Oral FXa Inhibitor Research Articles

3-Factor prothrombin complex concentrate versus 4-factor prothrombin complex concentrate for the reversal of oral factor Xa inhibitors.

Multiple agents exist for the reversal of oral Factor Xa inhibitor (FXa) associated bleeding, including Coagulation FXa Recombinant, Inactivated zhzo (andexanet alfa) and 4-factor prothrombin complex concentrate (4F-PCC). While classified as a 3F-PCC product, Profilnine contains up to 35IU of Factor VII (per 100IU of Factor IX) in addition to therapeutic levels of Factors II, IX, and X, and has demonstrated a similar impact on prothrombin time and blood product usage in non-warfarin related bleeding. This was a retrospective, multicenter study at four medical centers of adult patients who presented with major bleeding associated with oral FXa inhibitors and received either 4F-PCC (n = 64) or 3F-PCC (n = 61). The primary outcome was hemostatic effectiveness. Secondary outcomes included the incidence of thromboembolism, in-hospital mortality, and length of stay. The most common indication for reversal was intracranial bleeding. For the primary outcome, 84% of all patients were rated as effective with no difference noted between the groups (p = 0.81). No significant difference between groups was found in the multivariable analysis adjusting for baseline differences between groups including race, total body weight, type of bleeding, and the use of antiplatelet therapy. There was no difference in the length of stay, in-hospital mortality, or the incidence of thromboembolism between the groups. Overall, no significant differences were found in the effectiveness or safety of 4F-PCC and 3F-PCC use in the management of oral FXa inhibitor-associated bleeding. Further investigations are warranted to explore the use of 3F-PCC for this indication and its safety and effectiveness.

Factor XI inhibitors: a new option for the prevention and treatment of cancer-associated thrombosis

Venous thromboembolism (VTE) is a relatively common complication in cancer patients with potentially dire consequences. Anticoagulants are the mainstay of treatment of cancer-associated VTE. The anticoagulants most often used are low-molecular-weight heparin (LMWH) and direct oral factor (F) Xa inhibitors, which include apixaban, edoxaban, and rivaroxaban. Most guidelines recommend primary VTE prophylaxis with LMWH, apixaban, or rivaroxaban after abdominal or pelvic cancer surgery, or in high-risk ambulatory cancer patients. Both oral FXa inhibitors and LMWH have limitations. LMWH requires daily subcutaneous injections, and because of its renal clearance, its use may be problematic in patients with severe kidney disease. The risk of bleeding with oral FXa inhibitors may be higher than with LMWH in patients with intraluminal gastrointestinal or genitourinary cancers. Other problems with oral FXa inhibitors include potential drug-drug interactions and dosing issues in patients with thrombocytopenia or severe kidney or liver disease. Therefore, there remains a need for convenient and safer anticoagulants for VTE treatment in cancer patients. FXI has emerged as a potentially safer target for anticoagulants than FXa because FXI is essential for thrombosis, but mostly dispensable for hemostasis. This review summarizes the currently available therapeutic options for cancer-associated VTE, highlights knowledge gaps, and discusses the potential of FXI inhibitors to address key unmet clinical needs in this vulnerable patient population.

Minimally modified human blood coagulation factor X to bypass direct factor Xa inhibitors

BackgroundDirect oral factor (F)Xa inhibitors are widely used as alternatives to conventional vitamin K antagonists in managing venous thromboembolism and nonvalvular atrial fibrillation. Unfortunately, bleeding-related adverse events remain a major concern in clinical practice. In case of bleeding or emergency surgery, rapid-onset reversal agents may be required to counteract the anticoagulant activity. ObjectivesThe ability of FXa variants to bypass the direct oral FXa inhibitors was assessed. MethodsHuman FXa variants were generated through substitution of phenylalanine 174 (F174) for either alanine, isoleucine, or serine. FXa variants were stably expressed in HEK293 cells and purified to homogeneity using ion-exchange chromatography. ResultsF174-substituted human FX variants demonstrated efficacy in restoring thrombin generation in plasma containing direct FXa inhibitors (apixaban, rivaroxaban, edoxaban). Their ability to bypass the anticoagulant effects stems from a significantly reduced sensitivity for the direct FXa inhibitors due to a decrease in binding affinity determined using molecular dynamics simulations and free energy computation. Furthermore, F174 modification resulted in a partial loss of inhibition by tissue factor pathway inhibitor, enhancing the procoagulant effect of F174-substituted FX. Consequently, the F174A- and F174S-substituted FX variants effectively counteracted the effects of 2 widely used anticoagulants, apixaban and rivaroxaban, in plasma of atrial fibrillation and venous thromboembolism patients. ConclusionThese human FX variants have the potential to serve as a rescue reversal strategy to overcome the effect of direct FXa inhibitors in case of life-threatening bleeding events or emergency surgical interventions.

