Factor XI Activity Research Articles

Changes in Coagulation Factor XI Activity Levels in Patients with ST-Segment Elevation Myocardial Infarction Undergoing Primary PCI.

Background - Although Factor XI (FXI) inhibitors are currently tested for the prevention of thrombotic events, their early treatment could prevent thrombus consolidation in ST-segment elevation myocardial infarction (STEMI). This study aims to characterize coagulation FXI levels and their variations in patients with STEMI undergoing primary percutaneous coronary intervention (PCI). Methods - Patients with STEMI were prospectively enrolled between December 2023 and May 2024. FXI activity (FXIa) levels were measured at admission and after PCI (i.e., before discharge). Variations in FXIa levels were evaluated. Differences in indicators of thrombotic risk between groups with high and low FXIa variability were analyzed, and predictors of high FXIa variability were identified. Results - After screening, 54 patients with STEMI were included. The median FXIa level was 0.865 IU/mL (interquartile range [IQR] 0.554-0.978) at admission and 1.161 IU/mL (IQR 0.982-1.317) before discharge, with a median difference of +34.2% (p-value < 0.001). No significant differences were found in indicators of thrombotic risk between groups at high and low FXIa variability, except for the days intercurred between the essays (p-value = 0.016). Neither this nor other variables emerged as independent predictors of high FXIa variability. Conclusions - This study firstly reported an increase in FXIa levels from admission to discharge in STEMI patients undergoing PCI. Common indicators of thrombotic risk were not associated with FXIa levels or their variability. These findings aim to stimulate further research into anticoagulant therapies tailored to the patient's coagulative state and disease.

Laboratory and Molecular Diagnosis of Factor XI Deficiency.

The prevalence of factor XI (FXI) deficiency is 1 per 10 to 20,000 in the general population, much higher than that reported in most texts. The prevalence is higher in Ashkenazi Jews where it is about 1:20. Clinically, FXI deficiency presents as a mild bleeding disorder mostly associated with posttraumatic or postsurgical hemorrhages or unexplained minor bleeding. It is often discovered due to incidental finding of a prolonged activated partial thromboplastin time (aPTT) on routine laboratory screening. FXI deficiency is an autosomal recessive bleeding disorder with many causative F11 gene defects. Diagnosis is based on FXI activity, antigen levels, and molecular diagnostics. As FXI levels do not correlate with bleeding symptoms, identification of pathogenic genetic variants may be a more accurate predictor of bleeding risk and therefore aid in the clinical management of the patient. Two variants in the F11 gene account for most cases found in the Jewish and Arab populations. Patients with FXI deficiency can develop inhibitors to FXI although spontaneously acquired inhibitors are extremely rare. We will discuss laboratory and molecular assays used to diagnose FXI deficiency as well as interferences that can complicate diagnosis including new anticoagulants and acquired FXI inhibitors.

Factor XI activity in ST-segment elevation myocardial infarction

Abstract Background ST-segment elevation myocardial infarction (STEMI) is characterized by a significant thrombotic burden, where the coagulation cascade, including Factor XI (FXI) activity, plays a crucial role. In STEMI and other thrombotic conditions, FXI levels vary according to patient characteristics and timing from the event. Understanding the changes in FXI activity may offer valuable insights into the optimal patient selection and timing for use of investigational FXI inhibitors. Purpose Our aim was to characterize variations in FXI activity in STEMI patients. Methods In this prospective, observational study, patients admitted with STEMI underwent two measurements of plasma FXI activity, prior to coronary angiography and at discharge, using the HemosIL assay. The variation in FXI activity at these timepoints was compared between patients with and without coronary thrombus, as detected by an angiographic core-lab. Linear regression analyses were conducted to test the association of candidate predictors with FXI variation between admission and discharge (i.e., presence of coronary thrombus, PCI of the culprit artery, Thrombolysis in Myocardial Infarction flow post-PCI, bailout use of glycoprotein IIb/IIIa inhibitors, time to second measurement of FXI activity) and with FXI value at admission (i.e., age, sex, atrial fibrillation, late [&amp;gt;24 hours] STEMI presentation, presence of coronary thrombus). Results From November 2023 to February 2024, 49 STEMI patients were included. The median variation in FXI activity among admission and discharge was +24.8% (interquartile range [IQR] 11.8; 69.0; p&amp;lt;0.001). This variation was observed in both patients with (+41.6% [IQR 14.3; 74.6]; p&amp;lt;0.001) and without (+21.8% [IQR -2.3;40.7]; p=0.008) thrombus at coronary angiography, with significant interaction effects compared to the global STEMI population (p for interaction &amp;lt;0.001). An inverse linear regression relationship was observed between presence of coronary thrombus and changes in FXI activity (coefficient -33.0, 95% CI -59.5 to -7.2; p=0.012). Furthermore, in the model exploring predictors of FXI activity at admission, thrombus presence (coefficient -40.9, 95% CI -75.3 to -6.4; p=0.024) and subacute presentation (coefficient 90.8, 95% CI 21.9 to 159.8; p=0.015) emerged as significant predictors, albeit with contrasting effects on FXI activity. Figure 1 Conclusions Our findings reveal significant increases in FXI activity from admission to discharge in STEMI patients. The inverse relationship between the presence of coronary thrombus and FXI activity variations, along with the contrasting associations among initial FXI levels and subacute patients’ presentation or thrombus presence, support the importance of FXI in the coagulation process during STEMI. These findings suggest targeted FXI inhibition, especially early in STEMI management, could be beneficial, meriting detailed investigation in future studies.Multivariable linear regression analysis

First-in-human study to evaluate the safety, pharmacokinetics (PK), and pharmacodynamics (PD) of RBD4059, a GalNAc-siRNA drug targeting coagulation factor XI (FXI), in healthy subjects

