J. J. Van Veen

Thrombin generation testing in routine clinical practice: are we there yet?

Thrombin is the central enzyme in the coagulation cascade. Estimation of an individual’s potential to generate thrombin may correlate more closely with a hyper‐ or hypo‐coagulable phenotype, compared to traditional coagulation tests. The possible correlation and recent technical advances in thrombin generation measurement has caused a significant interest in the method and the development of commercial assays. Several variations of the assay exist depending on the defect to be investigated. Fluorogenic thrombin generation assays have acceptable intra‐laboratory variation but a higher inter‐laboratory variation. Variation in preanalytical variables makes comparisons between studies difficult. Thrombin generation is highly variable between individuals and there are suggestions that this may allow individualized treatment based on global haemostatic response in patients with bleeding disorders or on anticoagulant therapy. In patients with thrombotic disorders it may be possible to identify those at higher risk of recurrent thrombosis. For both scenarios, however, data from large prospective studies are lacking or inconclusive and a good relationship between thrombin generation and phenotype remains to be established. Further standardization of the assay is needed before large multicentre studies can be conducted and until then thrombin generation in routine clinical practice is not yet a reality.

Antiviral therapy for chronic hepatitis C in patients with inherited bleeding disorders: an international, multicenter cohort study

Summary.  Background: Hepatitis C is a major co‐morbidity in patients with hemophilia. However, there is little information on the efficacy of antiviral therapy and long‐term follow‐up after treatment.Objectives: To assess the effect of interferon‐based (IFN‐based) therapy on hepatitis C virus (HCV) eradication, to identify determinants associated with treatment response, and to assess the occurrence of end‐stage liver disease (ESLD) after completing antiviral therapy.Patients and methods: In a multicenter cohort study, 295 treatment‐naïve hemophilia patients chronically infected with HCV were included. The effect of therapy was expressed as sustained virological response (SVR). Determinants associated with treatment response were expressed as odds ratios (ORs). Cumulative incidence of ESLD was assessed using a Kaplan–Meier survival table.Results: Among human immunodeficiency virus (HIV) negative patients (n = 235), SVR was 29% (29/101) for IFN monotherapy, 44% (32/72) for IFN with ribavirin, and 63% (39/62) for pegylated IFN (PegIFN) with ribavirin. In patients co‐infected with HIV (n = 60), IFN monotherapy, IFN with ribavirin, and PegIFN with ribavirin eradicated HCV in 7/35 (20%), 1/2 (50%), and 11/23 (48%), respectively. SVR increased with genotype 2 and 3 [OR 11.0, 95% CI: 5.8–20.5], and combination therapy (IFN and ribavirin OR 3.7, 95% CI: 1.7–8.4), PegIFN and ribavirin (OR 4.2, 95% CI: 1.8–9.5). Up to 15 years after antiviral treatment, none of the patients with a SVR relapsed and none developed ESLD. In contrast, among unsuccessfully treated patients the cumulative incidence of ESLD after 15 years was 13.0%.Conclusions: Successful antiviral therapy appears to have a durable effect and reduces the risk of ESLD considerably.

Wide variation in thrombin generation in patients with atrial fibrillation and therapeutic International Normalized Ratio is not due to inflammation.

Atrial fibrillation (AF) is a common cardiac arrhythmia with a 5–20% annual risk of stroke. Warfarin reduces this risk by at least 60%. Despite adequate anticoagulation within the target International Normalized Ratio (INR) range of 2·0–3·0, some patients still experience thrombotic and bleeding events. It is now possible to assess the intensity of anticoagulation with automated thrombin generation (TG) tests, such as the calibrated automated thrombogram (CAT). These tests were compared and an inverse relationship was found between the INR and CAT in 143 elderly AF patients. There was equally good correlation between the concentration of factors II, VII, IX and X and the INR and TG parameters. The peak thrombin was most strongly associated with the concentration of prothrombin fragment 1 + 2 in plasma. There was wide variability in TG parameters in patients with identical INR values, sometimes up to a fourfold difference. This TG variability in individuals with the same INR is not due to inflammation, at least when the latter is measured as the concentration of factor VIII coagulant activity, von Willebrand factor antigen, high sensitivity C‐reactive protein and fibrinogen. It was concluded that, although the TG and INR were closely correlated there was wide variability in peak thrombin and endogenous thrombin potential in patients within the INR therapeutic range, the cause of which remains unclear.