Effects of the activated coagulation factor X (FXa) and its protease-activated receptor-2 (PAR2) on colon cancer cell growth in vitro and in vivo

Abstract Background Protease-activated receptors are a family of four (PAR1-4) G-protein-coupled receptors with a unique mechanism of activation. Serine proteases such as the activated coagulation factor X (FXa) can activate PARs, selectively PAR2, by proteolytic cleavage unmasking its tethered ligand which induces consecutive receptor activation. PAR2 has been linked to various pathophysiological processes such as cardiovascular or inflammatory diseases. Recent studies indicate a key role of PAR2 in certain cancer types. Purpose In this study, we investigated the impact of FXa, PAR2 and of the direct, oral FXa-inhibitor Apixaban in colon cancer cell growth and migration in vitro and in vivo. Methods FXa and PAR2-dependent effects were investigated in various colon cancer cell lines such as human HCT116 or murine MC38 cells in vitro using different assays for cell proliferation and migration, Western blotting and PCR analyses. A subcutaneous tumor model was utilized to compare the impact of Apixaban (5 and 50 mg/kg body weight) in C57Bl/6 wild type (WT) versus PAR2-deficient mice (PAR2-KO). Tumor cells were allowed to grow for up to 21 days. Apixaban was applied orally by gavage. All animal studies were approved by the responsible animal welfare committee. Results Incubation with FXa at concentrations up to 30 nM stimulated growth and migration in both cultured human and murine cells in vitro. These effects were mimicked by peptide-stimulated activation of PAR2. In comparison, thrombin or selective peptide-induced activation of PAR1 did not induce a marked response. In addition, we observed an activation of mitogenic signaling pathways such as p44/42, p38 and AKT by FXa suggesting a direct response for colon cancer cells to coagulation factor signaling. In vivo, PAR2-deficiency resulted in a significantly reduced tumor growth compared to wild-type mice. In comparison, oral treatment with Apixaban both at 5 mg or 50 mg/kg body weight did not markedly affect tumor development in vivo. This effect was reflected by a reduced tumor weight in PAR2-deficient mice after excision of the tumor at the end of the study. In addition, a significantly lower rate of complications was seen in PAR2-deficient mice during the experiment. Interestingly, the spleen was larger in relation to body weight in PAR2-deficient mice independently from Apixaban treatment. We observed no obvious histological differences within the tumor tissue between the treatment groups or animal strains. Conclusion Deficiently of PAR2 resulted in reduced growth of colon cancer cells in vivo, while oral treatment with Apixaban did not significantly affect tumor growth. In vitro, FXa and selective activation of PAR2 did stimulate cell growth and migration indicating a potential direct growth promotion effect of PAR2 in colon cancer cells. The pharmacological targeting of PAR2 may represent a future therapeutic strategy in the treatment of colon cancer.

Direct Oral FXa Inhibitors Binding to Human Serum Albumin: Spectroscopic, Calorimetric, and Computational Studies.

Direct FXa inhibitors are an important class of bioactive molecules (rivaroxaban, apixaban, edoxaban, and betrixaban) applied for thromboprophylaxis in diverse cardiovascular pathologies. The interaction of active compounds with human serum albumin (HSA), the most abundant protein in blood plasma, is a key research area and provides crucial information about drugs' pharmacokinetics and pharmacodynamic properties. This research focuses on the study of the interactions between HSA and four commercially available direct oral FXa inhibitors, applying methodologies including steady-state and time-resolved fluorescence, isothermal titration calorimetry (ITC), and molecular dynamics. The HSA complexation of FXa inhibitors was found to occur via static quenching, and the complex formation in the ground states affects the fluorescence of HSA, with a moderate binding constant of 104 M-1. However, the ITC studies reported significantly different binding constants (103 M-1) compared with the results obtained through spectrophotometric methods. The suspected binding mode is supported by molecular dynamics simulations, where the predominant interactions were hydrogen bonds and hydrophobic interactions (mainly π-π stacking interactions between the phenyl ring of FXa inhibitors and the indole moiety of Trp214). Finally, the possible implications of the obtained results regarding pathologies such as hypoalbuminemia are briefly discussed.

