Validation Studies on Coagulation Laboratory Assays Useful for Assessing the Plasma Levels of Apixaban
Authors: Hoppensteadt Debra , Goldford Marc , Bisset Anne , Syed Daneyal , Zuckerman Leon , Jeske Walter , Harenberg Job , Fareed Jawed 
Published: November 2019

Abstract
Apixaban(BMS/Pfizer), is clinically developed for DVT prophylaxis in patients undergoing hip and knee surgery, stroke prevention in patients with atrial fibrillation, and the prevention of acute ischemic events in patients with acute coronary syndrome. As claimed for other new oral anticoagulants (dabigatran, rivaroxaban), there appears to be no need to routinely monitor apixaban. However, there are clinical settings that would benefit from monitoring (e.g., patients who do not fit the enrollment criteria of those patients evaluated through the clinical trials). There are also clinical settings where assessing drug levels would be a critical component of patient care (e.g., patients admitted to the Emergency Department; stroke patients; patients on drug with failed anticoagulant protection or with bleeding; patients on drug who require interventional procedures or surgery, etc.). At this time a laboratory assay for apixaban has not yet been defined. Data from several laboratories demonstrates that the prothrombin time (PT) and activated partial thromboplastin time (aPTT) tests do not provide adequate sensitivity to apixaban at typical clinically obtained or higher concentrations. This was consistently found with multiple different manufacturers’ reagents for both the PT and aPTT. This collaborative study was designed to provide information on potential laboratory tests that maybe suitable for the clinical assessment of plasma levels of apixaban.
Material & Methods: Five coagulation laboratories were included in this collaborative study. Each laboratory received an apixaban standard at 1.0 ug/ml, normal human pooled plasma and unknown samples labeled A, B, C, D and E. Each day for 5 days, a standard 6 point curve in the range of 0-1.0 ug/ml and samples A-E were run in duplicate using the Technoclone anti-Xa assay (Technoclone, Vienna, Austria), Coamatic anti-Xa assay (Chromogenix; Instrumentation Laboratory; Bedford, MA) and the Prothrombin time using Innovin Siemens, Deerfield, IL). All date was sent to the Hemostasis & Thrombosis Research Laboratory, Loyola University Chicago for analysis.
Results: The measured apixaban levels as measured by various methods are shown in the following Table. TableSampleTechnochromCoamaticInnovin PT1000 ng/ml1051.50±48.2 ng/ml968.26±38.6 ng/ml362.06±138.5 ng/ml500 ng/ml525.1±32.5 ng/ml540.85±40.9 ng/ml944.09±245.4 ng/ml250 ng/ml239.3±28.8 ng/ml267.2±29.7 ng/ml468.5±21.60 ng/ml125 ng/ml140.4±24.6 ng/ml191.16±27.01 ng/ml678.55±147.8 ng/ml30 ng/ml25.5±8.9 ng/ml38.7±14.2 ng/ml155.19±93.3 ng/ml
The Technochrom anti-Xa and Coamatic anti-Xa assays were the most accurate in determining the concentration of apixaban in the plasma. There was a large standard deviation which was due to the variation in instrumentation between the laboratories. Each laboratory adapted the three assays to their current coagulation analyzers. The PT overestimated the apixaban concentration at <500 ng/ml and under estimated at higher concentration.
Conclusions: These studies suggest that the anti-Xa chromogenic methods from Technoclone and Chromogenix may be used to determine the concentration of apixaban in plasma. Each method needs to be validated on the specific coagulation instrument to be used to assess the apixaban concentration.

Disclosures
No relevant conflicts of interest to declare.