Abstract Background Patients at risk of thrombosis/thromboembolism, require effective and safe anti-coagulation with high compliance. Inhibiting FXI has emerged as a promising strategy with potential for lower bleeding risk compared to currently approved anti-coagulants. Here we report on the safety, PK and PD effects of the GalNAc-conjugated siRNA RBD4059, specifically targeting and repressing FXI protein synthesis in the liver. Purpose Investigate safety, PK, and PD of first-in-human dosing of RBD4059 in healthy subjects. Methods A randomized, single-blind, placebo-controlled, single ascending dose (SAD) phase I study in healthy subjects (NCT05653037), in cohorts of eight randomized 3:1 to receive escalating doses of RBD4059 (50, 150, and 400 mg) or placebo administered subcutaneously. Results As of January 25, 2024, 24 subjects (with follow-up to week 24) with demographics and baseline clinical characteristics shown in Table 1 were enrolled. RBD4059 was well tolerated (Table 2), all related AEs were grade 1. There was no evidence of any clinically relevant bleeding events. Plasma exposure of RBD4059 tended to increase in proportion to the dose. Maximum concentration in plasma (Cmax) was reached in 2 to 4 h and then declined in line with one-compartment kinetics. The mean plasma half-life (t1/2) ranged from 3.2 to 7.0 h. RBD4059 treatment resulted in dose dependent and sustained reductions in FXI activity and FXI antigen (Figure 1A, B, D). Observed mean maximum percentage change from baseline in FXI activity from the 50 mg, 150 mg, and 400 mg SAD cohorts were 67.5%, 81.0%, and 85.8%, respectively, recovering with a maintained effect observed on D141 (50mg: 50.6% inhibition; 150mg: 68.2% inhibition). The reduction in FXI antigen showed a very similar pattern as the reduction in FXI activity, and reductions were accompanied by a concomitant increase in APTT. (Figure 1C, D). PKPD modelling supports that RBD4059 may be dosed every 3 months or less frequent with a &amp;gt;90% FXI reduction throughout the dosing interval, and will be tested in the upcoming phase 2 study. Conclusion(s) RBD4059 demonstrated favorable safety, predictable PK, and pronounced durable PD effect in the dose range of 50 mg - 400 mg. This first-in-human SAD study supports the further development of RBD4059 as an anticoagulant that can maintain a high level of PD effect with a once every 3 month or less frequent dosing regimen, with potential for enhanced compliance in chronic anti-coagulant therapy.Table 1 &amp; Table 2Figure 1.Panels A-D

Preclinical characterisation and first-in-human results on the tolerability and pharmacodynamic effect of REGN9933, a monoclonal antibody targeting the factor XI/activated factor XI apple 2 domain

Abstract Background/Introduction Standard-of-care anticoagulants are associated with increased bleeding risk. Genetic/pharmacological data suggest coagulation factor XI (FXI) inhibition can prevent thrombosis with minimal increased bleeding risk. We present preclinical and first-in-human (FIH) data on REGN9933, a FXI-binding monoclonal Ab. Purpose To characterise REGN9933 epitope binding sites, affinities for FXI and activated FXI (FXIa), and effect on high molecular weight kininogen (HMWK)’s interaction with FXI. Also, to assess safety, tolerability and pharmacodynamics in healthy human participants (pts). Methods Epitope binding sites mapped with cryogenic electron microscopy, binding affinities determined with plasmon resonance techniques, effect on thrombin generation assessed with a thrombin generation assay, and impact on HMWK:FXI interaction analysed using ELISAs. In the FIH, Ph 1, randomised, double-blind, single-centre study, pts received a single dose of REGN9933 intravenously (IV; 3–300 mg) or subcutaneously (SC; 100 mg &amp; 300 mg), or placebo (PBO). Primary endpoint: incidence and severity of treatment-emergent AEs (TEAEs). Other endpoints: effect of REGN9933 on activated partial thromboplastin time (aPTT) and prothrombin time (PT) to assess intrinsic and extrinsic pathway-triggered coagulation, respectively; and FXI activity. Results REGN9933 was found to bind the FXI apple 2 domain, with subnanomolar binding affinities for FXI and FXIa at 37°C, resulting in reduced thrombin generation from the intrinsic (not extrinsic) pathway. REGN9933 competed with HMWK for FXI binding in a dose-related manner. In the Ph 1 study, 56 pts received REGN9933 (IV, n=30; SC, n=12) or PBO (n=14); 42.9% and 21.4% had ≥1 TEAE, respectively (most common with REGN9933: headache, 12%; oropharyngeal pain, 5%). No serious/severe TEAEs, or TEAEs of special interest (inc. moderate/severe bleeding) reported. Laboratory tests revealed no clinically meaningful differences in the number/type of treatment-emergent potentially clinically significant values with REGN9933 vs PBO. REGN9933 resulted in dose-dependent prolongation of aPTT; aPTT mean fold change from baseline was highest with IV 300 mg after 8 hrs (2.7) and remained &amp;gt;2 for 36 days (Fig 1). There was no detectable effect on PT or clinically meaningful effect on bleeding time. REGN9933 supressed FXI activity in a dose-dependent manner, with suppression to approx. the lower limit of assay quantification (5%) with IV ≥30 mg (Fig 2). Conclusion(s) REGN9933 binds the FXI apple 2 domain and inhibits HMWK:FXI interaction, preventing intrinsic pathway-triggered FXI activation. FIH data showed dose-dependent inhibition of the intrinsic coagulation pathway by REGN9933, without affecting the extrinsic or common pathways. Pharmacodynamic and safety findings support continued development of REGN9933 as an anticoagulant that may carry less bleeding risk than currently approved agents.

