Xa Inhibitors Research Articles

Effect of the FXa inhibitors Rivaroxaban and Apixaban on platelet activation in patients with atrial fibrillation

Rivaroxaban and Apixaban, increasingly used for stroke prevention in non-valvular atrial fibrillation (AF), might impact platelet reactivity directly or indirectly. By inhibition of Factor Xa (FXa) they preclude not only generation of relevant thrombin amounts but also block signalling of FXa via protease activated receptors. However, weather FXa-inhibition affects platelet haemostasis remains incompletely known. One hundred and twenty-eight patients with AF on chronic anticoagulation with either Rivaroxaban or Apixaban for at least 4 weeks were included in the study. In a time course group (25 on Rivaroxaban, 13 on Apixaban) venous blood samples were taken before NOAC medication intake in the morning as well as 2 and 6h afterwards. In 90 patients (Rivaroxaban n = 73, Apixaban n = 17) blood samples were drawn during left atrial RFA procedures before as well as 10 and 60min after the first heparin application (RFA group). Platelet reactivity analyzed by whole blood aggregometry (Multiplate Analyzer, Roche) in response to ADP, Collagen, TRAP and ASPI (arachidonic acid) was not altered by Rivaroxaban or Apixaban neither in the time course nor in the RFA group. Moreover, soluble P-selectin, Thrombospondin, von Willebrand Factor and beta thromboglobulin plasma levels, measured by ELISA, showed no statistically significant changes in both clinical settings for either FXa-inhibitor. The present study fails to demonstrate any significant changes on platelet reactivity in patients with AF under chronic Rivaroxaban or Apixaban medication, neither for trough or peak levels nor in case of a haemostatic activation in vivo as depicted by RFA procedures.

New Insights in Thrombin Inhibition Structure-Activity Relationships by Characterization of Octadecasaccharides from Low Molecular Weight Heparin.

Low Molecular Weight Heparins (LMWH) are complex anticoagulant drugs that mainly inhibit the blood coagulation cascade through indirect interaction with antithrombin. While inhibition of the factor Xa is well described, little is known about the polysaccharide structure inhibiting thrombin. In fact, a minimal chain length of 18 saccharides units, including an antithrombin (AT) binding pentasaccharide, is mandatory to form the active ternary complex for LMWH obtained by alkaline β-elimination (e.g., enoxaparin). However, the relationship between structure of octadecasaccharides and their thrombin inhibition has not been yet assessed on natural compounds due to technical hurdles to isolate sufficiently pure material. We report the preparation of five octadecasaccharides by using orthogonal separation methods including size exclusion, AT affinity, ion pairing and strong anion exchange chromatography. Each of these octadecasaccharides possesses two AT binding pentasaccharide sequences located at various positions. After structural elucidation using enzymatic sequencing and NMR, in vitro aFXa and aFIIa were determined. The biological activities reveal the critical role of each pentasaccharide sequence position within the octadecasaccharides and structural requirements to inhibit thrombin. Significant differences in potency, such as the twenty-fold magnitude difference observed between two regioisomers, further highlights the importance of depolymerisation process conditions on LMWH biological activity.

A highly regular fucosylated chondroitin sulfate from the sea cucumber Massinium magnum: Structure and effects on coagulation

A fucosylated chondroitin sulfate MM was isolated from the sea cucumber Massinium magnum. Structure of this polysaccharide was determined using chemical and NMR spectroscopic methods. The backbone of MM was shown to consist mainly of chondroitin sulfate E units with a small portion (about 10%) of chondroitin sulfate A fragments. Practically one type of branches Fuc3S4S attached to O-3 of GlcA residues was found in the polysaccharide molecules. The main repeating units of MM are →4)-[α-l-Fuc3S4S-(1→3)]-β-d-GlcA-(1→3)-β-d-GalNAc4S6S-(1→, whereas the minor repeating units are →4)-[α-l-Fuc3S4S-(1→3)]-β-d-GlcA-(1→3)-β-d-GalNAc4S-(1→. Anticoagulant activity of MM determined in APTT and TT tests was shown to be lower than that of heparin, but higher than that of enoxaparin. In the experiments with purified proteins MM effectively potentiated inhibition of thrombin and factor Xa by ATIII. Besides, MM did not induce platelets aggregation in platelets rich plasma.

Betrixaban: a direct oral inhibitor of activated factor X for the prophylaxis of venous thromboembolism in patients hospitalized for acute medical illness.

