Brynja R. Gudmundsdottir

The combination of recombinant factor VIIa and fibrinogen correct clotting ex vivo in patient samples obtained following cardiopulmonary bypass surgery.

Cardiac surgery involving cardio pulmonary bypass (CPB) may be associated with development of a coagulopathy that increases risk of bleeding. In the present ex vivo study we investigated the influence of fibrinogen and rFVIIa, alone or in combination, on whole blood coagulation thromboelastometry using pre- and postoperative blood samples from 18 consecutive adult patients undergoing CPB surgery. Dynamic thromboelastometric clotting profiles were recorded using citrated whole blood activated with trace amounts of tissue factor (Innovin, final dilution 1:17000). Blood samples were collected before surgery (control) and postoperative samples were obtained following in vivo neutralization of heparin with protamine sulphate. All blood samples were treated with heparinase to ensure neutralization of possible residual heparin effect. The post-operative blood samples were spiked with buffer, rFVIIa (2 microg/mL), fibrinogen (1 mg/mL), or the combination of rFVIIa and fibrinogen. Despite neutralization of heparin, CPB surgery left a measurable coagulopathy that was thromboelastometrically characterized by prolonged onset of clotting, reduced maximum velocity of clot formation (MaxVel), and decreased maximum clot firmness (MCF). Ex vivo spiking of the postoperative samples with rFVIIa shortened the clotting time. Fibrinogen also shortened the clotting time and, in addition, improved the MaxVel, and MCF. Finally, adding the combination of rFVIIa and fibrinogen to the postoperative samples corrected all thromboelastometric parameters to the preoperative range. In conclusion, the correction of whole blood clotting abnormalities that occurs with rFVIIa and/or fibrinogen suggests that future clinical trials on treatment of bleeding during CPB surgery should study the haemostatic effect of fibrinogen or possibly the combination of rFVIIa and fibrinogen.

Risk of excessive bleeding associated with marginally low von Willebrand factor and mild platelet dysfunction

Summary.  Background: Bleeding symptoms are so commonly reported that it is not known whether they associate causally or coincidentally with mild but measurable primary hemostatic defects. Objectives/patients/methods: In order to evaluate if the mild primary hemostatic defects are truly causative of increased bleeding symptoms, we surveyed a population of healthy teenagers for bleeding symptoms. Using a case–control approach, we then estimated the risk of excessive bleeding associated with low von Willebrand factor (defined as VWF below the 5th percentile of a normal reference population), and with mild platelet dysfunction [PD, defined as concurrent reduced platelet aggregation responses to two agonists (adenosine diphosphate and epinephrine)]. Results: Excessive bleeding was present in 63 out of 809 teenagers (7.8%). Among the 49 cases who were tested for VWF, low values by three measures were more commonly present than in 166 controls, specifically, ristocetin cofactor (RCo) activity [20.4% vs. 5.4%, odds ratio (OR) 4.5], collagen binding (14.3% vs. 4.2%, OR 3.8), and antigen level (20.4% vs. 6.0%, OR 4.0). The low RCo values ranged from 35 to 45 U dL−1 except for a single case with 26 U dL−1. Of the 47 teenagers with excessive bleeding who underwent platelet aggregation studies, reduced responses were more common than in controls (12.8% vs. 4.4%, OR 3.2). Twenty‐nine per cent of cases with excessive bleeding had either low RCo or PD. Conclusion: Almost one in three teenagers who report excessive bleeding is likely to have a measurable hemostatic disturbance manifested either by marginally low VWF (by three measures) or by mild PD.

Critical role of factors II and X during coumarin anticoagulation and their combined measurement with a new Fiix-prothrombin time

Vitamin K antagonists (VKA) are monitored with prothrombin time (PT) based assays that are equally sensitive to reductions in factors II, VII or X. We compared the effect of vitamin K dependent (VKD) coagulation factors on PT and also on rotational thromboelastometric (ROTEM) parameters. The PT was equally sensitive to reductions in factors II, VII or X but ROTEM parameters correlated poorly with the PT in anticoagulated patients´ plasmas. ROTEM parameters were more affected by mild and moderate reductions in FII or FX than by FVII or FIX which had little influence except at very low coagulant activity. We developed a modified PT that was sensitive only to reductions in factors II and X. The Fiix-PT (Fiix-INR) correlated well with PT (INR) but the Fiix-INR fluctuated less than the INR in an anticoagulated patient reflecting its insensitivity to FVII. The ROTEM results suggest that mild to moderate reductions in factors II or X are more important during clot formation than factors VII or IX. Reductions in FII and X may better reflect anticoagulation with VKA than FVII or IX. The new Fiix-PT may more accurately reflect the degree of therapeutic anticoagulation in patients treated with VKA than the current PT which is subject to a confounding variation caused by FVII.

