi³

et al. 2019

Journal of Thrombosis and Haemostasis

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Background Activated factor VII (FVIIa) is pertinent to the initiation of blood coagulation. Proteolytic and amidolytic activity of FVIIa are greatly enhanced by its cofactor, tissue factor (TF). Objective We aimed to generate a single‐domain antibody (sdAb) that recognizes free FVIIa rather than TF‐bound FVIIa. Methods A llama‐derived phage library was used to screen for anti‐FVIIa sdAbs. Results One sdAb, KB‐FVIIa‐004, bound to FVIIa, but not to its precursor FVII or to homologous proteins (prothrombin, factor X, or their activated derivatives). FVIIa amidolytic activity was inhibited by KB‐FVIIa‐004 (Ki = 28‐45 nM) in a competitive manner. KB‐FVIIa‐004 also inhibited FVIIa‐mediated FX activation (Ki = 26 nM). In contrast, KB‐FVIIa‐004 was inefficient in prolonging the clotting time of the prothrombin time‐test, which was prolonged by a maximum of 10 s at high sdAb concentrations (10 μM). Furthermore, FVIIa/TF amidolytic activity or FVIIa/TF‐mediated FX activation remained unaffected up to a 50‐fold to 1000‐fold molar excess of KB‐FVIIa‐004. These data suggest that KB‐FVIIa‐004 loses its inhibitory activity in the presence of TF. A KB‐FVIIa‐004/albumin fusion‐protein (004‐HSA) was generated for in vivo testing. By using 004‐HSA, we observed that this sdAb blocked the therapeutic capacity of FVIIa to correct bleeding in FVIII‐deficient mice. Discussion This observation is compatible with the view that FVIIa functions independently of TF under these conditions. In conclusion, we have generated a sdAb that specifically blocks TF‐independent activity of FVIIa. This antibody can be used to gain insight into the roles of TF‐bound and TF‐free FVIIa.

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A Novel Specific Immunoassay for Plasma Two-Chain Factor VIIa: Investigation of FVIIa Levels in Normal Individuals and in Patients With Acute Coronary Syndromes

Cited by 43 publications

References 20 publications

Inhibition of factor VIIa generation and prothrombin activation by treatment with enoxaparin in patients with unstable angina*

Inhibition of factor VIIa generation and prothrombin activation by treatment with enoxaparin in patients with unstable angina*

Longitudinal study on activated factors XII and VII levels during normal pregnancy

A single‐domain antibody that blocks factor VIIa activity in the absence but not presence of tissue factor

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