Venous thromboembolism (VTE), comprising deep vein thrombosis and pulmonary embolism, is a serious clinical and public health concern. Hospitalization is a major risk factor for developing VTE. Hospital-associated events account for more than 50% of all cases of VTE. Heparins have demonstrated to be efficacious in the prevention of VTE in medically ill patients. Despite the demonstrated efficacy and safety of the available direct oral anticoagulants in the prevention and treatment of different thromboembolic conditions, their net benefit in the prevention of VTE in hospitalized medically ill patients has not been fully confirmed. Betrixaban is an oral, specific and direct inhibitor of human coagulation factor Xa with demonstrated efficacy and safety for the prevention of VTE in patients undergoing total knee replacement and in patients with nonvalvular atrial fibrillation. Recent studies have successfully evaluated betrixaban 80 mg once daily in the prevention of VTE in acute medically ill patients in a large phase III trial. This review will address preclinical pharmacology and main aspects of the clinical development of betrixaban as an antithrombotic agent, with specific attention to recent studies on the prophylaxis of VTE in a specific population of patients hospitalized for acute medical illnesses.

Structure and anticoagulant property of a sulfated polysaccharide isolated from the green seaweed Monostroma angicava

An anticoagulant-active polysaccharide PF2 was extracted with boiling water from the green seaweed Monostroma angicava, further purified by anion-exchange and size-exclusion chromatography. PF2 was a rhamnan-type sulfated polysaccharide with molecular weight of about 88.1kDa. Results of chemical and spectroscopic analyses demonstrated that PF2 consisted of→3)-α-l-Rhap-(1→ and →2)-α-l-Rhap-(1→residues, with partially branches at C-2 of→3)-α-l-Rhap-(1→residues. Sulfate groups were substituted at C-3 of →2)-α-l-Rhap-(1→ residues. The sulfated polysaccharide PF2 had a high anticoagulant action, and the mechanism of anticoagulant activity mediated by PF2 was mainly attributed to strong potentiation thrombin by heparin cofactor II. PF2 also exhibited weak effect on antithrombin-dependent thrombin or factor Xa inhibition. The fibrin(ogen)olytic activity and thrombolytic activity of PF2 were also evaluated. The investigation revealed that PF2 was a novel sulfated rhamnan differing from previously described sulfated polysaccharides from green seaweed and could be a potential anticoagulant polysaccharide.

Porcine and Ovine Mucosal Heparins and Their Depolymerized Derivatives Are Comparable in Contrast to Their Bovine Equivalents

Introduction:The currently used unfractionated heparin (UFH) and low molecular weight heparins (LMWH) are mostly derived from Porcine mucosal tissue. Since the technology to manufacture heparin has advanced and the quality assurance practices are in place, improved products with high potency and purity are now available. Considering these factors the resourcing of heparins utilizing bovine (cow) and ovine (sheep) tissues is discussed at regulatory and pharmaceutical levels. The purpose of this study is to compare multiple individual batches of UFH obtained from Bovine, Ovine, and Porcine origin and their depolymerized product obtained by benzylation followed by alkaline hydrolysis representing enoxaparins.Materials and Methods:The molecular profile of the heparins and enoxaparins from various sources were determined using the size exclusion chromographic method. The anticoagulant potency was measured using clot based methods such as aPTT and Thrombin Time. The AT mediated anti-Xa and anti-IIa activities were also measured in defined biochemical sysytems. A comercially obtained chromogenic substrate based methods were used to determine the USP potency in terms of anti-Xa and anti-IIa activities (Hyphen Biomedical, Ohio, USA). The relative interaction of the heparins and enoxaparins with heparin induced thrombocytopenia (HIT) antibody induced aggregation of platelets were investigated using serum pool obtained from clinically confirmed HIT cases using aggregometry.Results:The molecular profile of multiple batches of the Bovine, ovine, and porcine heparins and enoxaparin were almost comparable and ranged from 15-18 kDa. The global anticoagulant and amidolytic protease assays for the bovine heparin were consistently lower than porcine and ovine samples. In the purified system the Porcine and Ovine preparations consistently showed lower IC50 values for both the thrombin and Xa inhibition in contrast to the bovine heparin. Similar trends were observed in the anti IIa assays. The USP potency of 28 batches of porcine ranged from 170-210 U/mg, whereas the 21 batches of ovine heparins exhibited comparable potencies in the range of 160-210 U/mL. In contrast the USP potency of 30 batches of bovine mucosal heparin was much lower and ranged from 110-140 U/mg. The anti-Xa - IIa ratio for the porcine and ovine heparins were comparable. However, the anti-Xa - IIa ratio of bovine heparin was somewhat lower. The ovine and porcine enoxaparin exhibited comparable potencies which ranged from 94-110 U/mg whereas bovine enoxaparin was slightly lower, ranging from 80-87 U/mg. However the antiXa and anti-IIa ratios of the enoxaparins derived from various species were comparable. In the HIT screening, there was no difference between the HIT responses in the heparins from different species. Similar results were obtained amongst enoxaparins of different origins.Conclusions:These studies show that heparins from bovine, ovine, and porcine origin exhibit comparable molecular profiles. While the porcine and ovine heparins exhibit similar biological potencies, the bovine heparin was found to be weaker. The enoxaparins derived from these species also exhibit similar molecular profiles, however, in functional assays, they exhibited similar trends where the bovine derived products were weaker. In the HIT screening profile, the heparins from all of these species produced stronger effects in comparison to their enoxaparin counterparts. These results suggest that heparin and enoxaparin derived from ovine mucosal tissue exhibit comparable biosimilar profiles. However, the bovine heparins and enoxaparin derived from bovine mucosal tissues are somewhat weaker. DisclosuresYao:Ronnsi Pharma: Employment.