Fiix-prothrombin time versus standard prothrombin time for monitoring of warfarin anticoagulation: a single centre, double-blind, randomised, non-inferiority trial

BACKGROUND Rapid fluctuations in factor VII during warfarin anticoagulation change the international normalised ratio (INR) but contribute little to the antithrombotic effect. We aimed to assess non-inferiority of anticoagulation stabilisation with a warfarin monitoring method affected only by factors II and X (Fiix-prothrombin time [Fiix-PT]) compared with standard PT-INR monitoring that includes factor VII measurement as well. METHODS The Fiix trial was a single centre, double-blind, prospective, non-inferiority, randomised controlled clinical trial. Ambulatory adults on warfarin with an INR target of 2-3 managed by an anticoagulation dosing service using software-assisted dosing at the National University Hospital of Iceland, Reykjavik, Iceland, were eligible for inclusion in this study. We excluded patients undergoing electroconversion and nursing home residents. Patients were randomly assigned (1:1) to either the Fiix-PT monitoring group or the PT monitoring group by block randomisation. A blinded research INR (R-INR) based on results of the respective test was reported to the dosing staff. Participants were contacted by a study nurse at 4-week intervals to elicit information about thromboembolism or bleeding otherwise unknown to the anticoagulation management centre. The primary efficacy outcome was a composite of objectively diagnosed non-fatal and fatal arterial or venous thromboembolism, including myocardial infarction and transient ischaemic attacks, assessed in all eligible patients who were randomised (intention-to-monitor population). The safety endpoint was major bleeding or other clinically relevant bleeding, assessed in the per-protocol population. We assumed a 3% annual thromboembolism incidence and a non-inferiority margin of 2·5%. This trial is registered with ClinicalTrials.gov, number NCT01565239. FINDINGS Between March 1, 2012, and Feb 28, 2014, we enrolled 1156 patients. 573 patients were assigned to Fiix-PT and 575 to PT-INR monitoring after exclusion of four patients from each group for various reasons. Median follow-up was 1·7 years (IQR 1·1-1·9). During days 1-720, ten (1·2% per patient year) thromboembolic events occurred in the Fiix-PT group versus 19 (2·3% per patient year) in the PT group (relative risk [RR] 0·52, 95% CI 0·25-1·13; pnon-inferiority<0·0001). Major bleeding occurred in 17 of 571 patients in the Fiix group (2·2% per patient year) versus 20 of 573 patients in the PT group (2·5% per patient year; RR 0·85, 0·45-1·61; pnon-inferiority=0·0034). Anticoagulation stability was improved with Fiix-PT monitoring as manifested by fewer tests, fewer dose adjustments, increased time in range and less INR variability than reported with standard PT monitoring. INTERPRETATION Monitoring of warfarin with Fiix-PT improved anticoagulation and dosing stability and was clinically non-inferior to PT monitoring. Results from this trial suggest that during vitamin K antagonist treatment INR monitoring could be replaced by Fiix-PT and that this would lead to at least a non-inferior clinical outcome compared with monitoring with PT-INR. FUNDING Innovation Center Iceland, University of Iceland Science Fund, Landspitali Science Fund and Actavis.

Quantification of menstrual flow by weighing protective pads in women with normal, decreased or increased menstruation

Objective. To investigate the association between menstrual flow weight measured from modern sanitary pads (converting liquid to non‐evaporating gel) and clinically assessed normal, increased or decreased menstrual flow. Design. Objective method development study. Setting. Outpatient clinic, University Hospital, Reykjavik. Population. One hundred and thirteen volunteers included 26 normally menstruating adult women and 52 normally menstruating teenagers not using oral or intrauterine contraception, seven normally menstruating women using oral contraception, 17 women with clinically diagnosed menorrhagia, five women using oral contraception for clinical menorrhagia, and six teenage girls claiming heavy menstrual flow. Methods. Menstruation length, menstrual flow weight and history of iron deficiency were assessed. During the menstruation following recruitment, all women collected their used protective pads in a hygienic manner and returned them to the laboratory for accurate weighing. Main outcome measures. Menstrual flow total weight measured in grams. Results. Mean menstrual flow total weight in the 78 asymptomatic women was 51 g (median 44, range 5–144). The mean flow in 17 women clinically diagnosed with menorrhagia was 217 g (median 207, range 63 – 402) (p<0.0001 compared to healthy women). The seven healthy women using oral contraceptives discharged 13 g (13–19) (p = 0.0004 compared with normals). Menstruation lasted

Clinical phenotype in heterozygote and biallelic Bernard‐Soulier syndrome—A case control study

