scholarly journals Andexanet Alpha Differentially Neutralizes the Anticoagulant, Antiprotease and Thrombin Generation Inhibitory Effects of Unfractionated Heparin, Enoxaparin and Fondaparinux

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1158-1158
Author(s):  
Fakiha Siddiqui ◽  
Alfonso J Tafur ◽  
Debra Hoppensteadt ◽  
Jeanine Walenga ◽  
Walter Jeske ◽  
...  
Keyword(s):  
Research Funding ◽  
Thrombin Generation ◽  
Surrogate Marker ◽  
Factor Xa ◽  
Bleeding Complications ◽  
Dependent Manner ◽  
Thrombin Time ◽  
Inhibitory Effects ◽  
Biologic Effects ◽  
Inhibition Of Thrombin

Introduction: Andexanet Alpha (Coagulation factor Xa recombinant, inactivated Zh-zo; AA, Portola Pharmaceuticals) is a recombinant factor Xa decoy protein which is designed to reverse the effects of apixaban and rivaroxaban and is approved for the control of bleeding complications associated with their use. The molecular modification in this recombinant protein involves the substitution of serine active site by alanine and the removal of the gamma-carboxyglutamic acid (GLA) domain to restrict its assemblage into prothrombinase complex. Beside the reversal of the effects of anti-Xa agents AA is also reported to neutralize the biologic effects of heparin and related drugs. Assay dependent variations in the neutralization profile of various factor Xa inhibitors by andexanet has been recently reported https://doi.org/10.1177/1076029619847524. Since heparin and related drugs also mediate their biologic actions by inhibiting factor Xa via AT complexation, it is hypothesized that AA may also inhibit their biologic effects as measured in various laboratory assays. It is the purpose of this study is to compare the relative neutralization profile of heparin (UFH), a low molecular weight heparin, enoxaparin (E) and a chemically synthetic pentasaccharide, Fondaparinux (F) by AA. Materials and Methods: API versions of UFH, E and F were commercially obtained in powdered forms and dissolved in saline at a working dilution of 1mg/ml. AA was dissolved in saline to obtain a 10mg/ml working solution. The anticoagulant profile of UFH, E and F was studied using the activated partial thromboplastin time (APTT) and thrombin time (TT) in a concentration range of 0 - 10 ug/ml in pooled human plasma. The anti-Xa and anti-IIa studies were carried out in amidolytic assays in the same concentration range. The thrombin generation inhibition was studied using calibrated automated thrombin generation systems (CAT, Diagnostica Stago). The effect of AA on the reversal of the anticoagulant and anti-protease and thrombin generation effects of each of these agents were studied by supplementing this agent at 100 ug/ml. The results are compared to determine the difference between pre and post AA neutralization settings. Results: All agents produce a concentration dependent effect in the anticoagulant and anti-protease assays with the exception of F which showed mild anticoagulant effects, and very weak anti-IIa actions and strong anti-Xa activity. In the anti-Xa assay the IC-50 for UFH was 2.1ug/ml (0.13 um), E 4.3 ug/ml (0.95 um) and F 0.7 ug/ml (0.41 um) upon supplementation of AA the IC50s for UFH was increased to 5 ug/ml (0.31 um) and for E 5 ug/ml (1.11 um). However, there was no neutralization of the anti-Xa effects of the F by AA and the IC50 remained the same for both pre and post andexxa studies. The anticoagulant effects of UFH as measured by aPTT and TT was strongly neutralized whereas E was only partially neutralized in the aPTT assay and almost completely neutralized in the thrombin time assay. At concentrations of up to 10 ug/ml F did not produced any significant anticoagulant effects, both in the presence and absence of AA. In the thrombin generation inhibition assays, UFH produced a complete inhibition of thrombin generation which was completely reversed by AA. Although both E and F produced strong inhibition of thrombin generation, AA did not completely neutralize these effects. The results are tabulated on table 1 for the studies carried out at 10 ug/ml of UFH, E and F. Conclusion: These results indicate that AA is capable of differentially neutralizing anticoagulant and anti-protease effects of UFH in an assay dependent manner. However, AA is incapable of neutralizing the anti-Xa effects of E and F. This may be due to the relatively differential affinities of enoxaparin and fondaparinux AT complex to factor Xa rendering it inhibited in the presence of AA. These studies also demonstrate that the primary surrogate marker anti-Xa activity for measuring the activities of anti-Xa agents is not proportional to the anticoagulant and thrombin generation inhibitory effects of these agents. A global clotting assay may be a better indication of the biologic effects of these agents and their reversal by AA. Disclosures Tafur: Recovery Force: Consultancy; Janssen: Other: Educational Grants, Research Funding; BMS: Research Funding; Idorsia: Research Funding; Daichi Sanyo: Research Funding; Stago: Research Funding; Doasense: Research Funding.