Andexanet Alfa Reverses Anticoagulation Effects of Enoxaparin and Associated Bleeding in a Rabbit Acute Hemorrhage Model

Introduction: Andexanet alfa (andexanet) is a modified recombinant human factor Xa (FXa), developed to reverse the anticoagulation effects of both direct and indirect FXa inhibitors. For antithrombin III (ATIII)-dependent FXa inhibitors, such as enoxaparin, andexanet binds to the ATIII-enoxaparin complex with high affinity and reverses the inhibition of coagulation factors Xa and IIa. This study evaluated the ability of andexanet to reverse anticoagulation effects and blood loss due to the indirect FXa inhibitor, enoxaparin, in a rabbit liver laceration model.Methods: In a rabbit model of surgically-induced, acute hemorrhage (liver laceration model), rabbits were randomized to receive enoxaparin (8 mg/kg; subcutaneous bolus) or enoxaparin vehicle. This route of administration and dose of enoxaparin was based on the pharmacokinetic profile of enoxaparin and pharmacodynamic activity in the rabbit (2-fold increase in blood loss relative to non-anticoagulated rabbits). Andexanet (15, 35, or 75 mg per rabbit) or andexanet vehicle was administered to anesthetized rabbits as a 5-minute bolus, 120 minutes after enoxaparin injection (from Time 120 to Time 125 minutes), prior to livery injury. The five treatment groups were enoxaparin vehicle + andexanet vehicle, enoxaparin + andexanet vehicle, or enoxaparin + andexanet (3 dose levels). For liver laceration, a standardized injury (10 1-cm-long and 2- to 3-mm-deep incisions) was made into two liver lobes with 5 incisions in each lobe, and bleeding was allowed to progress for 15 minutes (from Time 125 to Time 140 minutes). Study efficacy endpoints included evaluations of blood loss in rabbits anticoagulated with enoxaparin, and anti-FXa activity in plasma.Results: In rabbits anticoagulated with enoxaparin, andexanet significantly decreased blood loss 2-3 fold, relative to vehicle control (assessed at Time 140 minutes) at all doses of andexanet administered (15, 35, 75 mg per animal) (see Table 1). The mean blood loss in anticoagulated rabbits administered andexanet was similar to that in non-anticoagulated rabbits (enoxaparin vehicle + andexanet vehicle). In enoxaparin-anticoagulated rabbits, andexanet rapidly (5 minutes after the start of andexanet administration; Time 125 minutes) reduced mean anti-FXa activity, and the reduction was significant for the 35-mg and 75-mg doses of andexanet (see Table 1). Anti-FXa activity did not return to baseline levels for the duration of the study from Time 120 to Time 140 minutes. These results are consistent with previous studies in enoxaparin-anticoagulated rats where a similar decrease in anti-FXa activity was sufficient to significantly reduce blood loss. These findings with an indirect (ATIII-mediated) FXa inhibitor enoxaparin are distinct from those with a direct FXa inhibitor (e.g., rivaroxaban), which showed that larger reductions in anti-FXa activity were required for significant reductions in blood loss.Conclusions: Administration of andexanet resulted in reversal of the anticoagulant effects of enoxaparin as well as restoration of hemostasis in enoxaparin-anticoagulated rabbits, suggesting it could be clinically valuable for the management of major bleeding associated with indirect FXa inhibitors. This study suggests a potential difference in the extent of reduction in anti-FXa activity required for significant reduction in blood loss associated with indirect vs. direct FXa inhibitors. Further studies are ongoing to better understand correlations between anti-FXa activity and blood loss following andexanet administration in enoxaparin-anticoagulated animals. [Display omitted] DisclosuresPine:Portola Pharmaceuticals: Employment. Lu:Portola: Employment, Patents & Royalties. Canivel:Portola Pharmaceuticals: Employment. Pratikhya:Portola Pharmaceuticals: Employment. DeGuzman:Portola Pharmaceuticals: Employment. Karbarz:Portola Pharmaceuticals: Employment. Takeda:Portola Pharmaceuticals: Employment. Malinowski:Portola Pharmaceuticals: Employment. Leeds:Portola Pharmaceuticals: Employment, Equity Ownership, Patents & Royalties, Research Funding. Curnutte:Portola Pharmaceuticals: Employment, Equity Ownership, Patents & Royalties, Research Funding; 3-V Biosciences: Equity Ownership; Sea Lane Biotechnologies: Consultancy. Conley:Portola Pharmaceuticals: Employment, Equity Ownership, Patents & Royalties, Research Funding.

