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Background: Cardiovascular implantable devices, such as vascular stents, are critical for the treatment of cardiovascular diseases. However, their success is dependent on robust and often long-term antithrombotic therapies. Yet, the current standard-of-care therapies often pose significant bleeding risks to patients. Coagulation factor (F)XI and FXII have emerged as potentially safe and efficacious targets to safely reduce pathologic thrombin generation in medical devices., Objectives: To study the efficacy of monoclonal antibody-targeting FXII and FXI of the contact pathway in preventing vascular device-related thrombosis., Methods: The effects of inhibition of FXII and FXI using function-blocking monoclonal antibodies were examined in a nonhuman primate model of nitinol stent-related thrombosis under arterial and venous flow conditions., Results: We found that function-blocking antibodies of FXII and FXI reduced markers of stent-induced thrombosis in vitro and ex vivo. However, FXI inhibition resulted in more effective mitigation of thrombosis markers under varied flow conditions., Conclusion: This work provides further support for the translation of contact pathway of coagulation inhibitors for their adjunctive clinical use with cardiovascular devices., Competing Interests: Declaration of competing interests M.W. and E.I.T. are employees of Aronora Inc, and they as well as Oregon Health & Science University may have a financial interest in the results of this study. This potential conflict of interest has been reviewed and managed by Oregon Health & Science University. The remaining authors declare no competing financial interests., (Copyright © 2024 International Society on Thrombosis and Haemostasis. Published by Elsevier Inc. All rights reserved.)

Objective- Factor XI (FXI) contributes to thrombotic disease while playing a limited role in normal hemostasis. We generated a unique, humanized anti-FXI antibody, AB023, which blocks factor XIIa-mediated FXI activation without inhibiting FXI activation by thrombin or the procoagulant function of FXIa. We sought to confirm the antithrombotic activity of AB023 in a baboon thrombosis model and to evaluate the safety, tolerability, pharmacokinetics, and pharmacodynamics in healthy adult subjects. Approach and Results- In a primate model of acute vascular graft thrombosis, AB023 reduced platelet and fibrin accumulation within the grafts by >75%. To evaluate the safety of AB023, we performed a first-in-human study in healthy adult volunteers without any serious adverse events. Overall, 10 of 21 (48%) subjects experienced 20 treatment-emergent adverse events, with 7 of 16 (44%) subjects following active treatment and 3 of 5 (60%) subjects following placebo. AB023 did not increase bleeding or prothrombin times. Anticoagulation was verified by a saturable ≈2-fold prolongation of the partial thromboplastin time for over 1 month after the highest dose. Conclusions- AB023, which inhibits contact activation-initiated blood coagulation in vitro and experimental thrombus formation in primates, produced a dose-dependent duration of limited anticoagulation without drug-related adverse effects in a phase 1 trial. When put in context with earlier observations suggesting that FXI contributes to venous thromboembolism and cardiovascular disease, although contributing minimally to hemostasis, our data further justify clinical evaluation of AB023 in conditions where contact-initiated FXI activation is suspected to have a pathogenic role. Clinical Trial Registration- URL: http://www.clinicaltrials.gov . Unique identifier: NCT03097341.

Staphylococcus aureus infections can produce systemic bacteremia and inflammation in humans, which may progress to severe sepsis or septic shock, even with appropriate antibiotic treatment. Sepsis may be associated with disseminated intravascular coagulation and consumptive coagulopathy. In some types of mouse infection models, the plasma coagulation protein factor XI (FXI) contributes to the pathogenesis of sepsis. We hypothesize that FXI also contributes to the pathogenesis of sepsis in primates, and that pharmacological interference with FXI will alter the outcome of Staphylococcus aureus -induced lethality in a baboon model. Pretreatment of baboons with the anti-FXI antibody 3G3, a humanized variant of the murine monoclonal 14E11 that blocks FXI activation by FXIIa, substantially reduced the activation of coagulation, as reflected by clotting times and plasma complexes of coagulation proteases (FXIIa, FXIa, FIXa, FXa, FVIIa, and thrombin) with serpins (antithrombin or C1 inhibitor) following infusion of heat-inactivated S aureus 3G3 treatment reduced fibrinogen and platelet consumption, fibrin deposition in tissues, neutrophil activation and accumulation in tissues, cytokine production, kininogen cleavage, cell death, and complement activation. Overall, 3G3 infusion protected the structure and function of multiple vital organs, including lung, heart, liver, and kidney. All treated animals reached the end point survival (7 days), whereas all nontreated animals developed terminal organ failure within 28 hours. We conclude that FXI plays a role in the pathogenesis of S aureus -induced disseminated intravascular coagulation and lethality in baboons. The results provide proof of concept for future therapeutic interventions that may prevent sepsis-induced organ failure and save lives in certain forms of sepsis., (© 2019 by The American Society of Hematology.)