Optimizing warfarin reversal – an ex vivo study

Summary.  Background: Warfarin reversal is a common clinical situation. This is commonly performed using vitamin K and, depending on the urgency, fresh frozen plasma (FFP), prothrombin complex concentrates (PCCs), or activated factor VII. Even though PCCs are widely used, the ideal dosing regimen is far from established. Objectives: To verify differences in warfarin reversal patterns using FFP, recombinant FVIIa (rFVIIa), and PCC; and to test the hypothesis that supratherapeutic International Normalized Ratios (INRs) might not correlate with thrombin generation (TG) and identify the ideal concentrations of PCC required to reverse various INR thresholds. Methods: We studied the effects of FFP, rFVIIa and Beriplex P/N on the INR and TG, using the calibrated automated thrombography assay in ex vivo warfarinized plasma. Plasmas with different INRs were spiked with different concentrations of Beriplex P/N. Results: Beriplex P/N was the only agent that completely normalized TG and the INR. The endogenous thrombin potential (ETP) and the peak thrombin showed a significant negative correlation with all INRs. The ETP and velocity of TG reached a plateau at an INR of ∼ 4.0. A concentration equivalent to a dose of 30 IU kg−1 Beriplex P/N normalized the ETP, the INR, FII, FVII, FIX and FX of samples with INRs ≥ 4.0. Higher doses resulted in hypercoagulable TG patterns. A concentration equivalent to a dose of 20 IU kg−1 was sufficient to reverse warfarin at an INR range of 2.0–3.9, as judged by the same tests. Conclusions: Warfarin reversal algorithms could be simplified with the adoption of this strategy utilizing two doses of PCC, depending on the INR of the patient. This would also lead to cost reductions and, possibly, a reduction in thrombotic risk.

Routine preoperative coagulation tests: an outdated practice?