The first-in-human study to assess the safety, tolerability, pharmacokinetics, and pharmacodynamics of the factor XI monoclonal antibody SHR-2004 in healthy subjects

ABSTRACT Background Inhibiting the coagulation factor XI (FXI) is a novel strategy for prevention and treatment of thromboembolism without affecting extrinsic coagulation pathways. SHR-2004 is a humanized monoclonal antibody that selectively binds to FXI and factor XIa (FXIa). Research design & methods This randomized, double-blind, dose-escalation, placebo-controlled study evaluated SHR-2004 administered either intravenously (i.v.; Part A) or subcutaneously (s.c.; Part B). In Part A, 24 subjects received a single i.v. dose of SHR-2004 (0.1, 0.3, or 1.0 mg/kg) or placebo. In Part B, 40 subjects received a single s.c. dose of SHR-2004 (0.5, 1.0, 3.0, or 4.5 mg/kg) or placebo. Results SHR-2004 was well tolerated. Plasma exposure to SHR-2004 increased in a dose-dependent manner. The geometric mean half-time ranged from 11.6 to 13.0 days. FXI activity decreased, and the activated partial thromboplastin time (APTT) was prolonged after i.v. and s.c. administration in a dose- and time-dependent manner. FXI activity was nearly completely abolished immediately after administering the highest i.v. dose, with the average APTT prolonged to nearly three times of baseline. Conclusion SHR-2004 is a promising candidate for further development as an anticoagulant drug that exerts effective anticoagulation with minimal risk of bleeding. Clinical trial registration www.clinicaltrials.gov identifier is NCT05369767.

Pharmacokinetics, pharmacodynamics, and safety of fesomersen, a novel antisense inhibitor of factor XI, in healthy Chinese, Japanese, and Caucasian volunteers.

The inhibition of coagulation factor XI (FXI) presents an attractive approach for anticoagulation as it is not expected to increase the risk of clinically relevant bleeding and is anticipated to be at least as effective as currently available anticoagulants. Fesomersen is a conjugated antisense oligonucleotide that selectively inhibits the expression of FXI. The article describes three clinical studies that investigated the safety, pharmacokinetic (PK), and pharmacodynamic (PD) profiles of fesomersen after subcutaneous (s.c.) injection to healthy participants. The studies included participants from diverse ethnic backgrounds (Caucasian, Japanese, and Chinese). Fesomersen demonstrated good safety and tolerability in all three studies. No major bleeding events were observed. After single-dose s.c. injection, fesomersen was rapidly absorbed into the systemic circulation, with maximum fesomersen-equivalent (fesomersen-eq) concentrations (Cmax) in plasma observed within a few hours. After reaching Cmax, plasma fesomersen-eq concentrations declined in a biphasic fashion. The PD analyses showed that the injection of fesomersen led to dose-dependent reductions in FXI activity and increases in activated partial thromboplastin time (aPTT). The maximum observed PD effects were reached between Day 15 and 30, and FXI activity and aPTT returned to near-baseline levels by Day 90 after a single dose. The PK/PD profiles after a single injection were similar among the various ethnic groups. Collectively, the study results suggest that fesomersen has a favorable safety profile and predictable and similar PK and PD profiles across Chinese, Japanese, and Caucasian participants.

First-in-Human Study to Assess the Safety, Tolerability, Pharmacokinetics and Pharmacodynamics of Factor XI Monoclonal Antibody SHR-2004 in Healthy Subjects

Introduction: Thromboembolic disease is a major global health problem. Inhibition of coagulation factor XI (FXI) is a novel strategy for the prevention and treatment of thrombotic diseases without affecting exogenous coagulation pathways. SHR-2004 is ahumanized monoclonal antibody that selectively binds to FXI and FXIa, hindering the activation of FXI by FXIIa and thereby blocking the cascade amplification process of endogenous coagulation pathways and exerting anticoagulant effects. This first-in-human, phase I study aimed to assess the safety, tolerability, pharmacokinetics (PK), pharmacodynamics (PD) and immunogenicity of SHR-2004 in healthy Chinese subjects. Methods: This single-center, randomized, double-blind, dose-escalation, placebo-controlled study (NCT05369767; Figure 1) evaluated SHR-2004 adminstered intravenously (i.v., part A) or subcutaneously (s.c., part B). In part A, 24 healthy subjects (8 for each dose level) received single i.v. dose of SHR-2004 (0.1, 0.3 or 1.0 mg/kg) or placebo in a randomization ratio of 3:1. In part B, 40 subjects (10 for each dose level) received single s.c. dose of SHR-2004 (0.5, 1.0, 3.0 or 4.5 mg/kg) or placebo (4:1). The safety, tolerability, PK, PD and immunogenicity of SHR-2004 were assessed. Results: SHR-2004 was well tolerated at all tested doses, and all adverse events were mild. A total of 88 treatment-related adverse events (TRAEs) were reported in 35 of 50 subjects assigned to SHR-2004 (70.0% of subjects), and 20 TRAEs were reported in 8 of 14 subjects receiving placebo (57.1%). Two subjects in the SHR-2004 cohorts and one in the placebo cohort showed mild ecchymosis. There was no evidence of other clinically relevant bleeding events. The plasma exposure of SHR-2004 (maximum concentration, C max and area under the plasma concentration-time curve, AUC) increased in a dose-dependent manner for both i.v. and s.c. dose cohorts (Figure 2). The median time-to-peak concentration (T max) was 1.76 -2.79 hours after the start of i.v. infusion, and 4.0-6.5 days after s.c. injection, respectively. The mean elimination half-life (t 1/2) ranged from 12.4 to 13.2 days following i.v. infusion, and 11.7 to 12 days after s.c. injection. It was observed that FXI activity decreased and activated partial thromboplastin time (APTT) was prolonged after i.v. and s.c. administration in a dose- and time-dependent manner. In the 1.0 mg/kg i.v. cohort, FXI activity was almost completely inhibited immediately after completion of the infusion, with average APTT prolonged to nearly three times that of baseline, and these effects persisted until day 9. Following s.c. administration, more than 80% inhibition was achieved within 12h after injection in the 3 mg/kg and 4.5 mg/kg cohort, and the maximum inhibition was maintained to day 22 and day 29, respectively. Similarly, the APTT was prolonged more than 1.7-fold in a gentle and sustained manner from 12h post-dose to day 29 in the 3 mg/kg and 4.5 mg/kg dose levels. Conclusion: Single intravenous infusion (0.1-1.0 mg/kg) and subcutaneous injection (0.5-4.5 mg/kg) of SHR-2004 are safe and well tolerated. SHR-2004 showed predictable PK with a relatively slowly elimination, which supports administration once a month in the future clinical and market. The encouraging PD data demonstrated a fast and sustained anticoagulant effect. These data indicate that SHR-2004 is a promising candidate for further development as a potential anticoagulant drug, which is expected to minimize the risk of bleeding while maintaining effective anticoagulation.

