Factor XI replacement for inherited factor XI deficiency in routine clinical practice: results of the HEMOLEVEN prospective 3-year postmarketing study.

Factor XI (FXI)-deficient patients may develop excessive bleeding after trauma or surgery. Replacement therapy should be considered in high-risk situations, especially when FXI levels are below 20 IU dL(-1) . HEMOLEVEN is a human plasma-derived factor XI concentrate available in France since 1992, but there are few data regarding its use by physicians. This prospective study assessed the use, efficacy and safety of HEMOLEVEN in common clinical practice. HEMOLEVEN was evaluated in FXI-deficient patients in 13 French centres in a 3-year postmarketing study. Forty-four patients (30 females, 14 males) received 67 treatments. The median age was 37 years (8 months-91 years). Basal FXI levels were <1 to 51 IU dL(-1) (median: 5.5); 29 patients were severely FXI-deficient (<20 IU dL(-1) ). FXI was administered prophylactically before 43 surgical procedures, 10 invasive procedures, 8 vaginal deliveries, or as curative treatment for six bleeds. The efficacy was assessed as excellent/good in 63, moderate in two and undetermined in two treatments. Seven patients experienced seven adverse effects, including two rated as serious: one sudden massive pulmonary embolism with fatal outcome and one case of inhibitor to FXI. HEMOLEVEN is effective for bleeding prevention in FXI deficiency. However, considering the benefit/risk ratio observed in relation to dosage in this study; firstly, it should be used sparingly due to its potential prothrombotic effect; secondly, new prescription procedures should be defined to adapt the dosage, especially in patients with intrinsic and/or acquired risk factors for thrombosis.

Perioperative management and neuraxial analgesia in women with factor XI deficiency (<60 IU/dL): a French multicenter observational study of 314 pregnancies.

Background: Factor (F)XI deficiency is a rare bleeding disorder with a poor correlation between bleeding tendency and FXI level. Management of pregnant women with FXI deficiency is not clearly established, especially regarding neuraxial analgesia (NA). Objectives: A retrospective multicenter observational study was conducted in French hemostasis centers on pregnant women with FXI of <60 IU/dL. Methods: Data to report were (i) FXI levels before pregnancy and at time of delivery, (ii) type of NA and delivery management modalities, and (iii) possible complications related to NA and bleeding complications. Results: Three hundred fourteen pregnancies in patients with FXI deficiency of <60 IU/dL were reported (from 20 centers); among them, 199 NA procedures have been completed (137 epidurals and 61 spinals, 1 had both). The period of childbirth was mostly from 2014 to 2020 (281/314; 89.5%). Congenital FXI deficiency was established with certainty by investigators in 32.8% patients (n = 103). Previous bleedings were described in 20.4% of the patients (64/314; 45.3% cutaneous, 31.3% gynecologic, and 15.6% postsurgical). Thirteen deliveries had an NA procedure with FXI of <30 IU/dL, 42 with FXI of 30-40 IU/dL, and 118 with FXI of 40-60 IU/dL. Median FXI levels at delivery in the epidural and spinal groups were not significantly different but were significantly lower in the group without NA by medical staff contraindications. There were no complications related to NA. A 17.5% postpartum hemorrhage or excessive postpartum bleeding incidence was reported, which is consistent with previous data. Conclusion: Our data support the use of a 30 IU/dL FXI threshold for NA, as suggested by the French proposals published in August 2023.

[Appropriate laboratory investigation in women with menorrhagia]. / Bonne pratique et valeur diagnostique de la biologie : hémostase-hématologie.

Inherited bleeding disorders are potentially causes of menorrhagia and must be investigated if no specific cause is identified. The reported prevalence of inherited bleeding disorders is high in women with menorrhagia compared to the general population. The most frequent disorders reported are von Willebrand's disease and Factor XI deficiency. Menorrhagia is, also, a frequent finding in women with congenital bleeding disorders. Morever, menorrhagia represents the major cause of iron-deficiency anemia among women of reproductive age. Primary evaluation for an underlying disorder of hemostasis in a woman wih menorrhagia is a focused history for familiy and personal history of bleeding symptoms and a complete blood cell count. This will rule out thrombocytopenic bleeding and also assesses for the degree, if any, of anemia. Those women with a positive screen and normal platelet count should be evaluated with laboratory investigation including prothrombin time, activated partial thromboplastin time, factor VIII, VWF ristocetin cofactor and antigen. If initial hemostasis testing above is normal, then further hemostasis testing can be considered, especially in terms of platelet aggregation, in a multidisciplinary clinic.