Oral Anti-Factor Xa and Factor IIa Agent Mediated Inhibition Of Tissue-Factor Mediated Generation Of Thrombin In Prothrombin Complex Concentrates

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4810-4810
Author(s):  
Daneyal Syed ◽  
Debra Hoppensteadt ◽  
Daniel Kahn ◽  
Job Harenberg ◽  
Jawed Fareed
Keyword(s):  
Tissue Factor ◽  
Oral Anticoagulant ◽  
Thrombin Generation ◽  
Conflicts Of Interest ◽  
Factor Xa ◽  
Onset Time ◽  
Thrombin Inhibitors ◽  
Bleeding Complications ◽  
Prothrombin Complex Concentrates ◽  
Inhibition Of Thrombin

Introduction Several oral anti-factor IIa and factor Xa agents have recently been developed. These include the thrombin inhibitors Ximelagatran/Melagatran (M) and Dabigatran Etexilate/Dabigatran (D), which require endogenous conversion to the active agents D and M. The factor Xa inhibitors, Rivaroxaban (R) and Apixaban (A), are anti-Xa agents that do not require any endogenous activation. Ximelagatran was withdrawn from the market due to adverse reactions. Dabigatran, Rivaroxaban, and Apixaban are approved for various clinical indications. Antagonism of the anticoagulant effect may be required in bleeding complications. Contradictory results were reported for the efficacy of various prothrombin complex concentrates (PCCs) with these new oral anticoagulants (NOACs). The purpose of this study was to determine the differences in the thrombin generation inhibitory profiles of the newer oral anticoagulant agents. Methods Commercially available PCCs namely Octaplex and Beriplex, were used as a source of Factors II, VII, IX and X. To investigate the effect of each of these agents, a working solution of 1U/ml of both PCCs were supplemented in a graded concentration of 0-1250ng/ml with M, D, R and A. Thrombin generation studies were carried out using a thromboplastin activator (RC High, Technoclone Vienna, Austria). Total thrombin generated was measured in terms of nM’s. The IC-50 for each agent was calculated individually. The time course of thrombin generation was also measured following the kinetic profiles and AUC. Results Dabigatran and Melagatran produced relatively weaker inhibition of thrombin generation with the IC-50 values ranging from 410-110ng/ml in Beriplex and 350-1120ng/ml in Octaplex. Both Rivaroxaban and Apixaban produced strong inhibition of thrombin generation, with the IC-50 ranging from 58-62ng/ml in Octaplex; whereas, in Beriplex these values ranged from 48-50ng/ml. The onset time for thrombin generation and total thrombin formation was concentration dependent. The kinetics of thrombin generation with A and R were distinct from D and M. At concentrations below 310ng/ml the total amount of thrombin generated was comparable to the control; however, its formation was delayed. In both systems, D exhibited the weakest thrombin generation inhibitory potential. While the onset time of thrombin generation was delayed at concentrations below 310ng/ml the levels were comparable to or higher than the control. Discussion This data suggests that PCC’s such as Octaplex and Beriplex can be used to generate thrombin and it’s inhibition by new oral anticoagulant drugs. Octaplex generates much higher amount of thrombin than Beriplex at equivalent units. These results also show that in comparison to the oral anti-Xa agents, the oral anti-IIa agents are relatively weaker inhibitors of thrombin generation. These studies also suggest that the differential inhibition of the generation of thrombin through tissue factor by the anti-Xa and IIa agents may contribute to the potential neutralization profile of PCC’s for these drugs. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Assay-Based Differentiation in the Neutralization Profile of Unfractionated Heparin, Enoxaparin, and Fondaparinux by Andexanet Alfa

2020 ◽  
Vol 26 ◽  
pp. 107602961989512
Author(s):  
Fakiha Siddiqui ◽  
Alfonso Tafur ◽  
Emily Bontekoe ◽  
Omer Iqbal ◽  
Walter Jeske ◽  
...  
Keyword(s):  
Unfractionated Heparin ◽  
Thrombin Generation ◽  
Factor Xa ◽  
Dependent Manner ◽  
Thrombin Time ◽  
Clotting Time ◽  
Activated Clotting Time ◽  
Complete Reversal ◽  
Andexanet Alfa ◽  
Partial Neutralization

