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The view that clot time-based assays do not provide a sufficient assessment of an individual's hemostatic competence, especially in the context of anticoagulant therapy, has provoked a search for new metrics, with significant focus directed at techniques that define the propagation phase of thrombin generation. Here we use our deterministic mathematical model of tissue-factor initiated thrombin generation in combination with reconstructions using purified protein components to characterize how the interplay between anticoagulant mechanisms and variable composition of the coagulation proteome result in differential regulation of the propagation phase of thrombin generation. Thrombin parameters were extracted from computationally derived thrombin generation profiles generated using coagulation proteome factor data from warfarin-treated individuals (N = 54) and matching groups of control individuals (N = 37). A computational clot time prolongation value (cINR) was devised that correlated with their actual International Normalized Ratio (INR) values, with differences between individual INR and cINR values shown to derive from the insensitivity of the INR to tissue factor pathway inhibitor (TFPI). The analysis suggests that normal range variation in TFPI levels could be an important contributor to the failure of the INR to adequately reflect the anticoagulated state in some individuals. Warfarin-induced changes in thrombin propagation phase parameters were then compared to those induced by unfractionated heparin, fondaparinux, rivaroxaban, and a reversible thrombin inhibitor. Anticoagulants were assessed at concentrations yielding equivalent cINR values, with each anticoagulant evaluated using 32 unique coagulation proteome compositions. The analyses showed that no anticoagulant recapitulated all features of warfarin propagation phase dynamics; differences in propagation phase effects suggest that anticoagulants that selectively target fXa or thrombin may provoke fewer bleeding episodes. More generally, the study shows that computational modeling of the response of core elements of the coagulation proteome to a physiologically relevant tissue factor stimulus may improve the monitoring of a broad range of anticoagulants.

Bivalirudin, a synthetic analog of the carboxy-terminus of hirudin, is a reversible thrombin inhibitor used during coronary balloon angioplasty. The objective of this study was to evaluate the influence of bivalirudin on thrombin generation. Three in-vitro models (numerical simulations, synthetic coagulation proteome and whole blood) of contact pathway-independent blood coagulation triggered with tissue factor were used in this study. Increasing concentrations of bivalirudin prolong the initiation phase of thrombin generation in a concentration-dependent manner. At subpharmacologic bivalirudin concentrations (0.5-2 micromol/l), total thrombin generation was significantly increased. At a pharmacologic concentration (5 micromol/l), bivalirudin suppressed thrombin generation in the synthetic coagulation proteome; in numerical simulations and contact pathway-inhibited whole blood, no thrombin generation was detected over 1200-2000 s and platelet activation was inhibited by 80%. The addition of a pharmacologic concentration (9 micromol/l) of a naturally occurring protease inhibitor aprotinin in the presence of at least 0.5 micromol/l bivalirudin provided limited enhancement of the bivalirudin inhibitory effect. In conclusion, bivalirudin at pharmacologic concentrations is an efficient inhibitor of thrombin generation, platelet activation and clot formation, which acts not as a modulator but as an 'on-off' switch of blood coagulation.

Bleeding is the most feared and difficult to predict adverse event of anticoagulation. We sought to investigate whether calibrated automated thrombography (CAT) parameters are associated with minor bleeding (MB) in anticoagulated patients following venous thromboembolism (VTE). Enrolled were 132 patients on rivaroxaban, 145 on vitamin K antagonists (VKA) and 31 controls who stopped anticoagulation. Prior to the next dose of the anticoagulant, we measured CAT parameters, along with rivaroxaban concentration and INR. During a median follow-up of 10 months, we recorded minor and major bleedings. On rivaroxaban, 27 (20.5%) patients with MB had longer time to start thrombin generation, lower peak thrombin generation and lower endogenous thrombin potential compared with subjects without MB (all p < 0.001). All CAT parameters, except for peak thrombin generation (p = 0.049), were similar in VKA patients with (n = 25, 17.2%) vs. without MBs. By logistic regression, time to start thrombin generation (p = 0.007) and unprovoked VTE (p = 0.041) independently predicted MBs on rivaroxaban. Major bleedings were more frequent in patients with MBs (17.3% vs. 1.8%, p < 0.001). Abnormal CAT parameters characterize VTE patients prone to MBs on rivaroxaban, but not on VKA. Time to start thrombin generation measured about 24 h since the last rivaroxaban dose might help predict MBs. [ABSTRACT FROM AUTHOR]

Abstract: Tissue factor (TF) is the major physiological initiator of blood coagulation. It exists as an integral membrane protein that upon injury to the blood vessel becomes exposed to the blood stream and initiates a series of enzymatic reactions that cause blood to clot. TF is also found within circulating blood cells, requiring specific signaling events to promote its expression. In this review we will discuss current controversies concerning the structure–activity relationships of TF and contributions of TF to the hemostatic process, the potential roles of intravascular TF, including non-cell-bound TF and cell-expressed TF and the overall relationship between TF function and hemorrhage control. [Copyright &y& Elsevier]

Therapeutic agents that regulate blood coagulation are critical to the management of thrombotic disorders and surgical intervention. Many new compounds that were plausible in terms of target, relative specificity, mechanism and kinetic and thermodynamic properties have failed to perform as expected when assessed, at significant expense, in vivo. Our current deterministic mathematical model (Hockin et al. J. Biol. Chem. 2002, 277: 18322) describes the dynamic biochemical repair process that emerges in response to vascular injury and allows us to incorporate hypothetical inhibitors/enhancers at their proposed sites of interaction, thus providing an assessment of their efficacy. In conjunction with our coagulation proteome model (a synthetic reconstruction using purified protein of the components described in the computational model), we evaluated a set of anticoagulants currently in clinical use (unfractionated heparin, synthetic low molecular weight heparins, factor Xa inhibitors and thrombin inhibitors). The interactions of each anticoagulant with its target(s) were mathematically incorporated into the overall mathematical description of the reaction pathways. Outcomes for each anticoagulant at clinically relevant concentrations were generated and validated by comparisons with the synthetic coagulation proteome. Our results show that this type of mathematical methodology can lead to insights into drug performance and to rationalizing the differing properties that an ideal anticoagulant would require in therapeutic vs. prophylactic settings. [ABSTRACT FROM AUTHOR]