Marie-Hélène Horellou

Assessment of apixaban plasma levels by laboratory tests: suitability of three anti-Xa assays

While laboratory monitoring is not required in patients treated with apixaban, a direct factor-Xa inhibitor, assessment of its concentration is useful in some critical situations. However, few data are available on its effect on coagulation tests and on the suitability of anti-Xa assays for its quantification. It was the objective of this study to identify laboratory tests suitable for apixaban concentration assessment. Coagulation tests - PT and aPTT- and anti-Xa assays were performed in apixaban-spiked plasma samples. To evaluate the sensitivity of PT and aPTT to apixaban, we conducted a first monocenter part, with a wide range of concentrations (50-1,000 ng/ml), a large panel of reagents (20 reagents), and two coagulometers (STAR®, Stago and ACL TOP®, IL), and a second multicenter part involving 13 laboratories using either a common PT reagent (RecombiPlastin2G®) or the local PT and aPTT reagents. In the multicentre part, five blinded apixaban-spiked plasma samples (0/100/200/400/800 ng/ml - checked by HPLC-MS/MS) were used; apixaban concentrations were measured with three anti-Xa assays, apixaban calibrators and controls (Stago). PT and aPTT tests using a large panel of reagents displayed a low sensitivity to a wide range of apixaban concentrations. The concentrations to double PT ranged from 400 to >1,000 ng/ml with the 10 reagents. With the three anti-Xa assays, inter-laboratory precision and accuracy were below 11% and 12%, respectively. In conclusion, whereas PT and aPTT tests were not sensitive enough to detect apixaban, the three anti-Xa assays tested using lyophilised apixaban calibrators and controls allowed to reliably quantify a wide range of apixaban concentrations.

A vitamin K epoxide reductase complex subunit-1 (VKORC1) mutation in a patient with vitamin K antagonist resistance.

We would like to thank the Department of Clinical Epidemiology and Medical Technology Assessment, Academic Hospital Maastricht, University of Maastricht, The Netherlands, for providing blood samples of the patients included in that center for the MALT study, the personnel of the Diagnostic Experimental Vascular Medicine Laboratory for their expert technical assistance, and professor C.J.F. Van Noorden for his constructive comments.

At III content and antithrombin activity in oestrogen-progestogen and progestogen-only treated women

Serum antithrombin (AT) and AT 3 content in plasma and serum were measured in 204 women taking either 50 or 30 mg of estrogenprogestogen preparations, or a progestogen only pill, to determine the estrogen content on antithrombin levels. AT 3 determination was done with the Mancini technique, and serum AT activity was measured by the van Kaulla method. 140 controls were also observed before oral contraception. Results of the study showed a decrease in serum AT 3 and in serum AT activity with combined pills containing 50 mg of ethinyl estradiol, but the decrease was not time-dependent after the 3rd. month of treatment. A relevant decrease was also observed with pills with a 30 mg. of estrogen content, while progestin only pills did not induce a decrease in either AT 3 or AT activity.

Red blood cell aggregability in patients with a history of leg vein thrombosis: influence of post-thrombotic treatment

Summary. Reversible aggregation or red blood cells (RBC) plays an important role in determining blood flow properties, and it is this aggregation which increases blood viscositgy at low shear rates. The structure and sites of venous thrombi, as well as the fact that stasis is a major predisposing factor in venous thrombosis, suggest a strong association between vein thrombosis, slow blood flow and increased blood viscosity. RBC aggregation and disaggregation were measured (SEFAM erythroaggregameter, France) in 54 patients with a history of unexplained leg vein thrombosis. Results were compared to those of controls classified according to age. Increased RBC aggregability was observed in 41% of the patients, and the mean values indicated a significant elevation of RBC aggregability in patients when compared with controls (P < 0.05). Subgroups were compared to study the influence of thrombus recurrence and thrombosis type (deep versus superficial vein thrombosis) on the aggregation parameters. No significant difference was found between these subgroups. The use of compression stockings and veinotropic drugs tended to reduce the abnormalities in RBC aggregability (P < 0.05). An increase in RBC aggregability and in the shear resistance of RBC aggregates, by predisposing ro circulatory stasis, is likely to contribute to the evolution and complications of leg vein thrombosis.

Multiple genetic alterations in vitamin K epoxide reductase complex subunit 1 gene (VKORC1) can explain the high dose requirement during oral anticoagulation in humans.

L . BODIN ,* J . PERDU, M. DIRY ,* M. -H . HORELLOU and M. -A . LO R I OT *§ *Université Paris Descartes, INSERM U-775, Paris; Assistance-Publique Hôpitaux de Paris, Hôpital Européen Georges Pompidou, Département de Génétique, Centre National de Référence des Maladies Vasculaires Rares, Paris; Assistance Publique Hôpitaux de Paris, Hôpital Hôtel-Dieu, département d Hématologie Biologique, Paris; and §Assistance-Publique Hôpitaux de Paris, Hôpital Européen Georges Pompidou, Service de Biochimie, Pharmacogénétique et Oncologie Moléculaire, Paris, France

Pregnancy-associated venous thromboembolism (VTE) in combined heterozygous factor V Leiden (FVL) and prothrombin (FII) 20210 A mutation and in heterozygous FII single gene mutation alone.