OC-06 - Pro-thrombotic biomarkers in pancreatic diseases: are they specific of cancer?

INTRODUCTION: Venous thrombo-embolic events (VTE) occur frequently in patients with pancreatic cancer and contribute to elevated morbidity and mortality. Clinical risk factors for thrombosis such as cancer stage and tumor grade have been clearly identified. Recently, several biomarkers have been proposed which may help identifying cancer patients at high risk of thrombosis. Those biomarkers have been studied in heterogeneous cohorts of patients with different cancer types. AIM: To compare pro-thrombotic biomarkers in pancreatic cancer and in chronic pancreatitis to determine whether these biomarkers are related to cancer or inflammation and to validate their association with thrombotic risk in a pancreatic cancer population. MATERIALS AND METHODS: 45 patients with pancreatic cancer, 49 with intraductal papillary mucinous tumor of the pancreas (IPMN), a precancerous lesion, and 50 with chronic pancreatitis were recruited. Plasma levels of factor VIII, D-dimers, thrombin-antithrombin complexes, soluble p-selectin, tissue factor-dependent procoagulant activity of MP (TF-MP), free Tissue Factor Pathway Inhibitor (TFPI) and extracellular DNA were measured. Thrombin generation triggered by 1pM of TF was evaluated with the Calibrated Automated Thrombogram assay. RESULTS: Levels of factor VIII, D-dimers, TF-MP, TFPI and extracellular DNA were significantly higher in cancer patients compared to IPMN or chronic pancreatitis (Table 1). Patients with metastatic cancer (n=27) presented higher levels of D-dimers (mean±sd 1.77±1.28 vs 0.80±0.96 µg/ml, p=0.004) and MP-TF (54.3±53.2 vs 15.8±10.4 fM, p=0.02) compared to patients with localized lesions (n=18). Among cancer patients, 42 were followed for a median duration of 187 days (min 21-max 802 days). VTE occurred in 10 (23%) patients. All had metastatic cancer at the time of thrombosis. Only D-dimers were significantly elevated in cancer patients with VTE compared to patients without VTE (median 1.85 vs 0.7 µg/ml, p=0.02). CONCLUSIONS: Elevation of factor VIII, D-dimers, TF-MP, TFPI and extracellular DNA seems to be related to cancer process, not to local or systemic inflammation as these parameters differentiate cancer from chronic pancreatitis. Interestingly, D-dimers and TF-MP are related to the disseminated cancer stage, suggesting that vascular invasion is a prerequisite to the release of TF-MP from the primary tumor into the bloodstream and to coagulation activation. However, only D-dimers are associated with the occurrence of future VTE. We propose that D-dimers could be a useful tool to predict thrombotic events in pancreatic cancer patients. This should be confirmed in a larger population.

Mechanism of factor IXa inhibition by antithrombin in the presence of unfractionated and low molecular weight heparins and fucoidan.