The name human contact system is related to its mode of action, as "contact" with artificial negatively charged surfaces triggers its activation. Today, it is generally believed that the contact system is an inflammatory response mechanism not only against artificial material but also against misfolded proteins and foreign organisms. Upon activation, the contact system is involved in at least two distinct (patho)physiologic processes: i . the trigger of the intrinsic coagulation via factor XI and ii . the cleavage of high molecular weight kininogen with release of bradykinin and antimicrobial peptides (AMPs). Bradykinin is involved in the regulation of inflammatory processes, vascular permeability, and blood pressure. Due to the release of AMPs, the contact system is regarded as a branch of the innate immune defense against microorganisms. There is an increasing list of pathogens that interact with contact factors, in addition to bacteria also fungi and viruses bind and activate the system. In spite of that, pathogens have developed their own mechanisms to activate the contact system, resulting in manipulation of this host immune response. In this up-to-date review, we summarize present research on the interaction of pathogens with the human contact system, focusing particularly on bacterial and viral mechanisms that trigger inflammation via contact system activation.

Emerging evidence from animal models suggests that platelets may participate in a wide variety of processes including the immune response against infection. More than 200 whole blood samples from patients and healthy controls were run in the System XE-5000 analyzer, and plasma fractions were separated for the following tests by ELISA, Luminex and light scattering. We describe two mechanisms by which platelets may contribute to immune function against various bacterial pathogens based on increased mean platelet volume in gram-positive bacterial infections and increased platelet counts in gram-negative bacterial infections. Gram-negative bacteria activate platelets to recruit neutrophils, which participate in the immune response against infection. During this process, fractalkine, macrophage inflammatory protein-1β, interleukin-17A, tumor necrosis factor-α and platelet-activating factor were higher in patients infected with Escherichia coli; additionally, giant platelets were observed under the microscope. Meanwhile, we found that platelets played a different role in gram-positive bacterial infections. Specifically, they could actively adhere to gram-positive bacteria in circulation and transfer them to immune sites to promote antibacterial lymphocyte expansion. During this process, complement C3 and factor XI were more highly expressed in patients infected with Staphylococcus aureus; additionally, we detected more small platelets under the microscope. Platelets participate in the immune response against both gram-negative and gram-positive bacteria, although the mechanisms differ. These results will help us understand the complex roles of platelets during infections, and direct our use of antibiotics based on clinical platelet data.

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., (© 2015 John Wiley & Sons Ltd.)

Hemophilia is characterized by genetic mutations resulting in the deficiency of factors critical to the normal process of coagulation, sometimes resulting in spontaneous bleeding into soft tissue, joints, and internal organs. The 2 most common subtypes are hemophilia A, or factor VIII deficiency, and hemophilia B, or factor IX deficiency. Hemophilia affects an estimated 20,000 individuals in the United States. The diagnosis and management of patients with severe hemophilia is complex, and requires preventive treatment (prophylaxis) to avoid bleeding episodes and related complications and the use of replacement therapy with coagulation factors during acute bleeding episodes. To achieve optimal long-term results, the treatment of patients with hemophilia requires a comprehensive approach coordinated by a multidisciplinary team of specialists. Hemophilia imposes a substantial burden from economic, societal, and patient perspectives.

Background: High levels of activated protein–inhibitor complexes of the intrinsic coagulation proteins are associated with ischemic stroke (IS) but not with myocardial infarction (MI). This study was aimed at determining whether the antigen levels of coagulation factors(factor XII, FXII, and FXI and prekallikrein (PK)are associated with MI and IS, and whether this association is independent of levels of activated protein–inhibitor complexes., Patients and Methods: The RATIO study included young women (< 50 years) with MI (N = 205)and IS (N = 175), and 638 healthy controls. Antigen levels of FXII, FXI and PK were measured and expressed as percentages of of those in pooled normal plasmas. Odds ratios (ORs) and corresponding 99% confidence intervals (CIs) were calculated for high levels (i.e. ≥ 90th percentile of controls) as measures of rate ratios., Results: After adjustment for potential confounders, high levels of FXII antigen were not associated with MI risk or IS risk(OR(MI) 1.18, 99% CI 0.51–2.74; ORIS 1.03, 9% CI 0.41–2.55). High levels of FXI antigen were slightly associated with an increase in MI risk (OR(MI) 1.55, 9% CI 0.74–3.21), whereas there was a substantial association with IS risk (ORIS 2.65, 9% CI 1.27–5.56). PK antigen was slightly associated with MI risk but not with IS risk(ORMI 1.54, 9% CI 0.67–3.52; ORIS 0.90, 9% CI 0.35–2.33). All associations remained similar after adjustment for levels of protein–inhibitor complexes., Conclusion: Increased levels of FXI antigen were associated with an increase in IS risk, whereas they showed only a marginal association with MI risk. FXII antigen and PK antigen levels were not substantially associated with MI risk and IS risk.