Abstract WP203: Effects Of Oral Factor Xa Inhibitors On Ischemic Stroke In Patients Without Atrial Fibrillation: Systematic Review Of Randomized Clinical Trials

Background and Purpose: To assess the effects of oral factor Xa inhibitors for prevention of ischemic stroke in patients without a history of atrial fibrillation. Methods: Systematic review of randomized clinical trials (RCTs) testing oral factor Xa inhibitors in patients without a history of atrial fibrillation that reported ischemic stroke. Results: Twenty-two RCTs reported 1194 ischemic strokes: five RCTs testing apixaban (75 ischemic strokes), one RCT each testing betrixaban (52 ischemic strokes) and edoxaban (1 ischemic stroke), and 15 RCTs testing rivaroxaban (1066 ischemic strokes). Three double-blinded RCTs reported statistically significant reductions in ischemic stroke: Two comparing rivaroxaban 2.5mg twice daily versus placebo in patients with cardiovascular disease (COMPASS, hazard ratio (HR) 0.51, 95%CI 0.38-0.68 and COMMANDER-HF, HR 0.64, 95%CI 0.43-0.95), and a third trial compared betrixaban 80mg daily with enoxaparin followed by placebo in patients hospitalized for acute medical illness (APEX, relative risk 0.53, 95%CI 0.30-0.94). Compared with aspirin, rivaroxaban 5mg twice daily significantly reduced ischemic stroke in patients with chronic atherosclerosis in COMPASS (HR 0.69, 95%CI 0.53-0.90), but in contrast rivaroxaban 15mg daily vs. aspirin showed no reduction in ischemic stroke in patients with recent embolic stroke in the large NAVIGATE ESUS trial (HR 1.01, 95%CI 0.81-1.26). Comparisons from 18 other RCTs were inconclusive, although trends consistently showed numerically fewer ischemic strokes among those assigned oral fXa inhibitors compared with placebo. Conclusions: There is convincing evidence that oral factor Xa inhibitors reduce ischemic stroke in patients without a history of atrial fibrillation. The strongest evidence was for rivaroxaban 2.5mg twice daily when given with aspirin in patients with cardiovascular disease. Reduction in ischemic stroke by other dosages, by other factor Xa inhibitors, and when given without antiplatelet agents is less certain. The contribution of atrial fibrillation-related stroke unlikely accounts for the observed reductions.

Differential Neutralization of Apixaban, Betrixaban, Edoxaban, and Rivaroxaban by Andexanet Alfa as Measured by Whole Blood Thromboelastographic Analysis.

The available oral anti-Xa agents are routinely used for the management of thrombotic disorders. A molecularly modified recombinant coagulation FXa, also known as Andexanet Alfa (AA), that has been developed as an antidote to neutralize the bleeding effects of oral FXa inhibitors, such as Apixaban and Rivaroxaban. This study utilized thromboelastography (TEG 5000 Hemostasis System), to investigate the neutralizing effects of AA at different concentrations of oral FXa inhibitors measuring such parameters as R-Time, K-Time, Angle, and Max Amplitude (MA). Apixaban, Betrixaban, Edoxaban, and Rivaroxaban were obtained commercially in powdered form. Each of these drugs was supplemented with freshly drawn whole citrated blood at a concentration of 1 μg/mL. And subsequently mixed with AA at 50 or 100 μg/mL. At a concentration of 1 μg/mL, all FXa inhibitors produced variable anticoagulant effects in the order of Edoxaban > Betrixaban > Rivaroxaban > Apixaban. AA at 100 μg/mL produced a complete neutralization of these inhibitors whereas at 50 μg/mL relatively weaker neutralization as measured by various parameters. These results suggest that regardless of the variable anticoagulant effects exhibited by the FXa Inhibitors, AA at FC = 100 μg/mL fully neutralized these agents as measured by the TEG parameters. AA was shown to be more effective in neutralizing Betrixaban and least effective in Apixaban. The neutralization of various FXa inhibitors was dose and donor-dependent warranting dosage adjustment for optimal outcomes.

Factor Xa inhibitors: critical considerations for clinical development and testing

The selection of factor (F) X and its activated protease FXa for targeted inhibition to prevent and treat thrombotic conditions is based on an understanding of coagulation biochemistry, sequential steps that occur on tissue factor bearing cells and the interface of coagulation proteins, platelets, mononuclear cells and the nuclear constituents of inflammatory cells. The goal for developing direct oral FXa inhibitors was to achieve rapid, selective, predictable, safe and effective anticoagulation across a broad group of patients expected to derive benefit. The history and development in patient care are exemplars of knowledge, translation and collaboration between the public and private sectors.

