Dried plasma spot as an innovative approach to therapeutic drug monitoring of apixaban: Development and validation of a novel liquid chromatography-tandem mass spectrometry method

Abstract

Apixaban, a direct oral anticoagulant drug (DOAC), typically does not require routine therapeutic drug monitoring (TDM), yet recent guidelines propose its use in specific clinical scenarios. While various antifactor Xa (anti-FXa) chromogenic assays serve as useful proxies for measuring plasma exposure to apixaban in emergencies, they lack specificity compared with chromatographic methods. This research project is intended to the development and validation of a standardized protocol of liquid chromatography–tandem mass spectrometry (LC–MS/MS) in conformity with the ICH guidelines M10 for the measurement of apixaban in both plasma and dried plasma spots (DPSs). Samples preparation included protein precipitation after the addition of a deuterated internal standard (IS), and the chromatographic separation was carried out on a Thermo Scientific™ Accucore™ Polar Premium column (50 mm × 2.1 mm, i.d. 2.6 m). The newly developed LC–MS/MS method for apixaban mesurement from both plasma and DPS resulted linear over a wide concentration range (31.25–500 ng/mL), accurate, and reproducible without matrix effects, allowing for specific and rapid quantification. Stability was assessed on quality controls and a real sample, allowing the setting up of a robust TDM protocol that was applied to five anonymized plasma samples obtained from adult patients undergoing apixaban treatment at steady-state. In conclusion our novel LC–MS/MS method is adequate for accurate apixaban quantitation from both plasma and DPS matrixes, and may thus facilitate the guidelines suggested implementation of apixaban TDM, even in peripheral hospitals through shipment of DPS at reference laboratories.