Coagulation factor XI (FXI) is a promising target for anticoagulation, because of its major role in thrombosis and relatively minor role in haemostasis. This implies that inhibition of FXI can prevent thrombosis without causing bleeding. It was our aim to investigate the antithrombotic properties of two novel inhibitory anti-human FXI antibodies (αFXI-175 and αFXI-203). The in vitro properties of both antibodies were analysed using standard clotting assays and calibrated automated thrombography. For the in vivo model we used FXI knockout mice, in which FXI plasma levels were restored with purified human FXI. Thrombosis was induced by applying ferric chloride to the vena cava inferior, after which time to occlusion was analysed. A tail bleeding assay was used to investigate the safety of both antibodies. Using calibrated automated thrombography, both antibodies inhibited thrombin generation initiated via the intrinsic pathway. In contrast, upon tissue factor (TF)-initiated thrombin generation, αFXI-203 did not inhibit thrombin generation, while αFXI-175 inhibited thrombin generation only at low concentrations of TF. In the murine thrombosis model, the vena cava inferior remained patent for 25 minutes (min) in mice treated with αFXI-175 and for 12.5 min in αFXI-203 treated animals, which was significantly longer than in placebo-treated animals (5 min, p<0.05). Neither antibody caused severe blood loss in a tail bleeding assay. In conclusion, the two inhibitory antibodies against FXI prevented cessation of blood flow in a murine thrombosis model without inducing a bleeding tendency.

Purpose: To assess the risks of intraoperative and postoperative bleeding associated with cataract extraction without prophylactic treatment in patients with severe factor XI (FXI) deficiency., Design: Prospective interventional case series., Setting: Single institute., Study Population: Consecutive unrelated patients with severe FXI deficiency who underwent cataract extraction under topical anesthesia, with a clear corneal incision, phacoemulsification, and implantation of a foldable posterior chamber intraocular lens (PCIOL) were enrolled. Patients with associated intraocular conditions that could complicate the surgery were excluded., Intervention: Cataract extraction without prophylactic treatment for the FXI deficiency., Main Outcome Measures: Assessment of intraoperative and postoperative ocular bleeding and other related complications., Results: Seven patients ranging in age from 61 to 95 years (median, 79) underwent phacoemulsification and PCIOL implantation in 11 eyes. Five patients (71%) were homozygotes for type II mutation of the FXI gene (activity level of <1 U/dl), 1 patient was a homozygote for type III mutation (activity level of 11 U/dl), and 1 patient was a compound heterozygote for types II and III (activity level of 3 U/dl). Three of the patients (43%), all type II homozygotes, also had an inhibitor antibody to FXI. All 7 patients were followed for at least 1 week after the operation. The surgery was uneventful in all eyes, and neither major nor minor bleeding events were observed in any of the operated eyes during surgery and follow-up., Conclusions: Cataract extraction by phacoemulsification in uncomplicated eyes can be performed safely without prophylactic treatment in patients with severe FXI deficiency with or without inhibitor antibodies against FXI.

Objectives: Rare patients with systemic lupus erythematosus (SLE) patients exhibit anticoagulants that interfere in the earlier stages of the intrinsic coagulation pathway, such as those involving factor XI (FXI). The objectives of our study were to describe the presence of an acquired inhibitor to FXI causing a life-threatening bleeding disorder in an SLE patient and to review the association of this coagulopathy with SLE., Methods: We describe the clinical presentation of an SLE patient with an acquired FXI inhibitor. We reviewed the scientific literature using the MEDLINE database searching the following combinations of terms: "SLE and Factor XI," "SLE and Factor XI inhibitor," and "Factor XI inhibitor," from 1964 to 2007., Results: A 20-year-old woman with a 6-year history of SLE was admitted to the hospital because of severe life-threatening abdominal bleeding due to a ruptured ovarian cyst. This hemorrhagic event was related to the presence of an FXI inhibitor. We reviewed another 13 SLE patients with this condition, 8 of whom had bleeding events. Most patients had manifestations of active SLE, and prednisone was used as the primary treatment., Conclusions: SLE activity seems to be associated with the production of antibodies directed against FXI, which may cause important coagulopathies, especially bleeding events. The inhibitor disappeared after immunosuppressive therapy for SLE in most cases, suggesting that the appearance of this inhibitor is immune mediated. Although the majority of cases with the FXI inhibitor are not fatal, it should be suspected and investigated in SLE patients, especially those with abnormal clotting tests.

The protease thrombin is required for normal hemostasis and pathologic thrombogenesis. Since the mechanism of coagulation factor XI (FXI)-dependent thrombus growth remains unclear, we investigated the contribution of FXI to thrombus formation in a primate thrombosis model. Pretreatment of baboons with a novel anti-human FXI monoclonal antibody (aXIMab; 2 mg/kg) inhibited plasma FXI by at least 99% for 10 days, and suppressed thrombin-antithrombin (TAT) complex and beta-thromboglobulin (betaTG) formation measured immediately downstream from thrombi forming within collagen-coated vascular grafts. FXI inhibition with aXIMab limited platelet and fibrin deposition in 4-mm diameter grafts without an apparent increase in D-dimer release from thrombi, and prevented the occlusion of 2-mm diameter grafts without affecting template bleeding times. In comparison, pretreatment with aspirin (32 mg/kg) prolonged bleeding times but failed to prevent graft occlusion, supporting the concept that FXI blockade may offer therapeutic advantages over other antithrombotic agents in terms of bleeding complications. In whole blood, aXIMab prevented fibrin formation in a collagen-coated flow chamber, independent of factor XII and factor VII. These data suggest that endogenous FXI contributes to arterial thrombus propagation through a striking amplification of thrombin generation at the thrombus luminal surface.

