A-234 Protein S Activity Assay Reagent Unaffected by DOACs

Abstract

The use of direct oral anticoagulants (DOACs) for the prevention and treatment of venous thromboembolism (VTE) has been steadily increasing, establishing DOACs as a viable alternative to warfarin. However, it has been reported that DOACs can interfere with coagulation tests that rely on clotting assays. This interference is a significant challenge, as it may lead to false positives in tests assessing coagulation regulatory factors, including protein S (PS) activity. To accurately diagnose PS deficiency, a recognized risk factor for VTE, blood samples must either be drawn during periods of low DOAC levels or processed using a DOAC adsorbent. This requirement places an additional burden on both patients and medical technologists. Our reagent for testing PS activity, which utilizes a synthetic substrate method, is unaffected by DOACs and serves as an effective solution to this challenge. This reagent is based on the synthetic substrate method. In the initial reaction, activated protein C contained within the reagent degrades activated coagulation factor V (FVa) in the presence of PS as a cofactor within the sample. In the subsequent reaction, activated coagulation factor X converts prothrombin to thrombin using the residual FVa from the first reaction. The resultant thrombin then reacts with a chromogenic substrate in the reagent, leading to an increase in absorbance at 405 nm. Since the increase in absorbance is directly proportional to PS activity, this method facilitates the quantification of PS activity. The accuracy of this reagent was 0.74 IU/mL (96%) when compared to the international standard NIBSC 03/228 (0.77 IU/mL). The within-run reproducibility was 0.64 ± 0.020 IU/mL (CV 3.2%) for low PS samples and 1.17 ± 0.015 IU/mL (CV 1.3%) for normal PS samples. The between-run reproducibility was 0.67 ± 0.024 IU/mL (CV 3.6%) for low PS samples and 1.17 ± 0.015 IU/mL (CV 1.3%) for normal PS samples. Correlation analysis between this reagent (y) and a clotting-time reagent (x) yielded results of y = 0.984x + 0.097 (r = 0.94) when compared to clotting-time reagent A and y = 0.902x + 0.202 (r = 0.92) when compared to clotting-time reagent B. In addition, the effect of DOACs on this reagent was evaluated. The reagent was not affected by three DOACs (rivaroxaban, edoxaban, and apixaban). This reagent for testing PS activity effectively addresses clinical needs for patients undergoing treatment with DOACs, facilitating the ongoing evaluation of PS deficiency during anticoagulant therapy. By providing more reliable results, this reagent may enhance the rapid prevention, diagnosis, and treatment of VTE.