Routine coagulation tests have been used for many years in the preoperative setting in the belief that they identify patients who may have acquired or congenital bleeding disorders, and on the assumption that testing will predict perioperative bleeding allowing treatment to be given and prevent it. Coagulation ‘screens’ typically include the prothrombin time (PT) and the activated partial thromboplastin time (APTT). Both tests were originally developed to aid in the diagnosis of inherited bleeding disorders such as haemophilia and were not intended as screening tests. Activation of coagulation can be achieved through the intrinsic or the extrinsic pathway leading to activation of the common pathway and conversion of fibrinogen to fibrin leading to clot formation. Although the cascade model of coagulation is not physiological, it is useful as a means to understand the mechanism by which traditional coagulation tests detect coagulopathies. The APTT activates plasma typically with substances such as kaolin or silica which cause activation of the contact pathway and, subsequently, the intrinsic and common coagulation pathways. A deficiency in any of these pathways will therefore prolong the APTT. The PT is activated with supraphysiological concentrations of tissue factor and detects FVII deficiency and also deficiencies in the common pathway. Both tests can also be prolonged by the presence of a lupus anticoagulant, inhibitors such as anticoagulants, and acute conditions associated with an acquired bleeding state. Inherited coagulation defects are rare. The incidence of haemophilia A and B is estimated at 1:5000 2 and 1:30 000 male births, respectively. Severe, clinically relevant, deficiencies of FII, FV, FVII, FX, and fibrinogen are even rarer with an incidence varying between 1:300 000 and 1:2 000 000, although this may be higher in populations where consanguineous marriages are more common. FXI deficiency is common in Ashkenazi Jews with a prevalence of 8% but rare in a general population. The majority of patients with these bleeding disorders will be aware of their diagnosis through either a personal or family history of bleeding and will be registered at specialized haemophilia centres. Indiscriminate screening by routine coagulation testing will therefore only very rarely identify previously undetected individuals. In contrast, prolongation of the APTT is a common occurrence with the most common reasons being mild FXII deficiency and the presence of a lupus anticoagulant, neither of which is associated with a bleeding tendency. Moderate and severe FXII deficiency was found in 2.3% of otherwise healthy Austrian blood donors and in 10.3% of patients undergoing cardiac surgery. Lupus anticoagulant can be found in 1.2–3.8% of healthy individuals, but the incidence increases with age and chronic disease, and was found in up to 30% of patients with systemic lupus erythematosus. The ability of a lupus anticoagulant to prolong the APTT or PT depends on the combination of reagents and analysers used and may therefore vary between laboratories. Other causes of a prolonged APTT not associated with a bleeding tendency include high molecular weight kininogen deficiency and prekallikrein deficiency. Finally, a normal range is calculated by the mean +2 standard deviations of measurements in healthy, non-bleeding subjects and by definition 2.5% of measurements in normal individuals will show a prolonged clotting time. Therefore, if routine coagulation testing is done to identify previously undiagnosed bleeding disorders, it is much more likely to identify a prolonged routine coagulation test that is not associated with a bleeding tendency. In practice, this can lead to further unnecessary Volume 106, Number 1, January 2011

Management of peri-operative anti-thrombotic therapy.

The management of antithrombotic therapy in the peri‐operative setting is a common problem, balancing haemorrhagic risk with continued treatment and thrombotic risk when discontinued. High‐quality evidence is lacking regarding the optimal approach for patients on oral anticoagulants or antiplatelet agents. This review discusses the available evidence for the management of patients on warfarin, non‐vitamin K antagonist oral anticoagulant drugs, and antiplatelet therapy in the peri‐operative setting. Bridging therapy for patients on warfarin should be considered for those at highest risk of thrombosis, whereas it may not be necessary for those on non‐vitamin K antagonist oral anticoagulant drugs given the reduced time off anticoagulation and their more predictable pharmacokinetics. Aspirin can be continued for most procedures. Dual antiplatelet agents for patients with a recently inserted coronary artery stent should be continued if possible but decisions should be individualised and taken after multidisciplinary discussion.

The effect of tissue factor concentration on calibrated automated thrombography in the presence of inhibitor bypass agents

Thrombin generation has been suggested as a method to monitor treatment with factor eight inhibitor bypassing activity (FEIBA®) or recombinant FVIIa (rFVIIa). The sensitivity of the assay for individual coagulation factors is dependent on the tissue factor (TF) concentration. An inverse relation between the rFVIIa concentration needed to shorten the clotting time and TF concentration has been shown but the data on thrombin generation are inconsistent. Information on TF concentration in measurements with FEIBA® is limited. We studied the influence of TF concentration (1 and 5 pm) on thrombin generation through spiking experiments with rFVIIa and/or FEIBA® in the plasma of severe haemophilia A patients and after four and three treatment episodes, respectively, using the calibrated automated thrombin generation assay (CAT) in platelet poor plasma. Spiking with FEIBA® showed a linear relation with the endogenous thrombin potential (ETP)/peak at 1 pm but substrate depletion at 5 pm. Spiking with rFVIIa showed a near linear dose–response relation with the ETP/peak at 1 pm but only a shortening of the initiation phase at 5 pm. Similar effects were present in post‐treatment samples. FEIBA® acted synergistically with rFVIIa. This suggest a role for CAT in monitoring inhibitor bypass treatment but low TF concentrations are required to avoid substrate depletion with FEIBA® and to demonstrate the effect of rFVIIa.

