Effect on Plasma Protein S Activity in Patients Receiving the Factor Xa Inhibitors

Abstract

Aims: Measurement of protein S (PS) activity in patients taking direct oral anticoagulants (DOACs) using reagents based on a clotting assay results in falsely high PS activity, thus masking inherited PS deficiency, which is most frequently seen in the Japanese population. In this study, we investigated the effect of factor Xa (FXa) inhibitors on PS activity using the reagent on the basis of the chromogenic assay, which was recently developed in Japan.

Methods: The study enrolled 152 patients (82 males and 70 females; the average age: 68.5±14.0 years) receiving three FXa inhibitors (rivaroxaban, edoxaban, and apixaban). PS activity was measured using the reagents on the basis of the clotting and chromogenic assays.

Results: PS activity measured by the clotting assay reagents exhibited falsely high values depending on the plasma concentrations of FXa inhibitors in patients taking either rivaroxaban or edoxaban. However, none of the three FXa inhibitors affected PS activity when measured using the chromogenic assay.

Conclusion: In patients taking rivaroxaban or edoxaban, inherited PS deficiency is likely missed because the levels of PS activity measured using the reagents based on the clotting assay are falsely high. However, we report that three FXa inhibitors do not affect PS activity measured by the chromogenic assay. When measuring the levels of PS activity in patients undergoing DOACs, the principles of each reagent should be understood. Furthermore, plasma samples must be collected at the time when plasma concentrations of DOACs are lowest or the DOAC-Stop reagent should be used.

Keywords: Chromogenic assay; Clotting assay; Factor Xa inhibitor; Inherited protein S deficiency; Protein S Tokushima.

Fig.1. Principle of total PS activity measurement using a chromogenic assay

PS in the plasma sample is reacted with APC and FVa for a certain period of time, and FVa is inactivated by PS (reaction 1). Residual FVa forms a prothrombinase complex with FXa and phospholipid and then activates prothrombin to thrombin. Since the generated thrombin degrades the chromogenic substrate, PS activity is indirectly detected by colorimetric quantification of its coloration (reaction 2).

Fig.2. Effect of rivaroxaban, edoxaban, and apixaban on PT and APTT

A, D: Rivaroxaban, B, E: Edoxaban, C, F: Apixaban

Fig.3. Effect of rivaroxaban, edoxaban, and apixaban on the levels of PS activity and PS antigen

The graphs indicated PS activity measured using Chromogenic assay (●) and Clotting assay (○), and PS antigen (▲). A, D: Rivaroxaban, B, E: Edoxaban, C, F: Apixaban.

Fig.4. Plasma concentration of rivaroxaban and the changes in PT, APTT, and PS activity before and after rivaroxaban introduction

The graphs indicated the concentration of rivaroxaban (A), the change of PT (B) and APTT (C), and the changes in PS activity values by clotting assays or chromogenic assay before and after rivaroxaban introduction (D). “Stago STA Staclot Protein S,” “Siemens Protein S Ac,” and “HemoclotTM Protein S” are the clotting assay reagents, and “Shino-Test PS total activity” is the reagent based on the chromogenic assay.

Fig.5. Effect of rivaroxaban, edoxaban, and apixaban on AT activity

A: Rivaroxaban, B: Edoxaban, C: Apixaban, □: Xa-based assay, ■: Thrombin-based assay