The management of factor XI deficiency is not straightforward for three reasons: firstly, the role of this factor in the coagulation pathway is not clearly understood; secondly, the bleeding tendency, although mild, is unpredictable and does not clearly relate to the factor XI level; and thirdly, all treatment products, although available, have some potentially serious side effects. These factors (or enigmas) contribute to the variable management of patients with this coagulation factor deficiency, but recent research is helping to clarify some of these areas.

Autoimmune phenomena, most frequently autoimmune hemolytic anemia, is a well-known complication of lymphoproliferative diseases. We report a very rare association of a chronic lymphocytic leukemia with an acquired factor XI inhibitor. A 87-year-old man presented with auto-immune hemolytic anemia. He had untreated chronic lymphocytic leukemia for the past three years and renal insufficiency. Before surgical procedure for arteriovenous fistula, we discovered a very prolonged activated partial thromboplastin time (APTT), and an acquired factor XI inhibitor was detected. The patient was successfully treated with immunosuppressive therapy. Among patients with lymphoproliferative disorders the discovery of a prolonged APTT implies to search for rare autoimmune phenomena like acquired coagulation factor inhibitors.

In this paper, we report an inhibitor antibody to factor XI (FXI) in a woman with severe inherited FXI deficiency, induced by FXI present in an Rh immune globulin preparation. The patient is homozygous for the Glu117Stop mutation, associated with a FXI level of less than 1 U/dL. Unlike all previously described patients with severe FXI deficiency and an inhibitor, the patient had never been exposed to blood products. Following 3 injections of Rh immune globulin during pregnancy, she developed an inhibitor to FXI (8 Bethesda units) that was shown to bind specifically to FXI and inhibit factor IX cleavage by purified FXIa. The administered Rh immune globulin and 2 other similar products were shown to contain FXI. Clinicians should be aware of the potential for immunization of severely FXI-deficient patients by FXI present in Rh immune globulin preparations.

The ADAMTS13 (a disintegrin and metalloprotease with a thrombospondin type I domain 13) related markers were measured in the plasma of healthy volunteers and thrombotic microangiopathy (TMA) patients including thrombotic thrombocytopenic purpura (TTP) to examine their efficacy in the diagnosis of TTP. The plasma levels of the ADAMTS13 antigen and ADAMTS13-factor XI complex were significantly lower in TMA patients with a significant decreased ADAMTS13 activity (and these patients were considered to have TTP) than in the healthy volunteers. The plasma levels of ADAMTS13 antigens closely correlated with those of ADAMTS13-factor XI complex. Autoantibody for ADAMTS 13 was also positive in almost all TTP patients. In addition, the ratio of von Willebrand factor (VWF)/ADAMTS13 activity was significantly high in TTP suggesting that this ratio might be more useful for the differential diagnosis of TTP than the ADAMTS13 assay alone. These findings suggest that ADAMTS13 related markers are useful for the diagnosis and analysis of TTP.

Our aim was to establish whether differences in clotting times for recalcified blood and plasma samples might be explained by the use of different blood collection tubes. Samples obtained from different plastic vacuum tubes were recalcified and clotting times determined by free oscillation rheometry. The clotting times for blood collected in Vacutainer (Becton Dickinson, Rutherford, New Jersey, USA) or Vacuette (Greiner Bio-One, Kremsmünster, Austria) tubes decreased with time, with maximal effect after 30 min. Blood from Monovette (Sarstedt, Nümbrecht, Germany) tubes displayed longer clotting times, which did not decrease with time. Clotting times for plasma prepared after 1 h storage in Vacutainer or Vacuette tubes were unaffected by subsequent addition of corn trypsin inhibitor to inhibit factor XIIa, although an antibody against factor XI prolonged the clotting time markedly. In Monovette plasma, both corn trypsin inhibitor and anti-factor XI effectively prolonged the clotting time. When corn trypsin inhibitor or anti-factor XI was added to the tubes before blood collection, but both additions clearly prolonged the clotting times in all types of tubes, even though corn trypsin inhibitor was less effective in whole blood. Antibodies against human tissue factor did not affect the clotting times. The amounts of platelet or leukocyte microparticles in plasma were low and similar in all tubes. This indicates that blood collection in Vacutainer or Vacuette tubes induces a rapid activation of factor XII and factor XI.

Thrombin, generated through activation of factor XI (FXI) and/or tissue factor (TF)-factor VIIa, is essential for thrombosis and hemostasis. We investigated the role of FXI-dependent thrombus propagation under arterial flow conditions producing rapid thrombus growth that, after the initiation phase, could limit the availability of TF at the blood/thrombus interface. Thrombosis was initiated by knitted dacron or TF-presenting teflon grafts deployed into arteriovenous shunts in baboons treated with antihuman FXI antibody (aFXI). Although aFXI did not prevent thrombus initiation, it markedly reduced intraluminal thrombus growth on both surfaces. The antithrombotic effect of aFXI was comparable with that of heparin at doses that significantly prolonged the partial thromboplastin time (APTT), prothrombin time (PT), and bleeding time (BT). aFXI also prolonged the APTT, but the PT and BT were unaffected. Thus, antithrombotic targeting of FXI might inhibit thrombosis with relatively modest hemostatic impairment versus strategies targeting other coagulation factors.

