Selection of a Suitable Rapid Turn Around and Sensitive Test for the Laboratory Monitoring of Newer Oral Anticoagulants, Rivaroxaban, Apixaban and Dabigatran,

Abstract 3363

Recently, several of the newer oral anticoagulants such as Rivaroxaban (Bayer Healthcare) (R) Apixaban (BMS/Pfizer) (A) and Dabigatran (Boehringer Ingleheim) (D) have been approved for different indications in the US and European countries. Although the monitoring is not required or promoted to manage the approved dosages of these drugs in different indications, population based bleeding complications with some of these agents have been reported warranting the monitoring of these drugs to optimize therapy. Conventionally the Xa based drugs such as Rivaroxaban and Apixaban are monitorable by following the anti-Xa activity whereas the dabigatran can be followed by anti-IIa activity. The pharmacokinetics of these drugs is reportedly monitored by such sophisticated methods as the liquid chromatographic – mass spectrometric methods. The conventional clotting methods such the PT/INR and APTT assays provide widely variable results which are dependent on the type of reagents used. Heptest exhibits a higher sensitivity to the anti-IIa agents in comparison to the anti-Xa agents. Prothrombinase activated clotting time (PiCT) is a Xa/IIa based clotting assay which can be performed in using a 1 stage and 2 stage method. The 1 stage method is highly sensitive to newer anticoagulants in contrast to the 2 stage PiCT and other tests. For the routine monitoring of these newer oral anticoagulant drugs a simple and rapid assay with the linear sensitivity range of 0–500 ng/ml is optimal for various patient populations. The current study is designed to determine the relative sensitivity of the one stage PiCT test in comparison to 4 PT, 4 APTT, Heptest and thrombin generation assays. Normal human pooled plasma was supplemented with R, A and D at a concentration range of 0–10 μg/ml. PT/INR analysis was carried using Thromboplasin C+, Innovin, Neoplastin and Simplastin reagents. The APTT assays were carried out using Actin FS, Actin FSL, Stago APTT and Platelin reagents. Heptest and one stage and two stage PiCT assays were also performed. All of the clot-based assays were carried out on the ACL analyzer with the exception of Heptest and PiCT tests. The thrombin generation assay was carried out using a flourogenic substrate method (Technoclone). In the PT assay, all reagents showed assay based variabilities in the clotting times and were relatively insensitive to A, however R and D produced a concentration dependent anticoagulant effect. Which were relatively insensitive at concentrations below 1 μg/ml. Similarly in the APTT assays, while R and D produced a concentration dependent anticoagulant effect all 4 reagents were relatively insensitive to A. The Actin FSL reagent was found to be sensitive to both the R and D and concentrations of less than 100 ng/ml were measurable. D produced the strongest anticoagulant effects in both the PT and APTT assays. Heptest was relatively insensitive to R and A, however at concentrations of greater than 1 μg/ml it produced concentration dependent anticoagulant effects. In the one stage PiCT assay, all agents including A prolonged the anticoagulant effect which produced a linear response at submicrogram levels and was found to be highly sensitive to D. The two stage PiCT also exhibited a linear sensitivity in the 0–100 ng/ml range. In the thrombin generation generation assay, R exhibited the strongest inhibitory effect with an IC₅₀ of 160 ng/ml, D at 310 ng/ml and A produced a relatively weaker inhibitory effects with an IC₅₀ >1000 ng/ml. These studies clearly demonstrate the limitations of various laboratory assays in the monitoring of newer oral anticoagulant agents with the exception of 1 stage PiCT test which can be universally used in the monitoring of these agents in the projected clinical ranges. Furthermore the one stage PiCT test can be performed in such matrices as whole blood, PRP and PPP. Moreover this test can also be carried out in point of care settings. Additional results on the assay performance and the clinical utility of some of these tests will be presented with a recommendation for the monitoring of newer oral anticoagulant drugs in such clinical settings as the extended prophylaxis of DVT and atrial fibrillation.