67 Clinical Outcomes of Oral Factor Xa-Inhibitor Associated Gastrointestinal Bleeds Treated with or without 4F-PCC among US Medicare Patients

Prior to the availability of a specific oral factor Xa (fXa) inhibitor reversal agent, off-label replacement agents including 4-factor prothrombin complex concentrates (4F-PCC) were used to treat life-threatening hemorrhage including gastrointestinal (GI) bleeds. We compared outcomes of US Medicare patients hospitalized for oral fXa-inhibitor associated GI bleeds treated with or without 4F-PCC. We utilized a 20% random sample of Medicare fee-for-service beneficiaries who had Part A and D (medication) coverage. We included adult (≥ 18 years) patients experiencing their first hospitalization (index event) for major GI bleeding identified via ICD9/10 codes, between 10/2013 and 9/2017. Serious GI bleeds were defined by ICD9/10 codes developed based on the Cunningham and Joos et al. algorithms. Inclusion criteria were continuous medical and prescription coverage and a claim for apixaban, edoxaban, or rivaroxaban during the 6-months prior to the index event. Patients were divided into two cohorts based on treatment with or without 4F-PCC during the index GI bleed hospitalization. Fresh frozen plasma, cryoprecipitates, packed red blood cells, activated PCC, recombinant factor VIIa, desmopressin acetate, tranexamic acid and vitamin K administration were allowed in either cohort. Use of more than one agent in either group was defined as concomitant therapy. Endpoints included the 1) intensive care unit (ICU) treatment, 2) total hospital length of stay, 3) discharge disposition and 4) 30-day readmission. Among 8,169 patients hospitalized for GI bleeding, 62 (0.75%) patients were treated with 4F-PCC. Mean age was 78.4 years and 58.8% were female. Because of the low numbers the two groups could not be propensity matched. Overall in-hospital mortality was 1.7%. Concomitant therapy was 56.5% 4F-PCC-treated vs. 45.3% non-4F-PCC patients. Incidence of ICU was 71.0% 4F-PCC vs 39.4% non-4F-PCC patients. Median (Interquartile range) hospital stay was 6 (4, 10) for 4F-PCC vs 5 (4, 6) days non-4F-PCC patients. Discharge disposition home was 45.2% for 4F-PCC vs 57.8% non-4F-PCC patients. All-cause readmission at 30 days was 18.0% for 4F-PCC vs 18.8% non-4F-PCC patients (Table 1). In a large descriptive analysis of hospitalized Medicare patients with oral fXa-inhibitor associated major GI bleeds, those treated with 4F-PCC had higher concomitant medication use, longer LOS, and lower frequency of discharge to home. Whether this is a result of off-label use of 4F-PCC or other variables in these complex patients is uncertain and deserves further study.Table 1Characteristics of US Medicare patients hospitalized for oral fXai-associated GI bleedsAll (N=8,169)4F-PCC Treated (N=62)Non-4F-PCC Treated (N=8,107)Indication for fXa inhibitors (%)Atrial fibrillation75.879.075.8Deep vein thrombosis/venous thromboembolism24.025.824.0Any concomitant medication (%)45.456.545.3Packed RBCs (%)43.853.243.7Characteristics of index hospitalizationLength of stay in days (Median; interquartile range)5.0 (4.0-6.0)6.0 (4.0-10.0)5.0 (4.0-6.0)Index hospitalization as ICU (%)39.771.039.4Discharge status (%)Home57.745.257.8Discharge to a location other than home (including in-hospital death)*42.354.842.2*Data on in-hospital death alone was not reported by treatment cohort in order to comply with the Centers for Medicare and Medicaid Services’ Cell Size Suppression Policy Open table in a new tab

Emergent reversal of oral factor Xa inhibitors with four-factor prothrombin complex concentrate.

Controversy exists regarding first-line use of the recently approved reversal agent andexanet alfa due to limitations of the ANEXXA-4 study, thrombotic risks, and high medication acquisition cost. The purpose of this study was to evaluate the safety and effectiveness of 4F-PCC for the reversal of emergent oral fXa inhibitor-related bleeding. Furthermore, we aimed to evaluate a subgroup using strict ANNEXA-4 patient selection criteria. This was a retrospective study conducted utilizing chart review of adult patients that received 4F-PCC for oral fXa inhibitor-related bleeding. The primary endpoint was the rate of clinical success defined as achieving excellent or good hemostatic effectiveness following the administration of 4F-PCC. Secondary endpoints included in-hospital mortality and arterial/venous thromboembolism, and cost compared with andexanet alfa. A total of 119 patients were included, with 83 patients in the ANNEXA-4 criteria subgroup. Eighty-five of the 119 patients (71%) required reversal due to intracranial bleeding. Prior to reversal, 70 patients (59%) were taking apixaban and 49 patients (41%) were taking rivaroxaban. Clinical success was achieved in 106 of 119 patients (89%) and 74 of 83 patients (90%) in the strict criteria subgroup. Three of 119 patients (2.5%) had a thrombotic event during hospital stay and the overall mortality rate was 13%. The average cost increase of andexanet alfa compared to 4F-PCC would have been $29,500 per patient. Administration of 4F-PCC for the reversal of oral fXa inhibitors was effective with relatively low thrombotic risk. Further direct prospective comparison of 4F-PCC to andexanet alfa is warranted.