Factor XI deficiency, an injury-related bleeding disorder, is rare worldwide but common in Jews in whom 2 mutations, Glu117Stop (type II) and Phe283Leu (type III), prevail. Mean factor XI activities in homozygotes for Glu117Stop and for Phe283Leu are 1 and 10 U/dL, respectively. Inhibitors to factor XI in patients with severe factor XI deficiency have been reported in a small number of instances. This study was undertaken to determine the prevalence of acquired inhibitors against factor XI in patients with severe factor XI deficiency, discern whether these inhibitors are related to specific mutations, and characterize their activity. Clinical information was obtained from unrelated patients with severe factor XI deficiency, and blood was analyzed for factor XI activity, inhibitor to factor XI, and causative mutations. Immunoglobulin G purified from patients with an inhibitory activity was tested for binding to factor XI, effects on activation of factor XI by factor XIIa and thrombin, and activation of factor IX by exogenous factor XIa. Of 118 Israeli patients, 7 had an inhibitor; all belonged to a subgroup of 21 homozygotes for Glu117Stop who had a history of plasma replacement therapy. Three additional patients with inhibitors from the United Kingdom and the United States also had this genotype and were exposed to plasma. The inhibitors affected factor XI activation by thrombin or factor XIIa, and activation of factor IX by factor XIa. The results imply that patients with a very low factor XI level are susceptible to development of an inhibitor following plasma replacement.

Factor XI binds to high affinity sites on the surface of stimulated platelets where it is efficiently activated by thrombin. Here, we provide evidence that the factor XI binding site on platelets is in the glycoprotein (GP) Ibalpha subunit of the GP Ib-IX-V complex as follows. 1) Bernard-Soulier platelets, lacking the complex, are deficient in factor XI binding; 2) two GP Ibalpha ligands, SZ-2 (a monoclonal antibody) and bovine von Willebrand factor, inhibit factor XI binding to platelets; 3) by surface plasmon resonance, factor XI bound specifically to glycocalicin (the extracellular domain of GP Ibalpha) in Zn(2+)-dependent fashion (K(d)( app) approximately 52 nm). We then investigated whether glycocalicin could promote factor XI activation by thrombin, another GP Ibalpha ligand. In the presence of high molecular weight kininogen (45 nm), Zn(2+) and Ca(2+) ions, thrombin activated factor XI in the presence of glycocalicin at rates comparable with those seen in the presence of dextran sulfate (1 microg/ml). With higher high molecular weight kininogen concentrations (360 nm), the rate of thrombin-catalyzed factor XI activation in the presence of glycocalicin was comparable with that on activated platelets. Thus, factor XI binds to the GP Ib-IX-V complex, promoting its activation by thrombin.

Recent evidence shows that many antiphospholipid antibodies (aPL) to negatively-charged phospholipid (PL) do not target anionic PL per se, but are specific for anionic PL-binding plasma proteins, for example, beta(2)-glycoprotein I (beta(2)-GPI) and prothrombin. We also reported that certain antiphosphatidylethanolamine antibodies (aPE) are not specific for phosphatidylethanolamine (PE) per se, but are directed to PE-binding plasma proteins, high molecular weight kininogen (HK), and low molecular weight kininogen (LK). Additional studies have shown that certain aPE failed to recognize purified kininogens but continued to produce aPE ELISA reactivity in the presence of semipurified HK preparations containing the HK binding proteins, factor XI (FXI) and prekallikrein (PK). We therefore investigated if certain of these aPE recognized FXI and/or PK. In this study we observed that aPE can recognize contact proteins FXI and PK independently or in combination with HK. Since contact proteins such as HK, PK and factor XII (FXII) have anti-coagulant and profibrinolytic functions, the pathophysiological role of aPE has yet to be elucidated. We propose that aPE of different specificities may initiate or promote characteristics pathological conditions in patients with thrombosis or recurrent pregnancy losses., (Copyright 2001 Academic Press.)

A 75-year-old female known to have a chronic myelomonocytic leukaemia and an acquired FXI deficiency (FXI level, 5%) related to a FXI inhibitor (38 Bethesda units) was admitted to the hospital for acute pneumonia associated with a bulky pleural effusion. A therapeutic puncture was found to be essential for the patient. But, such a procedure is a haemostatic challenge which requires adequate preparation. A first treatment composed of intravenous immunoglobulins and immunosuppressive therapy failed to eradicate the inhibitor and to restore a normal FXI level. In this context, steroids or FXI concentrates were not recommended. Thus, small doses of recombinant activated factor VII were used to achieve haemostasis. The procedure was successful, the tolerance was good and no adverse events occurred.

Inhibitors against factor XI (FXI) have been frequently described in patients who acquired inhibitors (due to auto-immune disorders, malignancies or infections), but less often in those with a congenital deficiency of this factor, who had received plasma infusions. The present report concerns one such inhibitor found in the plasma of a patient with chronic myelomonocytic leukaemia and infected by B19 parvovirus, who was neither a heterozygote nor a homozygote for FXI deficiency, and who had no bleeding tendency despite a very low FXI level. Taking this case into account, we discuss and present the clinical and biological features of acquired FXI deficiency caused by an inhibitor.