Summary. The risk of venous thromboembolism (VTE) in the absence of prophylaxis was evaluated in a retrospective study of 47 women (84 pregnancies) with combined thrombophilia [heterozygous factor V Leiden (FVL) plus prothrombin (FII) 20210A mutation (group I)] and in 82 women (193 pregnancies) with the FII alone (group II). VTE was more frequent in group I than in group II [17·8% versus 6·2%, P = 0·003, relative risk (RR) 2·9, 95% confidence interval (CI) 1·4–5·9], ante partum (7·1% and 2·1%) and post partum (11·5% and 4·2%). The risk was higher in index cases than in family members (RR 2·5, 95% CI 1·2–5·2 and RR 2·1, 95% CI 0·2–22·3 respectively) Even women who had no history of VTE before pregnancy had an increased risk (RR 2·2, 95% CI 1·0–4·8). Our results suggest that, during ante partum, prophylaxis is indicated in women with combined thrombophilia and with a VTE before pregnancy. In those without VTE before pregnancy, prophylaxis might be decided for each individual case, taking into consideration all risk factors. In women with the FII mutation alone, the low risk may not justify prophylaxis in the absence of previous VTE. In post partum, prophylaxis is indicated in all cases.

Diagnosis And Clinical Characteristics Of Inherited Activated Protein C Resistance

The diagnostic strategies and clinical characteristics of thrombophilia associated with heterozygous or homozygous factor V Leiden mutation have been determined according to the literature and to a personal study in 51 families. Factor V mutation was present in the 51 propositi and in 84 out of 125 family members (81 heterozygous, 3 homozygous). Venous thrombosis was observed in all the propositi, in 17 of the 84 family members with the mutation and in 6 of the 41 with a normal APC resistance test and no mutation. An associated protein C or protein S deficiency was present in 5 families (10%). The most frequent clinical manifestations were superficial or deep vein thrombosis and/or pulmonary embolism, but also thrombosis at an unusual site (cerebral, mesenteric or central retinal vein). A causal relationship is frequently difficult to demonstrate. A precipitating factor was observed in 84% of cases and a recurrent thrombotic episode occurred in 50% of propositi. The risk of thrombosis associated with pregnancy was high in the post-partum period, especially in homozygous women. In the 28 homozygous subjects, markers of coagulation activation were frequently elevated in untreated patients. Finally, the efficacy of anticoagulant treatment is suggested but the long period often observed between treatment interruption and a recurrence does not militate in favour of long term treatment.

Risk assessment models for thromboprophylaxis of medical patients.

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 127 What is a risk factor and how to classify? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 128 Quantification of Risk in a Multifactorial Disease . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 128 RAMs Objectives and Process . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 128 Development of RAMs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 128 RAMs in the literature . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 128 Non-validated RAMS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 128 Validated RAMs in the literature . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 129 Non electronic alerts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 130 RAMs in medical outpatients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 130 Discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 131 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 131

Antiplatelet Effects of the Addition of Acetylsalicylic Acid 40 Mg Daily to Ticlopidine in Human Healthy Volunteers

Clinical studies have shown that acetylsalicylic acid (ASA) or ticlopidine (T) bring a partial clinical benefit in a subset of patients threatened by thrombosis on atherosclerotic plaques. Acetylsalicylic acid and T have different impacts on platelet function and the combination of the drugs seems logical to achieve a greater antiplatelet effect. Healthy volunteers were randomly allocated to any of the three treatment groups: T 250-placebo; T 250-T 250; placebo-placebo (treatment was administered in a double blind manner). Acetylsalicylic acid 40 mg was openly administered once a day to the subjects of the three groups after the first week of treatment. Simplate I bleeding time and platelet aggregation testing were performed before treatment and at the end of the two treatment periods. We confirmed that T alone prolongs bleeding time and impairs, in a dose-dependent manner, ADP-induced aggregation; platelet responses that depend on released ADP were also affected. Inhibition of thromboxane-dependent aggregation, associated with a doubling of the bleeding time, was observed after one week of low-dose ASA inhibition of thromboxane synthesis. An additive effect of ASA 40 mg to T 250 mg on the bleeding time was evidenced. There was also a wider alteration of platelet aggregation, with a trend towards an inhibition of the response to a high concentration of collagen as compared to ASA or T used alone. Such a powerful, logical drug combination is in good agreement with preliminary encouraging results obtained after coronary stent implantation and deserves further studies in patients at high risk for arterial thrombosis to define its benefit-risk profile. Key Words: Aspirin—Ticlopidine— Drug combination—Bleeding time—Platelet aggregation.

Laboratory Monitoring of a Low Molecular Weight Heparin (Enoxaparin) with a New Clotting Test (Heptest)

A new simple clotting test (Heptest) for low molecular weight heparins was compared to anti-Xa determination by an amidolytic assay in volunteers and in patients receiving standard calcium heparin or low molecular weight (LMW) heparin (Enoxaparin) by subcutaneous administration. The results obtained with both methods are in very good agreement. It seems that the Heptest, although influenced by various other clotting parameters, is nevertheless relatively specific for anti-Xa activity. Despite our positive first impression, we object to expressing the results in micrograms of heparin per milliliter or anti-Xa units. Until rigorous standardization is possible, we prefer to express the results as clotting times. Preliminary results warrant a more extensive study in order to assess the clinical relevance of Heptest in patients receiving unfractionated or LMW heparins.