Clinical relevance of pharmacokinetic and pharmacodynamic properties of edoxaban when treating patients with atrial fibrillation and heart failure

ABSTRACTIntroduction: Atrial fibrillation (AF) is an independent risk factor for stroke. It is most prevalent in the elderly and frequently coexists with heart failure (HF). The joint occurrence of AF and HF further worsens prognosis. The prevention of thromboembolism is crucial in the management of AF. In recent years, new oral anticoagulants (NOACs) have been licensed for the prevention of stroke and systemic embolism in patients with AF.Areas covered: This article reviews the key published studies on the pharmacology, clinical efficacy and safety of edoxaban, the latest NOAC to receive approval for the AF indication. This potent and selective inhibitor of factor Xa shows predictable pharmacokinetic and pharmacodynamic profiles. Its efficacy and safety have been demonstrated in the pivotal, phase III, warfarin-controlled ENGAGE AF-TIMI 48 trial in 21,105 AF patients.Expert opinion: NOACs will likely improve the management of AF, with or without HF. Edoxaban has a favorable pharmacokinetic profile that supports its use in special patient populations, including patients aged ≥75 years, with HF, renal impairment, poor adherence, and on polypharmacy. Proven strategies of edoxaban dose-reduction for optimal use in the presence of moderate renal impairment, and/or use of strong P-gp inhibitors are available.

The use of direct oral anticoagulants in inherited thrombophilia

To review the use of the direct oral anticoagulant (DOAC) agents in inherited thrombophilia based on the literature. MEDLINE, International Pharmaceutical Abstracts, and Google Scholar searches (1970-May 2016) were conducted for case reports, case series, retrospective cohorts, or clinical trials using the key words: protein C deficiency, protein S deficiency, antithrombin deficiency, activated protein C resistance, Factor V Leiden, hypercoagulable, NOACs, dabigatran, apixaban, rivaroxaban, betrixaban, edoxaban, Xa inhibitor, direct thrombin inhibitor. Results were limited to English-only articles. Clinical studies evaluating the use of DOACs for hypercoagulable states related to inherited thrombophilia were selected and evaluated. Thrombophilia, a predisposition to thrombosis, manifests predominantly as venous thromboembolism. Causes of inherited thrombophilia include antithrombin deficiency, deficiencies of proteins C and S, and Factor V Leiden mutation. Many patients with thrombophilia receive anticoagulant therapy for primary or secondary prevention of VTE, historically either warfarin or a heparin product. DOAC's have been considered as potential alternatives to traditional agents based on their pharmacologic activity. Case reports and a post-hoc analysis of a clinical trial have indicated positive results in patients with inherited thrombophilia and VTE. Positive results have been reported for the use of DOACs in inherited thrombophilia. Further robust studies are needed for definitive decision making by clinicians.