Factor XI Deficiency in Chinese Patients: Results from a Nationwide Multicenter Retrospective Study

Background: Factor XI(FXI) deficiency is a rare and relatively mild bleeding disorder. To date, data for China is limited. Our study aimed to investigate the clinical phenotype, F11 genotype and management of individuals diagnosed with FXI deficiency from throughout China. Methods: The study retrospectively analyzed patients with FXI activity (FXI:C) &amp;lt;50% registered from the National Hemophilia Registration System. The study collected patient characteristics, F11 gene mutations, clinical history, bleeding events, and treatment strategy by telephone follow-up. RESULTS: The study included 234 patients throughout 27 provincial districts registered in 38 Chinese hemophilia treatment centers. Among these patients, 192 (82.1%) had severe FXI deficiency (FXI:C &amp;lt;15%), and 42 (17.9%) had non-severe FXI deficiency (15%≤FXI:C&amp;lt;50%). The median age was 30 (1-87) years at diagnosis. There were 19 patients (8.1%) who reported a family history of bleeding. Five patients (2.1%) reported having consanguineous marriages in their families. There were 57 patients (24.4%) had their F11 gene tested. We detected 44 kinds of variants on 99 alleles, including nine that have never been reported to the F11 database. The most frequent variants were missense (44, 44.4%) and nonsense mutations (35, 35.4%). Among the genotyped subjects, F11 mutations were homozygous in 12 (21.0%), compound heterozygous in 29 (50.9%), and heterozygous in 16 (28.1%) patients. The FXI:C among patients with homozygous/compound heterozygous mutations was significantly lower than among patients with heterozygous mutations (2.1±2.3% vs. 14.8±12.9%, P&amp;lt;0.001). In total, 91 (38.9%) reported bleeding symptoms to varying degrees. Among these patients, 79 (33.8%) experienced spontaneous bleeding, with menorrhagia, epistaxis, and dermatorrhagia being the most prevalent manifestations. Twenty-one (26.6%) patients were administered on-demand replacement therapy. There were 129 patients underwent 220 invasive procedures with a total of 22 bleeding episode. The risk of bleeding was found to be significantly higher when sites with high fibrinolysis were involved, as compared to sites without fibrinolysis (Chi2=3.976, P=0.046). The prophylactic replacement treatment was administered to 50 cases (22.8%), and three bleeding events occurred, compared to 19 bleeding events in the 170 cases without prophylaxis. It was observed that prophylaxis did not reduce bleeding events (Chi2=1.150, P=0.283). A total of 83 deliveries were recorded in 58 female patients, and four bleeding events occurred. The blood group-O is a risk factor for bleeding [odds ratio (OR)=2.429, 95% confidence interval (CI) 1.139-5.182]. Conclusion: Most patients had no or mild bleeding symptoms. Most F11 variants were missense. The FXI:C among patients with homozygous/compound heterozygous mutations was significantly lower than among patients with heterozygous mutations. The risk of bleeding was higher when procedure sites with high fibrinolysis were involved. Prophylactic repalcement treatment could not reduce the risk of hemorrhages after invasive procedure. There is increased risk of bleeding in patients with blood group-O. Table 1 Table 2

Inhibition of Factor XI using a GalNAc-siRNA RBD5049: a novel antithrombotic drug with high potency and long duration

Abstract Introduction Bleeding risk remains a major concern of the existing anticoagulants. Inhibition Factor XI (FXI) using small molecule inhibitors, antibodies or antisense oligonucleotides (ASOs) to inactive intrinsic pathway of coagulation have been developed as a safer treatment option over the last decade. siRNA is a nucleic acid drug with class advantages of high selectivity, long duration and benign safety profiles. The current abstract summarizes key preclinical efficacy and safety data supporting the first-in-human study. Purpose To evaluate efficacy and safety, of a GalNAc-siRNA molecule RBD4059, for development of a novel antithrombotic treatment. Methods In vitro potency of several siRNAs were assessed and screened in HepG2 cells. Upon selection of the lead candidate, the siRNA was conjugated with a GalNAc molecule, using RIBO-GalSTAR liver targeting system. In vivo efficacy of RBD4059 was tested in C57BL/6J mice and cynomolgus non-human primates (NHPs). The antithrombotic effect of RBD4059 was determined in arterial and venous thrombosis models induced by FeCl3. Enoxaparin sodium (Lovenox) with clinical equivalent dose (4 mg/kg) was used as a control. Single- and repeated- dose GLP toxicology studies were performed in both CD-1 mice and NHPs. Results IC50 values of RBD4059 was 0.56 nM HepG2 cells. Dose-dependent and long-lasting effects of RBD4059 on the inhibition of plasma FXI activity (FXIa) were observed in mice and in NHPs. RBD4059 at a single dose of 9 mg/kg achieved a maximum of 89% plasma FXIa reduction and 1.38-fold APTT prolongation in mice, and the inhibitory effects could be maintained until Day 64. RBD4059 at 3 mg/kg or 9 mg/kg significantly prevented flow reduction due to FeCl3-induced thrombosis in both arterial and venous thrombosis models, and efficacy was superior than enoxaparin. In NHPs, the inhibitory effects of FXIa were maintained until Day 71 with the maximum effect of 77% FXIa reduction and 1.34-fold APTT prolongation. GLP toxicology studies showed good safety profile without any signs of increased bleeding despite supra-physiological doses and the NOAEL was considered 600 mg/kg in mice and 400 mg/kg in NHPs. Conclusions RBD4059 has demonstrated high potency/efficacy and long duration regarding FXI inhibition and with large safety margin. As the first siRNA based molecule targeting FXI, RBD4059 has now entered Phase 1 trial in human healthy volunteers.Figure 1

Coagulation abnormalities in a prospective cohort of 50 patients with PMM2-congenital disorder of glycosylation.