Coagulation signaling and cancer immunotherapy

The last decades have delineated many interactions of the hemostatic system with cancer cells that are pivotal for cancer-associated thrombosis, angiogenesis and metastasis. Expanding evidence shows that platelets, the tissue factor pathway, and proteolytic signaling involving protease-activated receptors (PARs) are also central players in innate and adaptive immunity. Recent studies in immune-competent mice have uncovered new immune-evasive roles of coagulation signaling networks in the development and growth of different preclinical tumor models. Tumor-type specific PAR1 signaling facilitates the escape from immune surveillance by cytotoxic T cells. In addition, tumor-associated macrophages produce factor X (FX) and cell autonomous FXa-PAR2 signaling emerges as a central mechanism for tumor-promoting macrophage polarization in the tumor microenvironment. Pharmacological targeting of this signaling pathway with tissue penetrating oral FXa inhibitor reprograms macrophage phenotypes, enhances tumor antigen presentation, and expands tumor-killing cytotoxic lymphocytes. Importantly, by specifically targeting innate immune cells, the oral FXa inhibitor rivaroxaban synergizes with checkpoint inhibitor therapy in enhancing antigen-specific antitumor immunity. In similar experiments, anticoagulation with heparin is inefficient to block extravascular coagulation signaling. Thus, antithrombotic therapy with oral FXa inhibitors may contribute to reversing tumor immune-evasive mechanisms and enhance the clinical outcome of targeted immuno-therapy regimens.

Trial of Andexanet in Patients Receiving an Oral FXa Inhibitor Who Require Urgent Surgery

Trial of Andexanet in Patients Receiving an Oral FXa Inhibitor Who Require Urgent Surgery

Prothrombin Complex Concentrates (PCCs) Have Limited Effect on TF-Initiated Thrombin Generation in FXa Inhibitor-Anticoagulated Plasma: In Vitro Comparison between Direct Reversal By Andexanet Alfa and “Work Around” By PCCs