Andexanet Alfa for Acute Major Bleeding Associated with Factor Xa Inhibitors

BackgroundAndexanet alfa (andexanet) is a recombinant modified human factor Xa decoy protein that has been shown to reverse the inhibition of factor Xa in healthy volunteers.MethodsIn this multicenter, prospective, open-label, single-group study, we evaluated 67 patients who had acute major bleeding within 18 hours after the administration of a factor Xa inhibitor. The patients all received a bolus of andexanet followed by a 2-hour infusion of the drug. Patients were evaluated for changes in measures of anti–factor Xa activity and were assessed for clinical hemostatic efficacy during a 12-hour period. All the patients were subsequently followed for 30 days. The efficacy population of 47 patients had a baseline value for anti–factor Xa activity of at least 75 ng per milliliter (or ≥0.5 IU per milliliter for those receiving enoxaparin) and had confirmed bleeding severity at adjudication.ResultsThe mean age of the patients was 77 years; most of the patients had substantial cardiovascular disease. Bleeding was predominantly gastrointestinal or intracranial. The mean (±SD) time from emergency department presentation to the administration of the andexanet bolus was 4.8±1.8 hours. After the bolus administration, the median anti–factor Xa activity decreased by 89% (95% confidence interval [CI], 58 to 94) from baseline among patients receiving rivaroxaban and by 93% (95% CI, 87 to 94) among patients receiving apixaban. These levels remained similar during the 2-hour infusion. Four hours after the end of the infusion, there was a relative decrease from baseline of 39% in the measure of anti–factor Xa activity among patients receiving rivaroxaban and of 30% among those receiving apixaban. Twelve hours after the andexanet infusion, clinical hemostasis was adjudicated as excellent or good in 37 of 47 patients in the efficacy analysis (79%; 95% CI, 64 to 89). Thrombotic events occurred in 12 of 67 patients (18%) during the 30-day follow-up.ConclusionsOn the basis of a descriptive preliminary analysis, an initial bolus and subsequent 2-hour infusion of andexanet substantially reduced anti–factor Xa activity in patients with acute major bleeding associated with factor Xa inhibitors, with effective hemostasis occurring in 79%. (Funded by Portola Pharmaceuticals; ANNEXA-4 ClinicalTrials.gov number, NCT02329327.)

Primary and key secondary results from the ROCKET AF trial, and their implications on clinical practice.

The safety and efficacy of the oral anticoagulant rivaroxaban were studied in the Rivaroxaban Once Daily Oral Direct Factor Xa Inhibition Compared with Vitamin K Antagonism for Prevention of Stroke and Embolism Trial in Atrial Fibrillation (ROCKET AF trial). A number of subanalyses of the ROCKET AF trial have subsequently analyzed the use of rivaroxaban in special patient populations. The outcomes of the ROCKET AF trial were reviewed. The use of rivaroxaban in higher risk populations, as determined by the presence of co-morbidities included in the CHADS2 criteria, was analyzed. Requirements for dose adjustment in patients with renal impairment and in East Asian patients were described. Finally, clinical management challenges, including interruptions in therapy, drug discontinuation, management of bleeding events, drug interactions, and management of patients requiring cardioversion/ablation were reviewed. Rivaroxaban is efficacious in high-risk populations, including elderly patients, patients with diabetes, heart failure, history of stroke, prior myocardial infarction, or peripheral arterial disease (PAD). Patients with PAD have a higher risk of bleeding with rivaroxaban compared with warfarin. East Asian populations do not require a dose adjustment for rivaroxaban, while a reduced dose of 15 mg daily is required for patients with moderate renal impairment. Rivaroxaban remains effective with temporary interruptions in therapy and in patients requiring cardioversion/ablation. Rates of major bleeding and subsequent outcomes were similar in patients on warfarin and rivaroxaban, although rates of gastrointestinal bleeding were higher with rivaroxaban. Concurrent use of antiarrhythmic therapy was not associated with adverse outcomes. Rivaroxaban represents an efficacious alternative to warfarin in high-risk patients with AF. Dose adjustment is required for patients with moderate renal impairment. Rivaroxaban can be used safely in a number of challenging clinical management scenarios although the concurrent use of amiodarone requires more study.

Safety of Non-Vitamin K Antagonist Oral Anticoagulants in Clinical Practice: Focus on Rivaroxaban in Stroke Prevention in Patients With Atrial Fibrillation.

