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Measurement of Factor V Activity in Human Plasma Using a Microplate Coagulation Assay

Measurement of Factor V Activity in Human Plasma Using a Microplate Coagulation Assay

Published on: September 9, 2012

NXT007-mediated hemostatic potential is suppressed by activated protein C-catalyzed inactivation of activated factor

Yuto Nakajima^1,2, Kenichi Ogiwara¹, Keito Inaba³

¹Department of Pediatrics, Nara Medical University, Kashihara, Nara, Japan.

Research and Practice in Thrombosis and Haemostasis

|December 20, 2023

Abstract

Background:

Activated protein C (APC) inactivates activated factor (F) V (FVa) and FVIIIa. NXT007, an emicizumab-based engineered therapeutic bispecific antibody, enhances the coagulation potential of FVIII-deficient plasma (FVIIIdef-plasma) to near normal levels. However, little is known about the effect of APC-induced inactivation in NXT007-mediated hemostatic function.

Objectives:

To investigate the contribution of APC-mediated reactions to NXT007-driven hemostasis.

Methods:

In pooled normal plasma (PNP) or FVIIIdef-plasma spiked with NXT007 (10 μg/mL), effects of APC (0-16 nM) were measured using a thrombin generation assay (TGA). The direct effects of APC on cofactor activity of NXT007 or FVIIIa in a FXa generation assay were also measured. The FVdef-plasma and FV Leiden plasma (FV_Leiden plasma) were preincubated with 2 anti-FVIII monoclonal antibodies (termed FVIII-depleted), and the APC effect in the presence of NXT007 in FVIII-depleted FVdef-plasma with the addition of exogenous FV (7.5-30 nM) or FVIII-depleted FV_Leiden plasma was investigated.

Results:

The APC dose-dependent suppression effect in TGA of FVIIIdef-plasma spiked with NXT007 was similar to that of PNP. FXa generation with NXT007 was not impaired by the addition of APC, suggesting that the APC-induced reaction in TGA with NXT007 was attributed to the direct inactivation of FVa. The addition of APC to FVIII-depleted FVdef-plasma, along with NXT007 and various FV concentrations, showed a similar attenuation to PNP. The NXT007-driven thrombin generation in FVIII-depleted FV_Leiden plasma was suppressed by APC, similar to the reaction in native FV_Leiden plasma.

Conclusion:

NXT007 did not impair APC-mediated downregulation of FVa in FVIIIdef-plasmas, regardless of the presence of FV mutation with APC resistance.

Keywords:

antibodies anticoagulants factor V factor VIII hemophilia A

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