Introduction: Direct oral FXa inhibitors (DOACs, e.g., apixaban [Apix] and rivaroxaban [Riva]) exert their anticoagulant activity by specifically targeting FXa with subnanomolar affinities. Reversal of DOAC-induced anticoagulation can be achieved by immediate and effective sequestration of the inhibitors to relieve inhibition of FXa activity and restore normal thrombin generation (TG). Andexanet alfa (AnXa) is a modified human FXa protein approved in the US and EU for patients treated with Apix or Riva, when reversal of anticoagulation is needed due to life-threatening bleeding. Factor replacement therapies (e.g., 4 factor-PCCs [4F-PCCs]) have demonstrated clinical benefit for vitamin K antagonist-anticoagulated patients by replacing factors II, VII, IX, and X rendered deficient by the anticoagulant. In contrast, the effectiveness and clinical benefit of 4F-PCCs are unclear for reversing DOACs because PCCs do not affect anti-FXa activity. In the present study, we evaluated the effect of individual coagulation factors (FIX, FX, FII), as well as 4F-PCCs on TF-initiated TG (TF-TG) in human platelet poor plasma (PPP), with or without Apix or Riva, and compared it with AnXa under the same conditions. Methods: TF-TG was measured using a calibrated automated thrombogram (CAT, 5 pM TF, Diagnostica Stago). Purified plasma proteins (FIX, FX, FII), PPP, and a commercially available 4F-PCC (Kcentra/Beriplex) were used. 4F-PCC was used with or without pretreatment with the heparinase cup (Haemonetics Corporation) at room temperature for 30 minutes. TF-TG was measured in PPP spiked with purified plasma proteins, 4F-PCC (0-1.0 IU/mL), 4F-PCC + Riva or Apix (0-250 ng/mL), or PPP with AnXa (0-4 μM) + Riva or Apix (0-2000 ng/mL). For comparison, reversal of warfarin anticoagulation by 4F-PCC was performed under similar conditions using individual plasma from warfarin-treated patients (INR = 1.5-6.9) spiked with 4F-PCC (0-1.0 IU/mL). Five CAT parameters were collected, and the endogenous thrombin potential (ETP) and Peak thrombin (Peak) were used for the comparisons. Results: In normal PPP, addition of FX (up to 1.0 IU/mL) had minimal effect on TF-TG parameters, whereas FIX (1.0 IU/mL) mainly increased Peak (~60%). In contrast, FII (1.0 IU/mL) increased both the ETP (~2x) and Peak (~50%). These observations are consistent with the relative plasma concentration and affinity of these coagulation factors as the substrate for the respective enzyme complexes, demonstrating the predominant role of prothrombinase activity in the TF-TG. Addition of 4F-PCC (1.0 IU/mL, equivalent to the therapeutic dose of 50 IU/kg) to PPP caused similar increases of ETP (~2.4x) and Peak (~40%). However, individual plasma protein (FIX, FX, FII) or 4F-PCC up to 1.0 IU/mL was unable to restore normal ETP or Peak in the presence of Apix or Riva (75-250 ng/mL). ETP and Peak increased only when the inhibitor concentrations were sufficiently low (<75 ng/mL), leading to less inhibition of FXa and increased FIIa formation, likely due to an increased FII level from 4F-PCC addition. Similar results were obtained with or without preincubation of 4F-PCC with heparinase prior to spiking in plasma. In contrast, AnXa (0-4.0 μM) dose-dependently reversed Apix- and Riva- (up to 2000 ng/mL) induced inhibition of TF-TG. In the same TF-TG assay, 4F-PCC dose-dependently reversed warfarin anticoagulation in warfarin-treated patients' plasma spiked with 4F-PCC. Conclusions: Specific reversal agents are effective for restoration of TF-TG by either direct sequestration of the FXa inhibitor and restoration of FXa activity (in the case of AnXa with FXa inhibitors) or by replacement of inactive factors due to warfarin treatment (in the case of 4F-PCCs). Addition of coagulation factors or use of 4F-PCCs had limited effect on restoration of normal TG in the presence of a FXa inhibitor. These data demonstrate that AnXa can normalize TF-TG over a wide range of FXa inhibitor concentrations (0-2000 ng/mL), in particular over the range observed in bleeding patients in the ANNEXA-4 study, where 77% of bleeding patients had inhibitor levels ≥75 ng/mL at baseline. In contrast, 4F-PCCs were only able to restore normal TF-TG at low inhibitor levels (<75 ng/mL). These in vitro data are consistent with ANNEXA-4, where AnXa demonstrated >90% immediate reversal of anti-FXa activity and >80% hemostatic efficacy with a wide range of FXa inhibitor levels and multiple bleed types. Disclosures Lu: Portola Pharmaceuticals: Employment, Equity Ownership. Lin:Portola Pharmaceuticals, Inc: Employment, Equity Ownership. Bui:Portola Pharmaceuticals, Inc.: Employment, Equity Ownership. Curnutte:Portola Pharmaceuticals: Employment, Equity Ownership. Conley:Portola Pharmaceuticals: Employment, Equity Ownership.

Lupus anticoagulant diagnosis in patients receiving direct oral FXa inhibitors at trough levels: A real-life study.

Direct oral factor Xa inhibitors (xabans) induce false positive results for lupus anticoagulant (LA) diagnosis. Consequently, it is suggested not to perform LA testing in xabans patients although it may be useful in selected patients. In this monocentric study, we evaluated xabans impact at trough levels (ie, just before the next intake) on LA diagnosis in treated patients using dilute Russell viper venom time (dRVVT) and two LA sensitive activated partial thromboplastin time (aPTT). Sixty patients receiving rivaroxaban (30) or apixaban (30) were included. Plasma concentrations were measured using specific anti-Xa assays. LA testing was performed using one dRVVT (LAC-Screening® /Confirm® ; Siemens) and two LA sensitive aPTT-based assays (Hemosil® Silica Clotting Time (SCT) Screen/Confirm; Werfen and Dade® Actin® Factor Sensitivity FSL/FS (Actin F); Siemens). Median [min-max] concentrations were 23 [<18-68] for rivaroxaban and 42ng/mL [19-99] for apixaban. dRVVT was positive in 93% of rivaroxaban and 40% of apixaban samples. SCT was positive in 40 and 30% and Actin F in 17 and 20% of samples respectively. Xabans affected more significantly dRVVT than aPTT-based assays (P<.001) with less false positive results with apixaban than with rivaroxaban samples irrespective of the assay used. lupus anticoagulant diagnosis in rivaroxaban and apixaban samples drawn at trough levels remains questionable whenever positive results are obtained. If LA testing in apixaban samples might be useful to rule-out LA using dRVVT and/or aPTT-based assays, the wide majority of rivaroxaban samples would give false positive results.