Two cases with congenital homozygous factor XI deficiency developed a factor XI inhibitor following repeated plasma transfusions. Case 1 was given cyclophosphamide, intravenous immunoglobulin, and steroids. The factor XI inhibitor disappeared on day 103 and cardiac catheterization was performed without complications after giving fresh frozen plasma. Case 2 was effectively managed by plasma exchange for cardiac catheterization and surgery. However, after five plasma exchange procedures, the same plasma volume exchange was not effective in shortening the activated partial thromboplastin time (APTT). A significant heparin rebound occurred 4 h after heparin neutralization with protamine sulphate for which the patient needed to have a blood clot evacuated from around the heart.

Platelet factor XI is an alternatively spliced product of the factor XI gene expressed specifically within megakaryocytes and platelets as an approximately 1.9-kb mRNA transcript (compared with approximately 2.1 kb in liver cells) lacking exon V. Flow cytometry with an affinity-purified factor XI antibody, with PAC1 antibody (to the GPIIb/IIIa complex on activated platelets), and with S12 antibody (to P-selectin, an alpha-granule membrane protein expressed on the platelet surface during secretion) on platelets activated with ADP, thrombin, thrombin receptor peptide (SFLLRN amide), or collagen at various concentrations exposed platelet factor XI and PAC1 antibody binding in parallel. Unactivated platelets expressed approximately 40% of total platelet factor XI but no PAC1 binding sites. Enhanced membrane exposure of platelet factor XI is independent of alpha-granule secretion, because ADP and collagen exposed platelet factor XI but no S12 binding sites. Platelets from four patients with plasma factor XI deficiency (<0.04 U/mL) had normal constitutive and activation-dependent expression of platelet factor XI. Well-washed platelets from normal and from factor XI-deficient donors incubated with low concentrations of thrombin (0. 05 to 0.1 U/mL) corrected the clotting defect observed with factor XI-deficient plasma. Thus, functionally active platelet factor XI is differentially expressed on platelet membranes in a tissue-specific manner both constitutively and in a concentration-dependent fashion by various agonists in the absence of detectable plasma factor XI.

Meningococcal septic shock (MSS) is complicated by activation of coagulation, fibrinolytic, and complement systems. We studied the contact system of the intrinsic pathway of coagulation in thirteen children with MSS. Activation was assessed upon admittance to the intensive care unit and 48 h thereafter, based on the measurement of factor XII- (FXII), prekallikrein- and factor XI (FXI) antigen levels, as well as on the detection of FXIa-FXIa inhibitor, FXIIa-C1-inhibitor, and kallikrein-C1-inhibitor complexes, respectively. Levels of FXII, prekallikrein and FXI were reduced to about 50% in all patients on admission, and were significantly higher 48 h later. FXIIa-C1-inhibitor complexes were elevated in 7 patients, and kallikrein-C1-inhibitor complexes in 2 patients. FXIa-alpha 1-antitrypsin complexes were elevated in all patients, FXIa-C1-inhibitor complexes in nine, and FXIa-antithrombin III complexes in one patient. We conclude that patients with MSS have activation of the contact system, which may contribute to activation of coagulation, and thus to morbidity and mortality.

We have produced a panel of ten monoclonal antibodies specific to coagulation factor XI. Western blot analysis demonstrates that 9 of these antibodies react with the heavy chain of factor XI and one with the light chain. Seven of these antibodies inhibit factor XI and factor XIa activity. We have used immobilised monoclonal antibody for the production of factor XI deficient plasma and to purify factor XI to homogeneity with high yield in a simple two-step procedure. These monoclonal antibodies were used to develop highly sensitive immunoassays capable of detecting less than 0.01 mu factor XI antigen ml-1. A strong correlation was found between antigen and activity levels in 11 patients with hereditary deficiency indicating that none was cross-reacting material positive. Cultured Hep G2 cells were found to synthesize small amounts of factor XI antigen and this could also be detected by functional assay and by western blot analysis.

Contact activation cofactor (CAC) facilitates the interaction of factors XI and XII. Patients lacking CAC have a coagulation defect and are deficient in high molecular weight kininogen. The coincidence of these two defects suggests that a single protein may be responsible for both physiologic functions. Immunologic and activity studies have been made on isolated CAC to clarify the relationship between CAC and kininogen. CAC forms a single precipitin line with anti-human kininogen, and antikininogen neutralizes CAC activity. CAC and high molecular weight kininogen show a reaction of identity on immunodiffusion against rabbit anti-CAC. Anti-CAC forms two precipitin lines with normal plasma which can be identified as high and low molecular weight kininogen. Monospecific immunoabsorbed anti-CAC forms a single precipitin line with plasma high molecular weight kininogen and neutralizes CAC activity. Cleavage of kinin fragment from CAC by insoluble trypsin or kalikrein does not proportionally reduce procoagulant activity. CAC neutralized by anti-CAC can release kinins on exposure to trypsin or kallikrein. The results support the conclusions that CAC procoagulant activity is a function of high molecular weight kininogen, that antigenic determinants unique to high molecular weight kininogen are shared by the CAC portion of the molecule, and that the clotting reactions may occur at a site removed from the kinin peptide.