Given the lack of reliable data on the prevalence of bleeding abnormalities and thrombotic episodes in PMM2-CDG patients, and whether coagulation abnormalities change over time, we prospectively collected and reviewed natural history data. Patients with PMM2-CDG often have abnormal coagulation studies due to glycosylation abnormalities but the frequency of complications resulting from these has not been prospectively studied. We studied fifty individuals enrolled in the Frontiers in Congenital Disorders of Glycosylation Consortium (FCDGC) natural history study with molecularly confirmed diagnosis of PMM2-CDG. We collected data on prothrombin time (PT), international normalized ratio (INR), activated partial thromboplastin time (aPTT), platelets, factor IX activity (FIX), factor XI activity (FXI), protein C activity (PC), protein S activity (PS) and antithrombin activity (AT). Prothrombotic and antithrombotic factor activities were frequently abnormal in PMM2-CDG patients, including AT, PC, PT, INR, and FXI. AT deficiency was the most common abnormality in 83.3% of patients. AT activity was below 50% in 62.5% of all patients (normal range 80-130%). Interestingly, 16% of the cohort experienced symptoms of spontaneous bleeding and 10% had thrombosis. Stroke-like episodes (SLE) were reported in 18% of patients in our cohort. Based on the linear growth models, on average, patients did not show significant change in AT (n=48; t(23.8)=1.75, p=0.09), FIX (n=36; t(61)=1.60, p=0.12), FXI (n=39; t(22.8)=1.88, p=0.07), PS (n=25; t(28.8)=1.08, p=0.29), PC (n=38; t(68)=1.61, p=0.11), INR (n=44; t(184)=-1.06, p=0.29), or PT (n=43; t(192)=-0.69, p=0.49) over time. AT activity positively correlated with FIX activity. PS activity was significantly lower in males. Based on our natural history data and previous literature, we conclude that caution should be exercised when the AT levels are lower than 65%, as most thrombotic events occur in patients with AT below this level. All five, male PMM2-CDG patients in our cohort who developed thrombosis had abnormal AT levels, ranging between 19% and 63%. Thrombosis was associated with infection in all cases. We did not find significant change in AT levels over time. Several PMM2-CDG patients had an increased bleeding tendency. More long-term follow-up is necessary on coagulation abnormalities and the associated clinical symptoms to provide guidelines for therapy, patient management, and appropriate counseling. Most PMM2-CDG patients display chronic coagulation abnormalities without significant improvement, associated with a frequency of 16% clinical bleeding abnormalities, and 10% thrombotic episodes in patients with severe antithrombin deficiency.

Abstract 132: Mechanism By Which Coagulation Factor XI Regulates Endothelial Cell Permeability And Barrier Function In Vitro And In Vivo

Introduction: Proteolysis of vascular endothelial (VE)-cadherin is a biomarker for disruption of endothelial cell (EC) barrier function, with elevated levels of soluble VE (sVE)-cadherin associated with atherosclerosis and sepsis. Factor XI (FXI) plays a regulatory role in inflammation, and while a relationship between FXI activity and VE-cadherin expression levels has been observed, the mechanism by which this process is mediated is unknown. Hypothesis: Activated FXI (FXIa) induces VE-cadherin proteolysis. Methods: Human umbilical vein ECs were stimulated with FXIa (30 and 5 nM) for six hours. Western blot analysis was used to study VE-cadherin proteolysis, disintegrin and metalloproteinase 10 (ADAM10) expression, FXIa-PAI-1 or -very low density lipoprotein receptor (VLDLR) interactions, and cell signaling events. SVE-cadherin in baboons challenged with Staphylococcus aureus was detected by ELISA. Results: FXIa (5 nM) generated a VE-cadherin fragment and increased ECs permeability; the serine protease inhibitor, PPACK, and an inhibitor of ADAM10 reversed these processes. FXIa induced the expression of active ADAM10 on ECs. The receptor-associated protein (RAP) blocked FXIa-induced VE-cadherin proteolysis by preventing FXIa interaction with PAI-1 and VLDLR on ECs. FXIa-VLDLR interaction induced Dab1 and Src kinases phosphorylation. FXIa activated the VEGFR2-PLCgamma1-ERK1/2 signaling pathway. The ERK1/2 inhibitor, LY3214996, and the Src kinases inhibitor, PP2, inhibited VE-cadherin proteolysis by FXIa. PLCgamma1 phosphorylation was also prevented with PP2. The VEGFR2 kinase inhibitor, SU1498, prevented the cleavage of VE-cadherin by FXIa; in contrast, a VEGF blocking antibody had no effect on FXIa-induced VE-cadherin cleavage. Finally, the inhibition of FXI activation in a baboon model of sepsis significantly decreased the levels of sVE-cadherin. Conclusion: The complex formation of FXIa with PAI-1 and VLDLR promoted Src kinase phosphorylation. Src activation triggered the VEGFR2-PLCgamma1-ERK1/2 signaling pathway, which induced the trafficking of active ADAM10 to the EC surfaces inducing the cleavage of VE-cadherin. FXI may be a druggable target to protect EC barrier function in inflammatory diseases.