Bernard‐Soulier syndrome (BSS) is a rare severe autosomal recessive bleeding disorder. To date heterozygous carriers of BSS mutations have not been shown to have bleeding symptoms. We assessed bleeding using a semi‐quantitative questionnaire, platelet parameters, PFA‐100 closure times, ristocetin response, GP Ib/IX expression and VWF antigen in 14 BSS patients, 30 heterozygote carriers for related mutations and 29 controls. Eight mutations in GP1BA, GP1BB or GP9 were identified including four previously unknown pathogenic mutations. Subjects with BSS reported markedly more mucocutaneous bleeding than controls. Increased bleeding was also observed in heterozygotes. Compared to controls, patients with BSS had lower optical platelet counts (P < 0.001), CD61‐platelet counts (P < 0.001) and higher mean platelet volume (17.7 vs. 7.8 fL, P < 0.001) and ristocetin response and closure times were unmeasurable. Heterozygotes had higher MPV (9.7 fL, P < 0.001) and lower platelet counts (P < 0.001) than controls but response to ristocetin and closure times were normal. The VWF was elevated in both BSS and in heterozygotes (P = 0.005). We conclude that heterozygotes for BSS mutations have lower platelet counts than controls and show a bleeding phenotype albeit much milder than in BSS. Both patients with BSS and heterozygote carriers of pathogenic mutations have raised VWF. Am. J. Hematol. 90:149–155, 2015.

Screening for anemia in patients on warfarin facilitates diagnosis of gastrointestinal malignancies and pre-malignant lesions

INTRODUCTION The prevalence and etiology of occult bleeding among patients on warfarin who are screened systematically for new anemia is largely unknown. We aimed to estimate the usefulness of following hemoglobin and mean red cell volume (MCV) with INR in order to screen for developing anemia as an indicator of occult bleeding. MATERIAL AND METHODS All patients on warfarin controlled at our institution had measurements of complete blood count (CBC) with INR during 18 months. Patients who fell>25 g/L and/or decrease of MCV over 5 fL or MCV<80 fL were contacted with instructions to undergo evaluation of anemia. RESULTS Overall 3218 patients on warfarin were monitored at our institution and 442 (13.7%) had anemia and 235 (7.3%) had unexplained anemia. A total of 163/235 (69%) who were notified contacted their doctors and 82/163 (50%) were referred for investigation with upper and/or lower endoscopies. Gastrointestinal malignancies were found in 11 patients (10 colorectal cancers, 1 esophageal) and pre-cancerous lesions among 14 other patients. Additional 25/82 patients (30%) had upper and/or lower bleeding lesions such as ulcers and angiodysplasia. Based on 3669 years of observation, 73 patients needed to be screened for one year in order to identify one gastrointestinal lesion causing occult bleeding. CONCLUSIONS Thirty percent of those endoscoped had malignant or pre-malignant diseases. Regular measurement of CBC concomitantly with INR in patients on warfarin therapy led to detection of otherwise asymptomatic diseases in a significant proportion of patients and might lead to earlier diagnosis of malignant and premalignant disease.

A single test to assay warfarin, dabigatran, rivaroxaban, apixaban, unfractionated heparin and enoxaparin in plasma

Essentials Simple and fast assaying of different anticoagulants (ACs) is useful in emergent situations. We used highly diluted prothrombin time (dPT) or highly diluted Fiix‐PT (dFiix‐PT) to assay ACs. Both tests could quantify target specific anticoagulants and warfarin anticoagulation. Improved results were consistently observed with the dFiix‐PT compared with the dPT.

Fiix-prothrombin time monitoring improves warfarin anticoagulation outcome in atrial fibrillation: a systematic review of randomized trials comparing Fiix-warfarin or direct oral anticoagulants to standard PT-warfarin

Monitoring warfarin with Fiix‐prothrombin time (Fiix‐PT), which is only affected by coagulation factors II and X, stabilizes anticoagulation and reduces thromboembolism compared to PT/INR monitoring. We compared outcome in nonvalvular atrial fibrillation (NVAF) patients treated with Fiix‐warfarin, direct oral anticoagulants (DOACs), or PT‐warfarin.

Complementary effect of fibrinogen and rFVIIa on clotting ex vivo in Bernard-Soulier syndrome and combined use during three deliveries

Abstract Women with Bernard-Soulier syndrome (BSS) are considered to be at high risk of serious bleeding during childbirth. Due to the frequently occurring platelet transfusion refractoriness, alternative prophylactic therapy is required. Using rotational thromboelastometry, we evaluated the whole blood coagulation profile of a pregnant woman with BSS before and after spiking ex vivo with different concentrations of recombinant activated factor VII (rFVIIa) and fibrinogen. As experiments suggested improved clotting with clinically applicable concentrations of both agents in a complementary manner, the findings were confirmed on blood from a non-pregnant woman and three men suffering from BSS. During delivery, bleeding refractory to platelets occurred and immediately following delivery she received both rFVIIa and fibrinogen intravenously. Immediate cessation of bleeding occurred, and no postpartum hemorrhage was seen. Another woman with BSS later also received the same rFVIIa and fibrinogen treatment prophylactically after delivery without any postpartum bleeding. Eventually, the first woman during her second delivery received the same treatment again prophylactically without any bleeding. No side effects were observed during these three deliveries. Our observations suggest that rFVIIa combined with fibrinogen may provide a beneficial clinical hemostatic effect partly by separate but complementary mechanisms.