Heparin exerts its anticoagulant activity by catalysing the inhibition of coagulation proteases by antithrombin (AT). Its main target is thrombin but it also catalyses the inhibition of the other serine-proteases of the coagulation cascade, such as factor IXa (fIXa). The aim of this study was to compare the catalysis of inhibition of blood fIXa by antithrombin in the presence of several sulfated polysaccharides with anticoagulant activity, i.e. heparin, three widely used in therapeutics low molecular weight heparins (LMWH) and fucoidan. Plots of the second-order rate constants of the fIXa-antithrombin reaction vs. the concentration of added heparin and LMWH are bell-shaped and fit the kinetic model established for thrombin-antithrombin reaction by Jordan R., Beeler D., Rosenberg R. (1979) J. Biol. Chem., 254, 2902-2913. In the ascending branch, the catalyst (C) binds quickly to the inhibitor (I) to form a catalyst-inhibitor (CI) complex which is more reactive towards the enzyme (E) than the free inhibitor, leading to the formation of an inactive enzyme-inhibitor complex (EI) and the release of free catalyst, in a rate-limiting second step. After a maximum corresponding to an optimal catalyst concentration, the decrease in the reaction rate was in keeping with the formation of a catalyst-enzyme (CE) complex, whose inactivation by the CI complex was slower than that of the free enzyme. Maximum second-order rate constants for the inhibition of fIXa by AT were 105, 6.8, 12.24 and 22 microM-1 min-1 with heparin, Enoxaparin, Fraxiparin and Fragmin, respectively, leading to 3500-, 225-, 405- and 728-fold increases in the inhibition rate in the absence of polysaccharide, respectively. Fucoidan yielded 23-fold increase in the fIXa-antithrombin interaction rate. The kinetic profiles obtained with this polysaccharide exhibited ascending branch which correlated well with the kinetic model based on the formation of binary complexes (CI or CE). Fucoidan was covalently conjugated with a fluorescent probe (DTAF) and used in conjunction with fluorescence anisotropy to follow its binding to antithrombin, heparin cofactor II (HCII), thrombin and fIXa. The binding of fucoidan to these proteins occurred with low affinities when compared to heparin and LMWH. Fucoidan had higher affinity for the inhibitor HCII compared to antithrombin and enzymes. These data suggest that binding of heparins and fucoidan to the inhibitor (CI) is required for the polysaccharide-dependent enhancement in the rate of neutralization of the enzyme by the inhibitor.

Limited proteolysis of human alpha-thrombin by urokinase yields a non-clotting enzyme.

Limited proteolysis of human alpha-thrombin by various proteases has been efficiently used to demonstrate the importance of two insertion loops located on the surface of this molecule. In the present study, we demonstrate that two-chain urokinase (tcu-PA) specifically cleaves the B chain of alpha-thrombin giving rise to a transient derivative, consisting of two non-covalently linked subunits. Although the thrombin derivative conserves its activity towards the synthetic substrate S-2238 (Km = 8.4 microM and kcat = 145 s-1 versus respectively 4.5 microM and 149 s-1 for alpha-thrombin), most of its coagulant activity is lost (140 NIH u/mg versus 3000 NIH u/mg) and its ability to activate platelets is considerably reduced (threshold for full platelet aggregation 2.5 nM versus 0.25 nM). The thrombin fragments were separated by HPLC and after reduction and S-carboxyamidemethylation were digested with a lysylendopeptidase; the resulting peptides were separated by HPLC and sequenced. One fragment corresponded to B chain fragment 1-73 and the second to B chain fragment 74-259 covalently linked to the A chain, indicating that tcu-PA cleaves selectively the peptide bond Arg 73-Asn 74 in the B chain. The proteolytic derivative obtained, designated beta u-thrombin, is therefore identical to the transient proteolytic derivative, beta 1-thrombin, produced by trypsin. Prolonged incubation with tcu-PA resulted in further conversion in a derivative analogous to gamma t-thrombin.(ABSTRACT TRUNCATED AT 250 WORDS)

Three new cases of dysfibrinogenemia: Poissy III, Saint-Germain I and Tahiti.

In order to identify unknown mutations, the FAMA method was used to rapidly screen the fibrinogen chain genes in individuals with dysfibrinogenemias. Chemical cleavage at mismatches on heteroduplexes DNA end-labeled with strand-specific fluorescent dyes reliably detects sequence changes in DNA fragments of up to 1.5 kb and locates them precisely. This method was successfully used for the detection of three new dysfibrinogenemias: Poissy III, Tahiti (heterozygous Aalpha Arg16His) and Saint-Germain I (heterozygous AalphaGly12Val). The mutations were confirmed by dideoxy sequencing.

Heparin-induced thrombocytopenia with thrombotic complications during prophylactic treatment with low-molecular-weight heparin.

Heparin-induced thrombocytopenia is very uncommon with low-molecular-weight heparin, especially when given for prophylaxis of venous thromboembolism. In two of the four published cases, with thrombotic complications, thrombocytopenia may have been related to cross-reactivity between unfractionated heparin and low-molecular-weight heparin. We report one patient who received low-molecular-weight heparin for prophylaxis against venous thromboembolism without any previous injection of unfractionated heparin, and experienced thrombocytopenia with thrombotic complications. Heparin-induced thrombocytopenia was confirmed by several laboratory assays. This observation emphasizes the need for platelet count monitoring during low-molecular-weight heparin therapy.

