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Monitoring Direct Thrombin Inhibitors and Oral Anti-Xa Antithrombotics

Presenter: Dorothy Adcock-Funk, MD
Panelists: Kandice Kottke-Marchant, MD, PhD; Marisa B. Marques, MD; John D. Olson, MD, PhD
Moderator: George A. Fritsma, MS MT (ASCP)

Precision BioLogic Laboratory Medicine Roundtable, June 20 and 21, 2008, Dartmouth, Nova Scotia

In addition to Wendy Porteous, Steve Duff and Michael Scott, I would like to acknowledge Dean Willett, Sandy Morrison, and all the Precision BioLogic Inc. folks who participated in our round-table discussion.

We have employed the time-honored partial thromboplastin time assay (PTT, activated partial thromboplastin time, APTT) to monitor unfractionated heparin (UFH) and the intravenous direct thrombin inhibitors (DTIs) Argatroban (Novostan®), Lepirudin (Refludan®) and Bivalirudin (Angiomax®). Though we have developed and published therapeutic target ranges, the PTT suffers from several interferences. It is prolonged by parallel warfarin therapy, lupus anticoagulant, and congenital or acquired factor deficiency. In these circumstances the PTT anticoagulant response is exaggerated and unreliable. Conversely, elevated factor VIII and fibrinogen shorten the PTT and underestimate anticoagulant effect, a circumstance called in vitro drug resistance. The PTT does not lend itself to inter-laboratory normalization, as a variety of reagent and instrument combinations generate diverse anticoagulant responses.

There are several alternatives to the PTT. In theory, the thrombin clotting time (TCT) could be used to monitor DTI therapy, however in its typical formulation using 1-2 IU/mL, the assay is exquisitely sensitive.

The ecarin clotting time (ECT) has been on the drawing board for years. Ecarin is a thrombin-like snake venom from Echinus carinatus that cleaves prothrombin to produce an intermediate activation product, meizothrombin, which stimulates fibrin polymerization. A well-designed ECT is reliably linear to DTI plasma concentration, even at small prophylactic doses. The ECT is unaffected by heparin or warfarin and cannot be used to monitor these, neither is it sensitive to low molecular weight heparin (LMWH) or Fondaparinux. This is also advantageous as the ECT can be used to monitor DTIs while patients are being initiated on warfarin therapy. Another advantage of ECT is that it is not sensitive to the presence of lupus anticoagulants. Current ECT reagents are poorly standardized with lot-to-lot variability, and no international calibrators are available. Few North American reference laboratories and no clinical providers offer the ECT assay.

Pentapharm’s (Centerchem) Prefakit prothrombinase-induce clotting test (PiCT®) applies a mixture of phospholipids, Russell viper venom factor V activator (RVV-v), and bovine Xa to monitor therapeutic inhibition of Xa and IIa. The assay is stable and reliable, however standard curves must be generated for each anticoagulant, and the assay is affected by fibrinogen, FII, and FV levels when they are less than or equal to 25% of normal. PiCT may be factitiously prolonged by warfarin and lupus anticoagulants.

Chromogenic anti FXa and anti FIIa assays rely on patient or reagent antithrombin and produce a colorimetric result inversely proportional to the drug plasma concentration. The anti-Xa has been applied successfully to UFH, LMWH and Fondaparinux monitoring, and has been enhanced so that both UFH and LMWH may be monitored on a single “hybrid” curve. These assays are not currently formulated to be DTI-sensitive. They could potentially be modified to assay currently investigational oral direct Xa inhibitors Apixaban (BMS, Inc), Rivaroxaban (Bayer, Inc; currently under FDA review), and at least five additional oral anti-FXa drugs under development. The anti-FXa assay may be ineffective in monitoring the developmental oral DTI Dabigatran (Pradax®, Boehringer Ingelheim), which, like Rivaroxaban, may be nearing release in North America. Release of oral anti-FXa and anti-FIIa may revolutionize antithrombotic therapy.

We wish there were a universal antithrombotic drug assay with a common standard. The recently reborn thrombin generation time assay (TGT, TGA) may be developed to meet these lofty requirements. TGT is an automatable, clot-based assay that lends itself to numerous modifications designed to detect a variety of anticoagulant effects and coagulopathies. TGT development will require a number of careful reagent standardizations and formulations.

Clinical trial data for investigational antithrombotics provide little data to generate therapeutic ranges. This may be by design, as developers prefer to emphasize there is little need for laboratory monitoring. Experience with LMWH and Fondaparinux however, teaches that laboratory monitoring becomes necessary under a variety of clinical circumstances. Upon their release, laboratory researchers may scramble to develop monitoring protocols that empirically determine therapeutic ranges. Perhaps there could be a concerted effort uniting laboratory scientists and pharmaceutical distributors towards the common goal of good anticoagulant monitoring for current and developmental drugs.

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