New progress in laboratory monitoring of the rivaroxaban therapy

Rivaroxaban is an oral direct factor FⅩa inhibitor with predictable pharmacokinetics and no routine monitoring, but the laboratory tests can help to assess the safety and effectiveness of rivaroxaban. The laboratory tests for rivaroxaban include liquid chromatography tandem mass spectrometry (LC-MS/MS), prothrombin time(PT), anti factor Ⅹ a activity(anti-Ⅹ a), thromboelastography(TEG) and rotational thromboelastography(ROTEM). LC-MS/MS can be used to quantitatively detect the blood of rivaroxaban with good specificity and sensitivity, but the instrument is expensive, technologically complex, and lack standardization, so it belongs to the laboratory developed tests(LDTs). Because of insufficient data of TEG and ROTEM, their clinical performance still needs to be verified. PT can detect treatment concentration of rivaroxaban, which can be used as a primary screening method to identify the overdose and the risk of severe bleeding, but the sensitivity of different reagents is different; anti-F Ⅹ a test can sensitively reflect the change of blood of rivaroxaban, and its clinical efficacy is similar to LC-MS/MS, and therefore it can be used as an effective method to guide doctors to use drugs rationally. Key words: Rivaroxaban; Chromatography, liquid; Tandem mass spectrometr; Prothrombin time; Factor Ⅹa; Clinical laboratory techniques

Safety, efficacy, and cost of four-factor prothrombin complex concentrate (4F-PCC) in patients with factor Xa inhibitor-related bleeding: a retrospective study.

Oral factor Xa (fXa) inhibitor-related bleeding is a concerning drug safety problem. There is considerable controversy surrounding available reversal strategies. The recently approved reversal agent andexanet alfa has limited data, an unclear safety profile, and imparts a substantial financial burden. This has led to the off-label use of four-factor prothrombin complex concentrates (4F-PCC) for this indication. This study aimed to assess the safety and efficacy of 4F-PCC for the management of major bleeding related to oral fXa inhibitors. This observational, retrospective study included adult patients admitted from 2014 to 2018 who received 4F-PCC (Kcentra®) for fXa inhibitor-related major bleeding. Efficacy was assessed using criteria described by Sarode et al. Secondary outcomes included the incidence of thromboembolism, mortality, and a cost analysis comparing 4F-PCC to andexanet alfa for reversal of oral fXa inhibitors. Thirty-one patients received 4F-PCC for major bleeding associated with apixaban (55%) or rivaroxaban (45%). Intracranial hemorrhage (58%) and pericardial effusion (16%) accounted for the majority of bleeding events. Most patients received a single weight-based 4F-PCC dose of 25 units/kg (38.7%) or 50 units/kg (51.6%). Effective hemostasis was achieved in 80.6% of patients. Five patients (16%) died due to acute bleeding and no thromboembolic events were observed. Administration of 4F-PCC was effective for most patients requiring emergent reversal of anticoagulation with apixaban or rivaroxaban and was associated with a low risk of thromboembolic events. Considerable cost differences limit the use of andexanet alfa and may warrant further study of 4F-PCC for fXa inhibitor reversal.

Linking Endogenous Factor Xa Activity, a Biologically Relevant Pharmacodynamic Marker, to Edoxaban Plasma Concentrations and Clinical Outcomes in the ENGAGE AF-TIMI 48 Trial.

We previously reported exogenous antifactor Xa (FXa) activity as a pharmacokinetic surrogate marker for edoxaban plasma concentrations. Inhibition of endogenous FXa activity is a more biologically relevant pharmacodynamic measure of edoxaban activity. Here we describe the value of endogenous FXa activity as a pharmacodynamic marker linking edoxaban concentrations and clinical outcomes in the ENGAGE AF-TIMI 48 trial (Effective Anticoagulation With Factor Xa Next Generation in Atrial Fibrillation-Thrombolysis in Myocardial Infarction Study 48). In ENGAGE AF-TIMI 48, edoxaban was administered in higher dose (60/30 mg QD) and lower dose (30/15 mg QD) regimens. Both regimens incorporated a 50% dose reduction in patients with characteristics known to increase edoxaban concentration. Pharmacokinetic-pharmacodynamic modeling was performed in a subgroup of 3029 patients who had samples collected for endogenous FXa activity (measured using an assay after endogenous FX was activated with Russell viper venom). Endogenous FXa activity decreased with increasing edoxaban concentrations of ≤440 ng/mL, indicating that inhibition of endogenous FXa activity is saturated above this concentration threshold. Baseline endogenous FXa activity averaged 92.1±20.9% (relative to normal control samples) and was lower with older age, with lower body weight, and in male patients. Model-predicted 24-hour average percentages of inhibition of endogenous FXa activity were 35.8±5.18, 29.1±3.92, 21.9±3.80, and 16.4±2.70 for the higher dose edoxaban regimen 60 mg, dose-reduced higher dose edoxaban regimen 30 mg, lower dose edoxaban regimen 30 mg, and dose-reduced lower dose edoxaban regimen 15 mg groups, respectively. A greater average percentage of inhibition of endogenous FXa activity was associated with a lower incidence of ischemic stroke or systemic embolism and a higher risk of major bleeding ( P<0.001). In a typical subject, the predicted risks for the 10th and 90th percentiles of inhibition of endogenous FXa activity were 1.04% and 0.57% for incidence of ischemic stroke or systemic embolism and 1.35% and 2.33% for major bleeding, respectively. The extent of inhibition of endogenous FXa activity is influenced by edoxaban dosing and clinical characteristics, and it is associated with both antithrombotic benefit and risk of bleeding. This approach of linking endogenous FXa activity to clinical outcomes may be used to guide dose selection in future clinical trials, monitor patients in certain clinical scenarios, or refine the doses of oral FXa inhibitors in patients who require precise anticoagulation therapy. URL: https://www.clinicaltrials.gov . Unique identifier: NCT00781391.