Gene Variants in Two Families with Inherited Coagulation Factor XI Deficiency and Identification of Mutations

Introduction: Mutations in the F11 gene can cause factor XI (FXI) deficiency, leading to abnormal coagulation activity and injury-related bleeding tendency. Therefore, identifying F11 gene mutations and studying the molecular basis will help us understand the pathogenesis of FXI deficiency. Methods: Coagulation tests and gene sequencing analysis of all members were performed. FXI wild-type and mutant expression plasmids were constructed and transfected into HEK293FT cells. The FXI protein expression level was evaluated by ELISA and Western blot. Results: The FXI activity (FXI:C) and FXI antigen (FXI:Ag) of proband-1 were decreased to 2% and 5%, respectively. FXI:C and FXI:Ag of proband-2 were reduced to 15% and 32%, respectively. Four mutations were found in the two unrelated families, including c.536C>T (p.T179M), c.1556G>A (p.W519*), c.434A>G (p.H145R), and c.1325_1325delT (p.L442Cfs*8). In vitro studies in transiently transfected HEK293FT cells demonstrated that p.T179M, p.W519*, and p.L442Cfs*8 mutations significantly lowered the FXI levels in the culture media. The FXI levels in the culture media and cell lysates of p.H145R mutation were similar to the wild type. Conclusion: Our results confirm that the four mutations in the F11 gene are causative in the 2 FXI deficiency families. Moreover, the p.H145R mutation is a cross-reactive material (CRM)-positive phenotype. The other three mutations are CRM-negative phenotypes and lead to FXI protein secretion disorder.

Abstract 9836: Variability of FXI and Relationship With Other Coagulation Factors - Sex Specific Differences and Impact of Diabetes Mellitus

Introduction: Novel anticoagulants targeting inhibition of Factor XI (FXI) are in clinical development and may have lower adverse bleeding risk compared to FX inhibitors. Population based preclinical studies have shown lower risk of stroke in patients with lower levels of FXI. We sought to examine the relationship of FXI with other coagulation factors and prothrombotic markers in a cohort of patients with cardiac risk factors or established cardiovascular disease. Hypothesis: FXI activity is associated with alterations of other coagulation factors. Methods: Plasma samples from subjects enrolled in a cardiovascular biobank (n=418) were analyzed by multiplex assay for FXI, FXII, FXIII, prothrombin, PAI-1, tPA, fibrinogen, antithrombin, c-reactive protein. Results: Increasing quintiles of FXI were associated with increases in fibrinogen, antithrombin, c-reactive protein, FXIII, and PAI-1, but decreases in prothrombin and FXII (p&lt;0.001). FXI levels were higher in women than men (149±54 vs. 134±48; p&lt;0.01) and in patients with diabetes mellitus (150±50 vs. 139±52; p&lt;0.05). In patients with subsequent spontaneous major bleeding during follow up (n=10), FXI was not significantly different from those without bleeding events (p=0.41). Conclusions: FXI activity may exhibit sex specific differences, and FXI activity may be increased in patients with diabetes, which is a strong risk factor for myocardial infarction and stroke. FXI activity is closely tied to activity of various markers of thrombosis and clotting factors. Future studies are warranted to examine effects of FXI inhibition on other coagulation factors.

Abstract 133: Coagulation FXI Regulates Endothelial Cell Barrier Function By Inducing VE-cadherin Proteolysis In An ADAM10-dependent Manner

Introduction: Proteolysis of vascular endothelial (VE)-cadherin is a biomarker for disruption of endothelial cell (EC) barrier function, with elevated levels of soluble VE-cadherin associated with atherosclerosis and sepsis. Factor XI (FXI), known for its role in the coagulation cascade, also plays a regulatory role in inflammation. While a relationship between FXI activity, VE-cadherin expression levels, and endothelial barrier function has been observed, the mechanisms by which this process is mediated are unknown. Hypothesis: Activated FXI (FXIa) regulates EC barrier function by inducing VE-cadherin proteolysis. Methods: Human umbilical vein endothelial cells were stimulated with FXIa (30 and 5 nM) for six hours. VE-cadherin proteolysis and expression levels were analyzed by Western blot and immunofluorescence. ECs permeability was quantified by measuring the flux of Evans blue-albumin across the ECs. Immunoprecipitation and Western blot analysis were used to study FXIa-PAI-1 or -very low density lipoprotein receptor (VLDLR) interactions. Results: FXIa (5 nM) generated a VE-cadherin C-terminal fragment and decreased VE-cadherin expression on ECs; this process was reversed by the serine protease inhibitor, PPACK. Thrombin, kallikrein, or FXIIa did not induce VE-cadherin proteolysis. FXIa increased ECs permeability, yet did not induce the expression of VCAM-1. In contrast, while VCAM-1 expression was upregulated by tumor necrosis factor alpha or vascular endothelial growth factor, neither of these induced VE-cadherin proteolysis. A pharmacological inhibitor of a disintegrin and metalloproteinase 10 (ADAM10) abrogated the effects of FXIa on ECs, including proteolysis of VE-cadherin. Similarly, the low density lipoprotein antagonist, receptor-associated protein (RAP), inhibited FXIa-induced VE-cadherin proteolysis by preventing FXIa complex formation with PAI-1 and VLDLR on ECs. Conclusion: These results suggest that FXIa disrupts VE-cadherin-mediated EC permeability by increasing ADAM10 activity through complex formation and internalization by PAI-1 and VLDLR. It remains to be seen whether this pathway represents a druggable target to protect EC barrier function in inflammatory diseases by inhibiting FXIa.