The interaction of Factor XIIa with Factor XI was investigated using two monoclonal antibodies, one (3Cl) directed against the heavy chain of Factor XIa and the other (5F4) against its light chain. 3C1 either as intact IgG or as Fab' fragment, enhanced the rate of Factor XIa generation in the fluid phase but inhibited it in the presence of kaolin and high molecular weight (HMW) kininogen. In contrast, the Fab' fragments of 5F4 inhibited only the fluid phase activation and had no effect on the surface-mediated activation. 3C1 was found to block the binding of Factor XI to HMW kininogen, whereas 5F4 did not. We conclude: a domain on the heavy chain region of Factor XI is essential for binding to HMW kininogen and for optimal surface-mediated activation by Factor XIIa; and binding of 3C1 to Factor XI changes its conformation rendering it a more favorable substrate for Factor XIIa in the fluid phase.

Coagulation factor XI was purified from human plasma using ion-exchange chromatography and affinity chromatography on high molecular weight kininogen-Sepharose. A monospecific precipitating antiserum was prepared and used to study factor XI antigen. Factor XI did not migrate during electrophoresis at pH 8.3. High molecular weight kininogen (HMWK), an alpha globulin, reversibly associates with factor XI. Complex formation between HMWK and factor XI was observed under conditions of crossed-immunoelectrophoresis. Using Laurell rocket immunoelectrophoresis, it was shown that the isolated alkylated light chain of kinin-free HMWK formed a complex with factor XI. In contrast to previous studies of prekallikrein, titrations of factor XI with increasing amounts of HMWK did not give a simple titration curve, suggesting that factor XI dissociates from the complex during electrophoresis. Prekallikrein and factor XI were shown to compete for the same HMWK molecules under the conditions of immunoelectrophoresis, and prekallikrein appeared to have a higher affinity for binding to HMWK than factor XI. Quantitative determinations of factor XI antigen in plasma by rocket immunoelectrophoresis were made. The average amount of factor XI measured in plasma samples from 20 normal individuals was 4.5 micrograms/ml (range 3-6). No factor XI antigen was detected in plasma from a patient deficient in factor XI. Normal factor XI antigen levels were detected in 3 different HMWK-deficient plasmas only if the plasmas were reconstituted with purified HMWK (2 U/ml). Addition of HMWK to normal plasma resulted in an increase of the factor XI antigen rocket. At HMWK levels of 2 U/ml, no further increase of the factor XI antigen rocket was observed. Therefore, accurate measurement of factor XI antigen by rocket immunoelectrophoresis is possible only if an excess of HMWK is present.

A 56 year old white man with psoriasis without serologic evidence of systemic lupus erythematosus was found to have a circulating anticoagulant against Factor XI with a low activity of Factor IX which could not be fully explained. The activity of the circulating anticoagulant was not potentiated on incubation after addition of normal plasma and it was still persisting after 15 months despite periodic treatment with methotrexate and topical steroid creams. This acquired anticoagulant neither protected the patient from myocardial infarction nor produced any bleeding complications.

Association of a circulating factor XI anticoagulant and disseminated intravascular coagulation (DIC) is described in a 33-year-old woman. Although the patient had rheumatoid arthritis and a bacterial infection treated with antibiotics, the anticoagulant was thought to be secondary to systemic lupus erythematosus. Curiously, the low levels of factor XI did not prevent the DIC from developing.

The results of studies in a patient with congenital deficiency of Factor XI who developed an inhibitor are presented. The patient presented with a severe, apparently spontaneous bleed into the thigh, which progressed despite infusion of fresh frozen plasma, but which responded promptly to activated prothrombin complex. During therapy with plasma his clotting time and Factor XI level were unresponsive and a Factor XI inhibitor titer of 6,000 U/ml was attained. The inhibitor was isolated and found to be polyclonal immunoglobulin G (IgG), predominantly of subclass 4. The specificity of the antibodies for Factor XI was shown by the ability of isolated inhibitor bound to polyacrylamide beads to remove Factor XI selectively from normal plasma. The binding of (125)I-labeled factor XI to the inhibitor was studied and an affinity constant of 1.65 x 10(10) liter/mol was found. Complexing of the antibodies with Factor XI was shown to block multiple activities of the clotting factor. Factor XI complexed with antibody did not bind to high molecular weight kininogen or undergo activation and cleavage by two-chain Factor XII. The complex of activated Factor XI with inhibitor prevented the cleavage and activation of Factor IX. Hence the inhibitor appears to act by binding to multiple sites on the Factor XI molecule and preventing its interaction with other molecules. Clinically these interactions of the inhibitor with Factor XI result in a state of severe Factor XI deficiency. The clinical circumstances of the case, with severe hemorrhage refractory to plasma infusion but readily responsive to an alternate clot-promoting agent, suggest that a defect of intrinsic system activation was critical, supporting the inference that Factor XI does participate in normal hemostasis. The clinical course of this patient, who has only had two documented hemorrhages in the presence of the inhibitor, is not as severe as that of patients with severe Factor VIII or IX deficiency. This suggests that physiologic activation of Factors XI and IX does not occur exclusively in series because deficiency of factors XII, XI, VIII, and IX should then have similar hemostatic consequences. We propose that independent mechanisms for bypass of Factors XII and XI are important in physiologic activation of coagulation.

