Comparison of Laboratory Assays to Measure Rivaroxaban and Dabigatran
Authors: Robert Severine , Mullier François , Mekouar Hassan , Polet Florence , Douxfils Jonathan , Biot Françoise , Lambert Marie-Christine , Baudar Justine , Devel Philippe , Chatelain Christian J. , Chatelain Bernard , Dogné Jean-Michel 
Published: October 2019

Abstract
Abstract 3332

Introduction:
Recently, 2 oral anticoagulants have been marketed: dabigatran, a direct thrombin inhibitor, and rivaroxaban, a direct FXa inhibitor. Thanks to their safety and efficacy profiles, routine coagulation monitoring is generally not required. However, the most commonly reported adverse reactions with these drugs are bleedings. A sensitive laboratory assay might be valuable to measure the pharmacodynamics of these 2 drugs in different situations including risks of drug interaction (i.e. strong P-gp inhibitors), overdose or to measure patient compliance. Due to their different mechanism of action, the measurement of their anticoagulant level in plasma may require different types of clotting assays.

Aims:
The aim of the present study was to determine which coagulation assay(s) could be used to measure the impact of rivaroxaban and dabigatran among a range of routine or more specific tests.

Methods:
Both drugs were spiked at increasing concentrations in pooled citrated normal human platelet-poor plasma (PPP). The final concentrations were ranging from 10 nM to 2 μM for dabigatran and from 25 nM to 5 μM for rivaroxaban. Such concentrations covered the plasma range in patients during orthopedic surgery. The following routine clotting assays were performed with the spiked plasma according to the manufacturer's protocols: Prothrombin Time (PT) with Neoplastin CI+ and Neoplastin R on STA-R (Roche Diagnostics), with Innovin on BCS (Siemens) and with Recombiplastin on ACL-TOP (Instrumentation Laboratories); activated Partial Thromboplastin Time (aPTT) with CK-Prest, PTT-A and Cephascreen on STA-R, with Actin FS on BCS and with Synthasil on ACL TOP and Thrombin Time (TT) on STA-R. More specific tests included Prothrombinase-induced Clotting Time (PiCT, Pefakit®, Pentapharm) and Ecarin Clotting Time (ECT) on STA-R; dilute PT (dPT) and activated Clotting time (ACT) on KC10. Thrombin Generation Tests (TGT) were also performed with Calibrated Automated Thrombogram (CAT) using PPP or PPP LOW reagents (Thrombinoscope). Sensitivity of a coagulation assay was defined as the concentration required to double clotting time (2XCT).

Results:

Dabigatran.
A concentration-dependent prolongation of PT, dPT and aPTT was observed. However, at high concentrations, sensitivity of aPTT decreased whereas sensitivity of PT increased. The results varied depending on the clotting reagent used. TT was too sensitive leading to high variability for concentrations higher than 250nM. PiCT showed a linear concentration coagulation time (CT) relationship until a plateau at 750nM. ECT showed a high sensitivity (2X CT &ap; 93 nM), a linear relationship in the whole concentration range and a very low variability (CV≤ 1,8%). ACT gave a similar profile to PT (Innovin) with a slightly higher sensitivity (2X CT &ap; 716 nM vs 842nM). Dabigatran induced a concentration-dependent delay and inhibition of the tissue factor-induced thrombin generation with 5 pM TF or 1 pM TF with 4 μM PL and 16.7 mM CaCl2. The drug strongly increased the lag time and Tmax whereas it slightly decreased the Cmax and ETP. The lag time was the most sensitive CAT parameter with a high sensitivity (2x lag time &ap; 145 nM) and a low variability (CV≤6%).

Rivaroxaban.
A concentration-dependent prolongation of PT, dPT and aPTT was observed. However, at high concentrations, sensitivity of aPTT increased. Sensitivity of dPT (2X CT &ap; 109 to 550 nM) and PT (2X CT &ap; 138 to 764 nM) were similar and superior to aPTT (2X CT &ap; 897 to 2050 nM). The results varied depending on the clotting reagent used. TT was insensitive to rivaroxaban until 2500nM. Both PiCT and ECT showed a low sensitivity (2X CT &ap; 2536 nM and 5750 nM, respectively). A concentration-dependent prolongation of ACT was observed until 2500nM (2X CT &ap; 2275 nM). Rivaroxaban induced a concentration-dependent reduction and delay of the TF-induced thrombin generation. The Cmax was more strongly decreased than the ETP whereas the Tmax was more prolonged than the lag time, showing a major influence on the amplification phase. The Cmax was the most sensitive CAT parameter with a high sensitivity (Cmax EC50 &ap; 50 nM) and a low variability (CV<1%).

Conclusions:
ECT and PT are the most appropriate tests to study the pharmacodynamic effects of dabigatran and rivaroxaban respectively, thanks to high sensitivity, linear relationship in the whole concentration range and low variability.

Disclosures:
No relevant conflicts of interest to declare.