Diagnostic value of two rapid and individual D-dimer assays in patients with clinically suspected pulmonary embolism: comparison with microplate enzyme-linked immunosorbent assay.

A semiquantitative enzyme-linked immunosorbent assay (ELISA) test (Instant IA D-dimer) and a new quantitative rapid and individual test (STA-Liatest DDi) were compared with the reference microplate ELISA (Asserachrom D-Di) for D-dimer testing in 142 patients clinically suspected of pulmonary embolism, on the basis of clinical symptoms and signs, electrocardiogram, blood gases and chest X-Ray abnormalities. The cut-off value for the quantitative tests was 500 ng/ml and Instant IA was interpreted by three readers. Pulmonary embolism was confirmed by lung scan or angiography in 60 patients (42%). The sensitivities of ELISA and STA-Liatest DDi were 92% [95% confidence interval (CI) 82-97%] and 93% (95% CI 84-98%), respectively. The three readings of Instant IA D-dimer disagreed in 27 (19%) of the patients and sensitivity varied from 83 to 93% according to the readers. In the 115 patients with concordant readings, sensitivity was 92% (95% CI 82-98%). These results suggest that STA Liatest DDi may be used instead of microplate ELISA for the exclusion of pulmonary embolism, whereas the use of Instant IA D-dimer for this purpose is limited by the number of discordant results.

Markers of hemostatic system activation in pulmonary embolism. Changes during and after cessation of anticoagulant treatment.

Plasma levels of prothrombin fragment 1+2 (Fl+2), thrombin-antithrombin complexes (TAT) and D-dimers were measured in 15 patients with pulmonary embolism during heparin therapy, oral anticoagulation, and after cessation of warfarin therapy. Each patient had a favorable outcome during anticoagulant therapy (3 months), but late venous thromboembolism occurred in six cases. The mean levels of the three markers were significantly increased on day 4 after the thrombotic event, and normalized during warfarin therapy. Nine months after the initial pulmonary embolism, mean levels of the three markers, as compared with a control population, were significantly higher in the patients with late recurrences, whereas only TAT were slightly higher in patients without recurrences as compared with controls. Only TAT levels were significantly higher in the patients with late recurrences than in those without late recurrences. Thus, the levels of the three markers 9 months after pulmonary embolism seem to be interesting to identify patients with high risk of recurrence and who might require longer anticoagulant treatment.

Acquired factor VII deficiency associated with pleural liposarcoma.

Isolated acquired factor VII (FVII) deficiency (0.15 U/ml) was identified in a 30-year-old man with pleural liposarcoma. The patient underwent surgery with continuous FVII concentrate infusion. No anti-FVII antibody or FVII/anti-FVII complex was detected. However, the short half-life and low recovery of FVII after concentrate infusion suggested the presence of an antibody. Whatever the mechanism, this FVII deficiency was related to the presence of the liposarcoma. FVII level normalized during tumour regression and fell again when the liposarcoma relapsed.

Fibrinogen Saint-Germain I: a case of the heterozygous Aalpha GLY 12 --> VAL fibrinogen variant.

A fibrinogen variant was suspected based on the results of routine coagulation tests in a 2-year-old asymptomatic child. Coagulation studies showed marked prolongation of both the thrombin and reptilase times, and discrepancy was noted between the level of plasma fibrinogen as measured by a kinetic versus immunological determination. Family studies revealed that the father beared the same abnormality. Studies of purified fibrinogen revealed an impaired release of both fibrinopeptides by thrombin. Fibrin monomer polymerization and fibrin stabilization were normal. DNA sequencing revealed a heterozygous G --> T point mutation in exon 2 of the gene coding for the Aalpha chain, which substituted a Gly for Val at position 12. Although the mutation is the same as in fibrinogen Rouen, fibrinogen Saint-Germain I shows a different fibrinopeptide release pattern and a mild factor V deficiency.

Recurrent thromboembolism in a patient with beta-thalassemia major associated with double heterozygosity for factor V R506Q and prothrombin G20210A mutations.