Despite direct oral anticoagulants (DOACs) have overcome the most relevant limitations of vitamin K antagonists, many patients with atrial fibrillation are not receiving the appropriate anticoagulant therapy. In addition, when patients are anticoagulated with DOACs, some of them are not taking the dose recommended in the summary of product characteristics. This may be related, at least in part, to the concern about the applicability of findings from randomized clinical trials to real-life patients. In this context, performing studies in daily clinical practice to assess the efficacy and safety of DOACs in real-world setting is mandatory. The aim of this review is to update the current evidence regarding safety of rivaroxaban in clinical practice. The Rivaroxaban Once Daily Oral Direct Factor Xa Inhibition Compared with Vitamin K Antagonism for Prevention of Stroke and Embolism Trial in Atrial Fibrillation (ROCKET-AF) showed that rivaroxaban was at least as effective as warfarin for preventing stroke or systemic embolism, with similar rates of major bleeding but with lesser risk of intracranial and fatal bleedings. Data from noninterventional studies and registries have confirmed the good results of ROCKET-AF. Even more, rates of bleeding may be inferior in real-life patients, since they have a lower risk profile than those included in ROCKET-AF.

Phosphatidylserine and Phosphatidylethanolamine Bind to Protein Z Cooperatively and with Equal Affinity.

Protein Z (PZ) is an anticoagulant that binds with high affinity to Protein Z-dependent protease inhibitor (ZPI) and accelerates the rate of ZPI-mediated inhibition of factor Xa (fXa) by more than 1000-fold in the presence of Ca2+ and phospholipids. PZ promotion of the ZPI-fXa interaction results from the anchoring of the Gla domain of PZ onto phospholipid surfaces and positioning the bound ZPI in close proximity to the Gla-anchored fXa, forming a ternary complex of PZ/ZPI/fXa. Although interaction of PZ with phospholipid membrane appears to be absolutely crucial for its cofactor activity, little is known about the binding of different phospholipids to PZ. The present study was conceived to understand the interaction of different phospholipids with PZ. Experiments with both soluble lipids and model membranes revealed that PZ binds to phosphatidylserine (PS) and phosphatidylethanolamine (PE) with equal affinity (Kd~48 μM); further, PS and PE bound to PZ synergistically. Equilibrium dialysis experiments revealed two lipid-binding sites for both PS and PE. PZ binds with weaker affinity to other phospholipids, e.g., phosphatidic acid, phosphatidylglycerol, phosphatidylcholine and binding of these lipids is not synergistic with respect to PS. Both PS and PE -containing membranes supported the formation of a fXa-PZ complex. PZ protection of fXa from antithrombin inhibition were also shown to be comparable in presence of both PS: PC and PE: PC membranes. These findings are particularly important and intriguing since they suggest a special affinity of PZ, in vivo, towards activated platelets, the primary membrane involved in blood coagulation process.

Interactions between depolymerized fucosylated glycosaminoglycan and coagulation proteases or inhibitors

Fucosylated glycosaminoglycan (FG) is a structurally novel glycosaminoglycan derivative, and it has potent anticoagulant activity. Depolymerized FG (dFG) is a selective factor Xase (FXase, FIXa-FVIIIa complex) inhibitor and it has antithrombotic action without major bleeding risks. In this study, we report the effects of dFG-3 (Mw ~14kDa) on the catalysis rates of factor IIa (FIIa), factor Xa (FXa) and factor IXa (FIXa) inhibition by antithrombin (AT), and the kinetic of the interactions between coagulation proteases or inhibitors and dFG-3 were also studied using biolayer interferometry (BLI) technology. We found that dFG-3 had much weaker catalysis activity of coagulation proteases inhibition by AT compared with heparin (UFH). The binding affinity of AT bound to dFG-3 was lower than UFH, and the UFH-AT interaction fitted well with biphasic-binding model while dFG-3-AT interaction was monophasic-binding, suggesting dFG-3 might not have allosteric activation effect on AT. The results are consistent with AT-independent inhibitory activities of dFG-3. dFG-3 could strongly bind to FIXa with much higher affinity than UFH, further explained the reason for its potent FXase inhibitory activity. Additionally, the binding ability of dFG-3 and FIXa decreased with decreasing molecular, and the fucose side chains and carboxyl groups of dFG-3 might be required for its high affinity binding with FIXa. Our data supports further the investigation of dFG-3 as a promising anticoagulant drug inhibiting the intrinsic FXase by binding to FIXa.

Rivaroxaban for secondary stroke prevention in patients with embolic strokes of undetermined source: Design of the NAVIGATE ESUS randomized trial.