Because human platelets participate in the contact phase of intrinsic coagulation and contain a Factor XI-like coagulant activity, the nature of the Factor XI-like activity was examined and compared with purified plasma Factor XI. The platelet factor XI-like activity was sedimented with the particulate fraction of a platelet lysate, was inactivated by heat (t(1/2) 3.5 min, 56 degrees C), was not a nonspecific phospholipid activity, and was destroyed by treatment with Triton X-100. Isolated platelet membranes were four-fold enriched in Factor XI activity and similarly enriched in plasma membrane marker enzymes. The Factor XI-like activity of platelet membranes was detected only when assayed in the presence of kaolin, which suggests that it is present in an unactivated form and can participate in contact activation. Concanavalin A inhibited the Factor XI-like activity of platelet lysates and platelet membranes but not of plasma or purified Factor XI. A platelet membrane-Factor XI complex was isolated after incubation of membranes with purified Factor XI. The Factor XI activity of the platelet membrane-plasma Factor XI complex was inhibited by concanavalin A, whereas unbound plasma Factor XI retained activity. An antibody raised against plasma Factor XI inhibited the in vitro Factor XI activity of plasma and of the platelet membrane-plasma Factor XI complex but had no effect on the endogenous Factor XI-like activity of washed lysed platelets or isolated platelet membranes. Washed platelets and isolated platelet membranes obtained from a Factor XI-deficient donor without a history of excessive bleeding had normal quantities of platelet Factor XI-like activity and normal behavior in the contact phase of coagulation (collagen-induced coagulant activity). These results indicate that platelet membranes contain an endogenous Factor XI-like activity that is functionally distinct from plasma Factor XI.

Since optimal rates of factor IX activation by factor XIa require the presence of calcium ions and the heavy chain of the enzyme as well as the active-site-containing light chain, we have studied the effects of calcium ions and the heavy chain on the reaction kinetics. Whereas the amidolytic activities of factor XIa and of its active-site-containing light chain were almost indistinguishable, the two enzymes behaved quite differently when factor IX was the substrate. Factor XIa was 100-fold more potent in the presence of Ca2+ than in its absence. On the contrary, the presence or absence of Ca2+ made very little difference in the case of the isolated light chain of factor XIa. Moreover, the enzymatic activity of the light chain was almost identical with that of intact factor XIa when Ca2+ was absent. Using an optimal concentration of Ca2+, we studied the activation in the presence of various concentrations of two monoclonal antibodies, one (5F4) directed against the light chain of factor XIa and the other (3C1) against its heavy chain. Analysis of 1/V vs. 1/S plots showed that whereas inhibition by 5F4 was noncompetitive, 3C1 neutralized the enzyme in a classical competitive fashion. We conclude that in the calcium-dependent activation of factor IX by factor XIa the heavy chain of the enzyme is involved in the binding of the substrate and this is essential for optimal reaction rates.

During the initiation of intrinsic coagulation factors XI and XIa interact intimately with several other coagulation proteins (factor XIIa, high Mr kininogen, and factor IX) as well as with the platelet surface. To help elucidate these complex intramolecular interactions, we have prepared a collection of monoclonal antibodies directed against various epitopes in factor XI. We have utilized these reagents to isolate factor XI and the light chain of factor XIa on affinity columns, and to probe structure-function relationships involved in the interactions of factor XIa with factor IX. The isolated light chain of factor XIa retained greater than 90% of its amidolytic activity against the oligopeptide substrate pyro-Glu-Pro-Arg-pNA (S-2366), but only 3.8% of its clotting activity in a factor XIa assay and 1% of its factor IX activating activity in an activation peptide release assay. This suggests that regions of the heavy chain are required for development of coagulant activity and specifically for the interaction of factor XIa with factor IX. To test this hypothesis, the effects of three of the monoclonal antibodies (5F4, 1F1, and 3C1) on the function of factor XIa were examined. The results show that in a clotting assay the light chain-specific antibody (5F4) inhibits 100% of the factor XIa activity, whereas of the heavy chain-specific antibodies, one (3C1) inhibits 75% and another (1F1) only 17%. Similarly in the factor IX activation peptide release assay, antibody 5F4 inhibits 100% of the factor XIa activity, whereas 3C1 inhibits 75% and 1F1 inhibits 33%. We conclude that regions located in the heavy chain, in addition to those in the light chain, are involved in the interaction of factor XIa with factor IX and in the expression of the coagulant activity of factor XI.

A relatively potent antiserum against highly purified, unactivated human factor XI antigen was raised in a rabbit. This antiserum, after concentration, neutralized 50% of the factor XI clotting activity of a standard normal plasma at an antiserum dilution of 1/900. The antiserum was used in a neutralization-inhibition assay to study the relation between factor XI clotting activity and factor XI antigen in plasma from ten unrelated patients with homozygous factor XI deficiency and from 12 heterozygous family members of these patients. No evidence of factor XI antigen significantly in excess of factor XI activity was found in either group. All data to date have been consistent with the hypothesis that hereditary factor XI deficiency represents a genetic disorder resulting from the absence of factor XI molecule. Severity of bleeding in factor XI deficiency could not be correlated with the level of factor XI activity or factor XI antigen.