Effect of extremes of body weight on drug level in patient treated with standard dose of rivaroxaban for venous thromboembolism; real life experience

Venous thromboembolism (VTE) is a leading cause ofmorbidity and mortalityworldwide. Appropriate anticoagulation is crucial in the treatment and prevention of recurrent VTE. Overdosing of an anticoagulant could result in life-threatening bleeding while under-dosing could result in lack of efficacy leading to recurrence of thrombosis. It is possible that “extremes in body weight” [EBW] (b50 kg or N120 kg) may alter the exposure profile of an anticoagulant and its benefit: risk ratio. Monitoring of anticoagulant intensity of direct acting oral anticoagulants (DOACs) is not routinely required due to their predictable anticoagulant effects but assessment of drug levels may be useful in some situations including EBW as recommended by both International Society for Thrombosis and Haemostasis (ISTH) [1] and the British Committee for Standards in Haematology (BCSH) guidelines [2]. Patients with EBW, both very low and very high, are underrepresented in clinical trials and in preclinical dose-finding studies. The mean weight was around 84 kg; with the majority of participants in phase III clinical studies weighing between 60 and 100 kg [3]. Sub-analysis of EINSTEIN DVT and PE studies in which weight was categorised into three groups as 50 kg, 50–100 kg and N100 kg, foundno association between body weight and the risk of recurrent VTE and similar rates of recurrent VTE by treatment group (2.3% on rivaroxaban vs 2.0% on VKA in patients N100 kg) [4]. However, none of the phase III trials with DOACs reported the patients with body mass index (BMI) N40 kg/m or their clinical outcome. The guidance from the SSC of the ISTH on use of DOACs in obese patients 2016 [3], recommended against the use of DOACs in patients with BMI N40 kg/m2 or the weight N 120 kg due to limited clinical data in this group of patients. We assessed the effect of EBWon rivaroxaban levels and clinical outcome with standard dose of rivaroxaban (15 mg BD for three weeks followed by 20 mg once daily) in the treatment of venous thromboembolism (VTE) in a routine clinical practice. The study was untaken as a part service evaluation project in a tertiary haemostasis and thrombosis centre in UK and approved by the trust clinical effectiveness unit. An unselected group of 219 patients with VTE (53% deep vein thrombosis [DVT], 41% pulmonary embolism [PE] and 6% PE and DVT: 51% male, 49% female, mean age 59 years) were studied. Rivaroxaban levels were measured in blood samples taken 2–4 h from the last dose of rivaroxaban in patients on rivaroxaban 20 mg daily after an initial

Limitation of the activated partial thromboplastin time as a monitoring method of the direct thrombin inhibitor argatroban

Argatroban is licensed for patients with heparin‐induced thrombocytopenia and monitoring is conventionally by activated partial thromboplastin time (APTT) ratio with a target of 1.5–3.0 and not exceeding 100 s. The APTT may be influenced by coagulopathies, lupus anticoagulant and raised FVIII levels. Variable but not clinically significant sensitivity of APTT reagents to argatroban has been highlighted in other studies.

Reduced cardiovascular mortality in hemophilia despite normal atherosclerotic load

See also Zwiers M, Lefrandt JD, Mulder DJ, Smit AJ, Gans ROB, Vliegenthart R, Brands‐Nijenhuis AVM, Kluin‐Nelemans JC, Meijer K. Coronary artery calcification score and carotid intima–media thickness in patients with hemophilia. This issue, pp 23–9; Biere‐Rafi S, Tuinenburg A, Haak BW, Peters M, Huijgen R, de Groot E, Verhamme P, Peerlinck K, Visseren FLJ, Kruip MJHA, Laros‐van Gorkom BAP, Gerdes VEA, Buller HR, Schutgens REG, Kamphuisen PW. Factor VIII deficiency does not protect against atherosclerosis. This issue, pp 30–7.