Embolic strokes of undetermined source comprise up to 20% of ischemic strokes. The stroke recurrence rate is substantial with aspirin, widely used for secondary prevention. The New Approach riVaroxaban Inhibition of Factor Xa in a Global trial versus ASA to prevenT Embolism in Embolic Stroke of Undetermined Source international trial will compare the efficacy and safety of rivaroxaban, an oral factor Xa inhibitor, versus aspirin for secondary prevention in patients with recent embolic strokes of undetermined source. In patients with recent embolic strokes of undetermined source, rivaroxaban 15 mg once daily will reduce the risk of recurrent stroke (both ischemic and hemorrhagic) and systemic embolism (primary efficacy outcome) compared with aspirin 100 mg once daily. Double-blind, randomized trial in patients with embolic strokes of undetermined source, defined as nonlacunar cryptogenic ischemic stroke, enrolled between seven days and six months from the qualifying stroke. The planned sample size of 7000 participants will be recruited from approximately 480 sites in 31 countries between 2014 and 2017 and followed for a mean of about two years until at least 450 primary efficacy outcome events have occurred. The primary safety outcome is major bleeding. Two substudies assess (1) the relative effect of treatments on MRI-determined covert brain infarcts and (2) the biological underpinnings of embolic strokes of undetermined source using genomic and biomarker approaches. The New Approach riVaroxaban Inhibition of Factor Xa in a Global trial versus ASA to prevenT Embolism in Embolic Stroke of Undetermined Source trial is evaluating the benefits and risks of rivaroxaban for secondary stroke prevention in embolic strokes of undetermined source patients. Main results are anticipated in 2018.

Use of Dual Antiplatelet Therapy and Patient Outcomes in Those Undergoing Percutaneous Coronary Intervention: The ROCKET AF Trial.

The authors assessed the use of dual antiplatelet therapy (DAPT) and outcomes in patients undergoing percutaneous coronary intervention (PCI) during the ROCKET AF (Rivaroxaban Once Daily Oral Direct Factor Xa Inhibition Compared with Vitamin K Antagonism for Prevention of Stroke and Embolism Trial in Atrial Fibrillation). The frequency, patterns, and outcomes when adding DAPT to non-vitamin K antagonist oral anticoagulants in the setting of PCI in patients with AF are largely unknown. The study population included all patients in the treatment group of the ROCKET AF trial divided by the receipt of PCI during follow-up. Clinical characteristics, PCI frequency, and rates of DAPT were reported. Clinical outcomes were adjudicated independently as part of the trial. Among 14,171 patients, 153 (1.1%) underwent PCI during a median 806 days of follow-up. Patients treated with rivaroxaban were significantly less likely to undergo PCI compared with warfarin-treated patients (61 vs. 92; p=0.01). Study drug was continued during PCI in 81% of patients. Long-term DAPT (≥30 days) was used in 37% andsingle antiplatelet therapy in 34%. A small number switched from DAPT to monotherapy within 30 days of PCI (n=19 [12.3%]) and 15% of patients received no antiplatelet therapy after PCI. Rates of stroke/systemic embolism and major bleeding events were high in post-PCI patients (4.5/100 patient-years and 10.2/100 patient-years) in both treatment groups. In patients with AF at moderate to high risk for stroke, PCI occurred in<1% per year. DAPT was used in a variable manner, with the majority of patients remaining on study drug after PCI. Rates of both thrombotic and bleeding events were high after PCI, highlighting the need for studies to determine the optimal antithrombotic therapy.

Apixaban to prevent stroke in patients with atrial fibrillation: a review.

Atrial fibrillation is a common, costly and morbid cardiovascular arrhythmia. Stroke prevention remains the mainstay of treatment for atrial fibrillation, and the recent advent of novel oral anticoagulants with direct factor IIa or factor Xa inhibition has significantly revolutionized this aspect of treatment for atrial fibrillation patients. This review focuses on the tolerability and efficacy of apixaban and tackles the generalizability of the findings with apixaban to broader patient populations than those primarily enrolled in the clinical trials, drawing from the AVERROES and ARISTOTLE trials and their subsequent secondary analyses. Taken together, findings from these trials show that apixaban is superior to warfarin in preventing stroke with a lower risk of major bleeding in the general population of patients with atrial fibrillation as well as in several key high-risk patient subgroups.