First-In-Human Study to Assess the Safety, Pharmacokinetics, and Pharmacodynamics of SHR2285, a Small-Molecule Factor XIa Inhibitor in Healthy Subjects

Background: Targeting factor XI (FXI) is a promising therapeutic strategy for the treatment and prevention of thrombosis without increasing the risk of bleeding. Here, we assessed the safety, pharmacokinetics (PK), and pharmacodynamics (PD) of SHR2285, a novel FXIa inhibitor, in healthy subjects. Methods: In this randomized, double-blinded, placebo-controlled, dose-ascending single-dosing trial (NCT03769831), eligible volunteer subjects receive either SHR2285 or placebo in a 3:1 ratio. Subjects assigned to the SHR2285 group received a single oral dose of SHR2285 at 50 mg, which was subsequently escalated to 100 mg, 200 mg, and 400 mg. Safety, pharmacokinetics, and pharmacodynamics parameters were assessed. All subjects were followed for 6 days. Results: SHR2285 was well tolerated. All adverse events were grade 1, and there was no evidence of bleeding events. The PK results revealed a rapid onset of action of SHR2285 (median time to maximum plasma concentration [Tmax] in different dose groups ranged 3.0–4.0 h) and the mean half-life ranged from 7.6 to 15.8 h. The metabolite SHR164471 had a slightly longer Tmax than the parent SHR2285, reaching a peak at a median of 6.0–7.0 h, and its mean half-life were 10.1–14.7 h in different dose groups. The sums of the area under the concentration–time curve from zero to time infinity of SHR2285 and SHR164471 in the 200 and 400 mg groups were similar, indicating the sum pharmacological activity of SHR2285 and SHR164471 showed a saturation trend between 200 and 400 mg. PD analysis showed that the inhibition of FXI activity was synchronized with prolonged activated partial thromboplastin time after SHR2285 administration, but the serum prothrombin time and international normalized ratio levels were not affected by SHR2285. Conclusion: SHR2285 demonstrated favorable safety, PK, and PD profiles in the dose range of 50 mg–400 mg. This first-in-human study supports the further development of SHR2285 for indications requiring anticoagulation. Clinical Trial Registration: https://clinicaltrials.gov/ct2/show/NCT03769831, identifier [NCT03769831].

Phase 2 Study of the Factor XI Antisense Inhibitor IONIS-FXIRx in Patients With ESRD

IntroductionPatients with end-stage renal disease (ESRD) requiring hemodialysis (HD) have an increased risk of thrombotic events and bleeding. Antisense reduction of factor XI (FXI) with IONIS-FXIRx is a novel strategy that may safely reduce the risk of thrombotic events.MethodsThis multicenter study enrolled 49 patients receiving HD in 2 parts. First, 6 participants (pharmacokinetics [PK] cohort) received 1 open-label 300 mg dose of IONIS-FXIRx both before and after HD. Subsequently, 43 participants were treated in a double-blind, randomized design with 200 mg or 300 mg IONIS-FXIRx or placebo for 12 weeks. The PK, pharmacodynamics (PD), and adverse events of IONIS-FXIRx were evaluated (ClinicalTrials.gov: NCT02553889).ResultsThe PK of IONIS-FXIRx was consistent with previous studies and similar whether injected before or after HD. No accumulation of IONIS-FXIRx was observed after repeat administration. By day 85, mean levels of FXI activity fell 56.0% in the 200 mg group, 70.7% in the 300 mg group, and 3.9% in the placebo group compared with baseline. FXI antigen levels paralleled FXI activity. Dose-dependent prolongation of activated partial thromboplastin time (aPTT) was observed, with no changes in international normalized ratio (INR). IONIS-FXIRx was not associated with drug-related serious adverse events. In the randomized phase of the study, major bleeding events occurred in 0 (0.0%; 200 mg), 1 (6.7%; 300 mg), and 1 (7.7%; placebo) patients and were not considered related to treatment.ConclusionIONIS-FXIRx reduced FXI activity in patients with ESRD receiving HD. Further studies are needed to determine the benefit-risk profile of FXI as a therapeutic target for patients who require HD.

The Association between Factor XI Deficiency and the Risk of Bleeding, Cardiovascular, and Venous Thromboembolic Events.

The objective of this study was to assess the relationship between factor XI (FXI) deficiency and the risks of bleeding and cardiovascular (CV) events. We conducted a retrospective cohort study using data from Maccabi Healthcare Services (MHS). We identified adults with FXI deficiency (severe: <15%, partial: 15 to <50%, any deficiency: <50%) that had been tested for FXI between 2007 and 2018 and matched to patients from the general MHS population. We estimated 10-year risks of outcomes using the Kaplan–Meier approach. Using Cox proportional hazards regression, we compared outcomes among patients with versus without FXI deficiency. Less than 10% of patients tested for FXI activity had activity levels <50% (mean age: 39 years; 72.2% females). Compared with the general population, patients with any FXI deficiency were at higher risk of severe bleeding (adjusted hazard ratio [aHR]: 2.56, 95% confidence interval [CI]: 1.13–5.81; 10-year risk: 1.90%, 95% CI: 0.50–3.20% vs. 0.90%, 95% CI: 0.50–1.30%) and clinically relevant nonsevere bleeding (CRNSB) (aHR: 1.45, 95% CI: 1.08–1.97; 10-year risk: 11.60%, 95% CI: 8.30–14.80% vs. 9.20%, 95% CI: 8.00–10.40%). Severe FXI deficiency was associated with a greater risk of CRNSB. While few CV events ( N = 2) and venous thromboembolisms (VTE) ( N = 1) were observed in the FXI overall deficient group, there was a nonsignificant negative association between any FXI deficiency and CV events (aHR: 0.55; 95% CI: 0.13–2.36) and VTEs (aHR: 0.45; 95% CI: 0.06–3.47). Overall FXI deficiency was associated with an increased risk of severe bleeding and CRNSB. Further research is warranted to explore the lower risk of CV and VTE among patients with FXI deficiency compared with the general population.

Phenotype and genotype analysis of patients with severe factor XI deficiency in Shaanxi Province, China.