Double heterozygosity for factor V R506Q and prothrombin G20210A mutations was identified in a 24-year-old man with beta-thalassemia major. The patient experienced a first thrombotic event at the age of 19 years and three recurrent thromboses in a short time interval, the third occurring while the patient was receiving long-term anticoagulant treatment. This case suggests that patients with major thalassemia and congenital thrombophilic mutations need intensive and long-lasting anticoagulant treatment. Thus, even if thrombotic events could be explained by a hypercoagulable state observed in patients with major thalassemia, after a first thrombotic event has occurred these patients should be screened for acquired and congenital thrombophilia.

[Anticoagulant clinics are they effective in France? Performance evaluation of six anticoagulant clinics concerning the management of vitamin K antagonists]. / Les cliniques d'anticoagulants sont-elles efficaces en France ? Évaluation des performances de six cliniques dans la gestion des traitements par antivitamines K.

PURPOSE: Recent data show that the quality of anticoagulation evaluated in patients receiving vitamin K antagonists (VKA) is not optimal in France. The aim of this retrospective study was to estimate the performances of six French anticoagulant clinics that manage VKA treatments over a 3-year period, from 2009 to 2011. METHODS: All clinics used the same rule based software. We determined the time spent in the therapeutic range (TTR), a surrogate end-point of quality of treatment with VKA. RESULTS: The overall duration of follow-up was 2755 patient-years concerning 2385 patients. The time spent in the therapeutic range 2 to 3 assigned for 89% of the patients, was 73%. On the other hand the time spent in the therapeutic range for the other two INR ranges (2.5-3.5 and 3-4.5) concerning 11% of patients with prosthetic heart valve was lower (63.7% and 68.8% respectively) with an imbalance in favour of the time below the range. In this study, warfarin (Coumadine(®)) and fluindione (Previscan(®)) allowed an equivalent quality of anticoagulation. The 1728 patients of age ranged from 60 to 100 years spent more time in TTR than the 651 younger patients. The percentage of time spent with an INR greater than 5 was extremely reduced which is a guarantee of safety. CONCLUSION: These results prove that anticoagulant clinics in France have the same good performances as their counterparts abroad. It can be assumed that a high TTR contributes to a low incidence of both bleedings and thrombosis.

Clinical practice guidelines on menorrhagia: management of abnormal uterine bleeding before menopause.

BACKGROUND: Normal menstrual periods last 3-6 days and involve blood loss of up to 80ml. Menorrhagia is defined as menstrual periods lasting more than 7 days and/or involving blood loss greater than 80ml. The prevalence of abnormal uterine bleeding (AUB) is estimated at 11-13% in the general population and increases with age, reaching 24% in those aged 36-40 years. INVESTIGATION: A blood count for red cells+platelets to test for anemia is recommended on a first-line basis for women consulting for AUB whose history and/or bleeding score justify it. A pregnancy test by an hCG assay should be ordered. A speculum examination and Pap smear, according to the French High Health Authority guidelines should be performed early on to rule out any cervical disease. Pelvic ultrasound, both abdominal (suprapubic) and transvaginal, is recommended as a first-line procedure for the etiological diagnosis of AUB. Hysteroscopy or hysterosonography can be suggested as a second-line procedure. MRI is not recommended as a first-line procedure. TREATMENT: In idiopathic AUB, the first-line treatment is medical, with efficacy ranked as follows: levonorgestrel IUD, tranexamic acid, oral contraceptives, either estrogens and progestins or synthetic progestins only, 21 days a month, or NSAIDs. When hormone treatment is contraindicated or immediate pregnancy is desired, tranexamic acid is indicated. Iron must be included for patients with iron-deficiency anemia. For women who do not wish to become pregnant in the future and who have idiopathic AUB, the long-term efficacy of conservative surgical treatment is greater than that of oral medical treatment. Placement of a levonorgestrel IUD (or administration of tranexamic acid by default) is recommended for women with idiopathic AUB. If this fails, a conservative surgical technique must be proposed; the choices include second-generation endometrial ablation techniques (thermal balloon, microwave, radiofrequency), or, if necessary, first-generation techniques (endometrectomy, roller-ball). A first-line hysterectomy is not recommended in this context. Should a hysterectomy be selected for functional bleeding, it should be performed by the vaginal or laparoscopic routes.