S. Performance of coagulation tests in patients on therapeutic doses of dabigatran: a cross-sectional pharmacodynamic study based on peak and trough plasma levels. J Thromb Haemost 2013; 11: 1493-1502.Summary. Background: Knowledge of anticoagulation status during dabigatran therapy may be desirable in certain clinical situations. Objective: To determine the coagulation tests that are most useful for assessing dabigatran's anticoagulant effect. Methods: Peak and trough blood samples from 35 patients taking dabigatran 150 mg twice daily, and one sample each from 30 non-anticoagulated individuals, were collected. Mass spectrometry and various coagulation assays were performed. 'Therapeutic range' was defined as the range of plasma dabigatran concentrations determined by mass spectrometry between the 2.5th and 97.5th percentiles of all values. Results: The therapeutic range was 27-411 ng mL À1. The prothrombin time (PT) and activated partial thromboplastin time (APTT), determined with multiple reagents, and activated clotting time (ACT) were insensitive to therapeutic dabigatran: 29%, 18% and 40% of samples had a normal PT, APTT, and ACT, respectively. However, normal PT, ACT and APTT ruled out dabigatran levels above the 75th percentile. The thrombin clotting time (TCT) correlated well and linearly with dabigatran levels below the 50th percentile, but was unmeasurable above it. The dilute thrombin time, ecarin clotting time and ecarin chromogenic assay showed linear correlations with dabigatran levels over a broad range, and identified therapeutic and supratherapeutic levels. Conclusions: The prothrombin time, APTT and ACT are often normal in spite of therapeutic dabigatran plasma levels. The TCT is useful for detecting minimal dabigatran levels. The dilute thrombin time and chromogenic and clotting ecarin assays accurately identify therapeutic and supratherapeutic dabigatran levels. This trial is registered at www.clinicaltrials.gov (#NCT01588327).

SCD patients with PHT have higher levels of markers of endothelial activation and other inflammatory markers than patients without PHT. A trend towards an increased level of markers of coagulation activation was observed in SCD patients with PHT compared with that in patients without PHT. Markers of hemolysis are associated with coagulation activation and endothelial dysfunction in SCD patients. Clinical trials of anticoagulants and anti-inflammatory agents are warranted in SCD patients with PHT.

The role of the factor IXa heparin-binding exosite in coagulation was assessed with mutations that enhance (R170A) or reduce (R233A) stability of the proteasefactor VIIIa A2 domain interaction. After tissue factor (TF) addition to reconstituted factor IX-deficient plasma, factor IX R170A supported a 2-fold increase in velocity index (slope) and peak thrombin concentration, whereas factor IX R233A had a 4-to 10-fold reduction relative to factor IX wild-type. In the absence of TF, 5 to 100 pM of factor IXa increased thrombin generation to approach TF-stimulated thrombin generation at 100% factor IX. Factor IXa R170A demonstrated a 2-to 3-fold increase in peak thrombin concentration and 5-fold increase in velocity index, whereas the response for factor IXa R233A was blunted and delayed relative to wild-type protease. In hemophilia B mice, factor IX replacement reduced the average time to hemostasis after saphenous vein incision, and the time to occlusion after FeCl 3 -induced saphenous vein injury. At 5% factor IX, the times to occlusion for factor IX wild-type, R170A, and R233A were 15.7 minutes, 9.1 minutes (P < . IntroductionThrombin is the penultimate product of the coagulation cascade, generated in an explosive burst on stimulation of plasma with limiting concentrations of tissue factor. 1,2 The measurement of plasma thrombin generation has merits as a global test of coagulation, and enhanced levels of thrombin generation have been associated with increased risk of recurrent venous thrombosis. 3 Thus, the rate and magnitude of thrombin generation may be predictive of the coagulation phenotype of patients. 4,5 In vitro and ex vivo modeling of the coagulation cascade indicates that factor X activation by the intrinsic tenase complex (factor IXa-factor VIIIa) is the rate-limiting step for thrombin generation. 1,2,6 Intrinsic tenase activity is unstable because of the diffusional loss of the noncovalently bound factor VIIIa A2 domain. 7,8 The instability of this enzyme complex is presumed to be an important regulator of the coagulation response.The mechanism(s) for activation of factor IXa within the intrinsic tenase complex are poorly understood. Factor VIII circulates as a heterodimer of A1-A2-B and A3-C1-C2 peptides with domain structure and metal-binding functions similar to ceruloplasmin. 9 Factor VIII undergoes proteolytic activation by thrombin, resulting in an unstable, metal-dependent A1/A2/A3-C1-C2 heterotrimer with cofactor activity. 10,11 Factor VIII or factor VIIIa light chain (A3-C1-C2 domains) bind to factor IXa on the phospholipid surface with an affinity that approaches intact factor VIIIa but lack cofactor activity. 12,13 The isolated factor VIIIa A1 domain also lacks cofactor activity. In contrast, the factor VIIIa A2 domain directly modulates the catalytic activity of factor IXa, which is further enhanced by the A1 domain, markedly increasing the k cat for factor X activation. 14,15 Although the isolated A2 domain binds with low affinity to factor IXa, it contributes significantly to protease-c...

Abstract`Thrombin aptamers' are based on the 15-nucleotide consensus sequence of d(GGTTGGTGTGGTTGG) that binds specifically to thrombin's anion-binding exosite-I. The effect of aptamer^thrombin interactions during inhibition by the serine protease inhibitor (serpin) heparin cofactor II (HCII) and antithrombin (AT) has not been described. Thrombin inhibition by HCII without glycosaminoglycan was decreased V Vtwo-fold by the aptamer. In contrast, the aptamer dramatically reduced thrombin inhibition by s 200-fold and 30-fold for HCII^heparin and HCII^dermatan sulfate, respectively. The aptamer had essentially no effect on thrombin inhibition by AT with or without heparin. These results add to our understanding of thrombin aptamer activity for potential clinical application, and they further demonstrate the importance of thrombin exosite-I during inhibition by HCII^glycosaminoglycans. ß

Summary.  Background: In an effort to improve interlaboratory agreement in the monitoring of unfractionated heparin (UFH), the College of American Pathologists (CAP) recommends that the therapeutic range of the activated partial thromboplastin time (APTT) be defined in each laboratory through correlation with a direct measure of heparin activity such as the factor Xa inhibition assay. Whether and to what extent this approach enhances the interlaboratory agreement of UFH monitoring has not been reported. Objectives: We conducted a cross‐validation study among four CAP‐accredited coagulation laboratories to compare the interlaboratory agreement of the anti‐FXa‐correlated APTT with that of the traditional 1.5–2.5 times the midpoint of normal (1.5–2.5:control) method for defining the therapeutic APTT range. Patients and methods: APTT and FXa inhibition assays were performed in each laboratory on plasma samples from 44 inpatients receiving UFH. Results: Using the anti‐FXa‐correlation method, there was agreement among all four laboratories as to whether a sample was subtherapeutic, therapeutic or supratherapeutic in seven (16%) patient samples. In contrast, consensus was achieved in 23 (52%) samples when the 1.5–2.5:control method was employed. Conclusions: The anti‐FXa‐correlation method does not appear to enhance interlaboratory agreement in UFH monitoring as compared with the traditional 1.5–2.5:control method. Adoption of the anti‐FXa‐correlation method produces considerable disparity in UFH dosing decisions among different centers, although the clinical impact of this disparity is not known.