Congenital coagulation factor XI (FXI) deficiency is a rare bleeding disorder with a heterogeneous haemorrhagic phenotype and various hotspot gene mutations associated with race and geography. Studies on FXI deficiency in Shaanxi Province, China, are scarce. In this study, seven patients with severe FXI deficiency and several family members were analysed. The International Society on Thrombosis and Hemostasis-Bleeding Assessment Tool (ISTH-BAT) was applied to assess bleeding symptoms. FXI activity was determined using a one-stage method, and the FXI antigen was measured by enzyme-linked immunosorbent assay. Targeted capture next-generation sequencing and Sanger sequencing were applied to detect FXI gene mutations. The bleeding phenotype varied, although none of the participants had a history of spontaneous bleeding. One maternal received replacement therapy during the perinatal period, one female patient presented with menorrhagia, one male patient experienced severe postoperative bleeding and others were asymptomatic. Family members with heterozygous mutations were all asymptomatic. The FXI activity of all the patients ranged from less than 1 to 3.1 IU/dl, and a synchronous decrease in the FXI antigen was observed. Two missense mutations (p. Gly350Glu and p. Cys482Trp), one nonsense mutation (p. Gln384∗) and one novel frameshift mutation (p. Ser225Phefs∗16) were detected. The bleeding manifestations and severity of severe FXI deficiency varied and were not related to its activity. Three reported mutations and one novel frameshift mutation were identified, thus extending the mutation spectrum of FXI deficiency.

Pharmacokinetics and Pharmacodynamics of Ionis-FXIRx, an Antisense Inhibitor of Factor XI, in Patients with End-Stage Renal Disease on Hemodialysis

BACKGROUNDPatients with end-stage renal disease (ESRD) on hemodialysis are at high risk of both thrombotic and bleeding events. Experimental and clinical data indicate that reducing factor XI (FXI), a key component of the intrinsic pathway, prevents thrombosis without causing bleeding. FXI activity can be lowered with IONIS-FXIRx, a 2nd generation antisense oligonucleotide that specifically reduces human FXI mRNA expression in the liver. This study sought to determine the safety, pharmacokinetics (PK) and pharmacodynamics (PD) of IONIS-FXIRx in participants with ESRD. METHODSIn this Phase 2 multicenter study, we enrolled a total of 49 ESRD patients: (A) 6 participants receiving chronic in-center hemodialysis (HD) to receive an open-label single-dose of 300 mg IONIS-FXIRx either 5 minutes before or 10 minutes after HD and assessed the effects of HD on peak (Cmax) and the extent of exposure (AUC0-24hr) parameters of IONIS-FXIRx and (B) another 43 participants in a double-blind design to one of two multiple-dose regimens (200 mg or 300 mg) IONIS-FXIRx or placebo for 12 weeks with 12 weeks of additional follow up. In the double-blind portion, IONIS-FXIRx was administered subcutaneously ~10 minutes after HD. For PK, plasma trough concentration and apparent elimination half-life (t1/2λz) were evaluated. The PD parameters including FXI activity and antigen, aPTT, prothrombin time (PT), and INR were evaluated in the per-protocol population (N= 34) that was defined as all participants who received the complete protocol-specified administration of IONIS-FXIRx or placebo through week 8 and who did not have any major protocol violations. The rate and frequency of clotting on the HD filters and circuit were qualitatively measured with a clotting scale as an exploratory PD analysis. RESULTSThe PK cohort included 6 males (5 white) with a mean age of 61 years. The plasma PK profile of IONIS-FXIRx was not significantly altered whether injected before or after HD. After multiple dosing, plasma Cmax values were dose-dependent and similar between Day 1 (first dose) and Day 78 (last dose), suggesting no accumulation of IONIS-FXIRx in ESRD participants after 12-weeks of dosing. After treatment cessation, IONIS-FXIRx had an apparent elimination half-life of approximately 2 weeks. In the double-blind portion of the study, 22 (51.2%) of the participants were male, 24 (55.8%) were white and the mean age was 59 years. Attenuation of FXI activity was associated with a reduction in the incidence of severe clotting events on both the air filter and dialyzer (Table). The reduction in severe clotting events was observed when FXI activity was < 0.4 U/mL. Consistent with prior observations in healthy subjects, IONIS-FXIRx prolonged aPTT in a time and dose-dependent manner with maximum mean percent changes from baseline of 53.3% (p < 0.05) for the 200 mg dose group and 75.6% (p < 0.05) for the 300 mg dose group at Week 14, but had no effect on PT. Treatment emergent adverse events (TEAEs) were mostly mild; participants reporting serious TEAEs were 1 (16.7%) for the PK cohort, 4 (30.8%) for placebo and 3 (20.0%) each for the 200 mg and 300 mg IONIS-FXIRx dose groups. There were no drug-related SAEs and no drug-related major or clinically-relevant non-major bleeding events. Minor bleeds were primarily observed in the 300 mg multiple-dose group at AV fistula/graft sites but did not correlate with reduction of FXI activity. No clinically significant reductions in platelet counts were reported during the study. CONCLUSIONSThis study demonstrated that (i) HD had no effect on IONIS-FXIRx PK, (ii) IONIS-FXIRx was well tolerated and produced sustained, and dose-dependent reductions in FXI antigen and activity, and (iii) IONIS-FXIRx reduced severe dialysis circuit clotting events beyond standard heparin use. Collectively, these data support further evaluation of IONIS-FXIRx as a potentially safe, effective antithrombotic therapy in ESRD patients on HD. [Display omitted] DisclosuresBethune:Ionis Pharmaceuticals: Employment. Walsh:Population Health Research Institute: Employment; Ionis Pharmaceuticals: Consultancy. Jung:Ionis Pharmaceuticals: Employment. Yu:Ionis Pharmaceuticals: Employment. Geary:Ionis Pharmaceuticals: Employment. Bhanot:Ionis Pharmaceuticals: Employment.