Inhibition of coagulation proteases Xa and IIa decreases ischemia–reperfusion injuries in a preclinical renal transplantation model

Coagulation is an important pathway in the pathophysiology of ischemia-reperfusion injuries. In particular, deceased after circulatory death (DCD) donors undergo a no-flow period, a strong activator of coagulation. Hence, therapies influencing the coagulation cascade must be developed. We evaluated the effect of a new highly specific and effective anti-Xa/IIa molecule, with an integrated innovative antidote site (EP217609), in a porcine preclinical model mimicking injuries observed in DCD donor kidney transplantation. Kidneys were clamped for 60minutes (warm ischemia), then flushed and preserved for 24 hours at 4°C in University of Wisconsin (UW) solution (supplemented or not). EP217609-supplemented UW solution (UW-EP), compared with unfractionated heparin-supplemented UW solution (UW-UFH) or UW alone (UW). A mechanistic investigation was conducted invitro: addition of EP217609 to endothelial cells during hypoxia at 4°C in the UW solution inhibited thrombin generation during reoxygenation at 37°C in human plasma and reduced tumor necrosis factor alpha, intercellular adhesion molecule 1, and vascular cell adhesion molecule 1 messenger RNA cell expressions. Invivo, function recovery was markedly improved in the UW-EP group. Interestingly, levels of thrombin-antithrombin complexes (reflecting thrombin generation) were reduced 60minutes after reperfusion in the UW-EP group. In addition, 3 months after transplantation, lower fibrosis, epithelial-mesenchymal transition, inflammation, and leukocyte infiltration were observed. Using this new dual anticoagulant, anti-Xa/IIa activity during kidney flush and preservation is protected by reducing thrombin generation at revascularization, improving early function recovery, and decreasing chronic lesions. Such an easy-to-deploy clinical strategy could improve marginal graft outcome.

Anti-Xa activity in apixaban overdose: a case report

Introduction: Apixaban is a novel oral anticoagulation agent that exerts its effect through direct factor Xa inhibition. We present a case of multi-drug overdose including apixaban with associated apixaban concentrations.Case: A 53 year-old man presented to our metropolitan hospital following a deliberate self-poisoning with 200 mg apixaban, 35 mg ramipril, 105 mg bisoprolol, 280 mg atorvastatin, 6 mg colchicine, 37.4 mg magnesium, 4 × 500 mg paracetamol/9.5 mg codeine/5 mg phenylephrine and alcohol. He developed hypotension that was treated with noradrenaline. His initial and peak apixaban concentration was 1022.6 ng/ml and was associated with only minor bleeding from his femoral central line insertion site, which improved with local compression. Vitamin K 10 mg (at 9 h post-ingestion) and Prothrombinex-VF 2000 units (at 13 h post-ingestion) were also administered without any observed effect on coagulation studies. Apixaban elimination appeared to display first-order kinetics with an elimination half-life of 7.4 h. His plasma apixaban concentration was within the therapeutic dose range 10 h post-ingestion and he recovered uneventfully.Conclusion: A case of apixaban overdose with associated apixaban concentrations is presented. There was rapid resolution of anticoagulation with no demonstrable benefit of currently available clotting factor replacement.

Laboratory monitoring of rivaroxaban and assessment of its bleeding risk

Background: The aims of this study were to investigate the effects of rivaroxaban on routine coagulation assays using our local, widely available, reagents and to study the relationship between sensitive coagulation assays and bleeding risk caused by rivaroxaban.Methods: Prothrombin time (PT), activated partial thromboplastin time (APTT) and anti-factor Xa (FXa) chromogenic assays (Biophen DiXaI) and inhibition of FXa activity were performed in normal pooled plasma (NPP) spiked with rivaroxaban and plasma samples from patients treated with rivaroxaban.Results: In vitro, the linear correlation coefficient of measured concentrations of rivaroxaban, by Biophen DiXaI, and spiked concentrations of rivaroxaban was 0.99. PT and APTT showed good linear correlation with rivaroxaban concentrations, while other assays showed poor correlation. In vivo, PT showed a moderate linear correlation with rivaroxaban concentrations while APTT had a weak correlation with rivaroxaban concentrations. In vitro and in vivo, the rivaroxaban concentrations, measured by Biophen DiXaI, always showed good correlation with the inhibition of FXa activity, and PT values showed moderate correlation with the inhibition of FXa activity.Conclusions: Biophen DiXaI can be considered as a quantitative method to monitor the anticoagulation activity of rivaroxaban, and could be used to evaluate bleeding risk caused by rivaroxaban. The PT reagent (Thrombosis S) could be considered as a rough method to monitor the anticoagulation activity of rivaroxaban and evaluate bleeding risk caused by rivaroxaban.