Effect of a new inhibitor of factor Xa zifaxaban, on thrombosis in the inferior vena cava in rabbits.

In recent years, oral factor Xa inhibitors have become a research focus as anticoagulant drugs. Zifaxaban is the first oral FXa inhibitor to enter clinical trials in China. The aim of this study was to determine the inhibitory effect of zifaxaban on thrombosisthrough a model ofinferior vena cava (IVC) thrombosis in rabbits. IVC thrombosis model was established by electrical injury and stenosis, and zifaxaban was administered (p.o.) for 5 consecutive days, then coagulation indicators and bleeding were observed. The results showed that zifaxaban had obvious inhibitory effects on FXa, and had a significant inhibitory effect on IVC thrombosis induced by electrical damage and stenosis. The effect of zifaxaban was similar to that of rivaroxaban, but the bleeding side-effects of zifaxaban were less severe than those of rivaroxaban. Zifaxaban could prolong the prothrombin time and activated partial thromboplastin time of plasma similar to that of other oral FXa inhibitors. Zifaxaban had a significant inhibitory effect on FXa, but it had no obvious effect on other coagulation factors, major anticoagulant factors or fibrinolytic indices. Our results suggest that zifaxaban had specific inhibitory effects on FXa and inhibited IVC thrombosis in rabbits with its hemorrhagic effect was less than that of rivaroxaban. Zifaxaban is ecpected to be developed as a new drug for the prevention of deep venous thrombosis, providing more medication options for patients with such disease, more research is required to support it in the future.

Coagulation Factor Xa and Protease-Activated Receptor 2 as Novel Therapeutic Targets for Diabetic Nephropathy.

The role of hypercoagulability in the pathogenesis of diabetic nephropathy (DN) remains elusive. We recently reported the increased infiltration of macrophages expressing tissue factor in diabetic kidney glomeruli; tissue factor activates coagulation factor X (FX) to FXa, which in turn stimulates protease-activated receptor 2 (PAR2) and causes inflammation. Here, we demonstrated that diabetes mellitus increased renal FX mRNA, urinary FXa activity, and FX expression in glomerular macrophages. Administration of an oral FXa inhibitor, edoxaban, ameliorated DN with concomitant reductions in the expression of PARs (Par1 and Par2) and of proinflammatory and profibrotic genes. Diabetes mellitus induced PAR2, and lack of Par2 ameliorated DN. FXa or PAR2 agonist increased inflammatory cytokines in endothelial cells and podocytes in vitro. We conclude that enhanced FXa and PAR2 exacerbate DN and that both are promising targets for preventing DN. Alleviating inflammation is probably more important than inhibiting coagulation per se when treating kidney diseases using anticoagulants.

X-ray powder diffraction data for 5-chloro-N-(4-(5,6-dihydro-3-(4-morpholinyl)-2-oxo-1(2H)-pyridinyl)phenyl)-pentanamide, C20H26ClN3O3

Apixaban (Eliquis®) is a novel oral pyrazole-based direct FXa inhibitor. The title compound is an intermediate in the synthesis of the anticoagulant, Apixaban. X-ray powder diffraction data for the title compound, are reported [a = 9.511(5) Å, b = 18.539(2) Å, c = 5.645(3) Å, α = 90°, β = 101.813(1)°, γ = 90°, unit-cell volume V = 974.28 Å3, Z = 2, ρcal = 1.336 g cm−3, and space group P2]. All measured lines were indexed and are consistent with the P2 space group. No detectable impurities were observed.