Andexanet alfa is a recombinant factor Xa decoy protein, designed to reverse bleeding associated with oral anti-Xa agents. Andexanet alfa is also reported to neutralize the effects of heparin-related drugs. This study focused on the neutralization profiles of unfractionated heparin (UFH), enoxaparin, and, a chemically synthetic pentasaccharide, fondaparinux by andexanet alfa. Whole blood clotting studies were carried out using thromboelastography (TEG) and activated clotting time (ACT). The anticoagulant profile of UFH, enoxaparin, and fondaparinux was studied using the activated partial thromboplastin time (aPTT), thrombin time (TT), and amidolytic anti-Xa, and anti-IIa methods. Thrombin generation inhibition was studied using the calibrated automated thrombogram system. Reversal of each of these agents was studied by supplementing andexanet alfa at 100 µg/mL. In the TEG, andexanet alfa produced almost a complete reversal of the anticoagulant effects of UFH and enoxaparin; however, it augmented the effects of fondaparinux. In the ACT, aPTT, and TT, UFH produced strong anticoagulant effects that were almost completely neutralized by andexanet alfa. Enoxaparin produced milder anticoagulant responses that were partially neutralized, whereas fondaparinux did not produce any sizeable effects. In the anti-Xa and anti-IIa assays, UFH exhibited partial neutralization whereas enoxaparin and fondaparinux did not show any neutralization. All agents produced varying degrees of the inhibition of thrombin generation, which were differentially neutralized by andexanet alfa. These results indicate that andexanet alfa is capable of differentially neutralizing anticoagulant and antiprotease effects of UFH and enoxaparin in an assay-dependent manner. However, andexanet alfa is incapable of neutralizing the anti-Xa effects of fondaparinux.

Inhibition of Thrombin Generation in Plasma by Inhibitors of Factor Xa

1997 ◽  
Vol 78 (04) ◽  
pp. 1215-1220 ◽  
Author(s):  
D Prasa ◽  
L Svendsen ◽  
J Stürzebecher
Keyword(s):  
Thrombin Generation ◽  
Factor Xa ◽  
Thrombin Inhibitors ◽  
Thrombin Inhibitor ◽  
Thrombin Generation Assay ◽  
Ic50 Values ◽  
Fxa Inhibitor ◽  
20 Nm ◽  
Tick Anticoagulant Peptide ◽  
Inhibition Of Thrombin

SummaryA series of inhibitors of factor Xa (FXa) were investigated using the thrombin generation assay to evaluate the potency and specificity needed to efficiently block thrombin generation in activated human plasma. By inhibiting FXa the generation of thrombin in plasma is delayed and decreased. Inhibitor concentrations which cause 50 percent inhibition of thrombin generation (IC50) correlate in principle with the Ki values for inhibition of free FXa. Recombinant tick anticoagulant peptide (r-TAP) is able to inhibit thrombin generation with considerably low IC50 values of 49 nM and 37 nM for extrinsic and intrinsic activation, respectively. However, the potent synthetic, low molecular weight inhibitors of FXa (Ki values of about 20 nM) are less effective in inhibiting the generation of thrombin with IC50 values at micromolar concentrations.The overall effect of inhibitors of FXa in the thrombin generation assay was compared to that of thrombin inhibitors. On the basis of similar Ki values for the inhibition of the respective enzyme, synthetic FXa inhibitors are less effective than thrombin inhibitors. In contrast, the highly potent FXa inhibitor r-TAP causes a stronger reduction of the thrombin activity in plasma than the most potent thrombin inhibitor hirudin.

scholarly journals Effect ofToona microcarpaHarms Leaf Extract on the Coagulation System

2014 ◽  
Vol 2014 ◽  
pp. 1-7 ◽  
Author(s):  
Hao Chen ◽  
Min Jin ◽  
Yi-Fen Wang ◽  
Yong-Qing Wang ◽  
Ling Meng ◽  
...  
Keyword(s):  
Leaf Extract ◽  
Factor Xa ◽  
Adenosine Diphosphate ◽  
Coagulation System ◽  
Dependent Manner ◽  
Thrombin Time ◽  
Antithrombotic Agent ◽  
Thrombin Activity

Toona microcarpaHarms is a tonic, antiperiodic, antirheumatic, and antithrombotic agent in China and India and an astringent and tonic for treating diarrhea, dysentery, and other intestinal infections in Indonesia. In this study, we prepared ethyl-acetate extract from the air-dried leaves ofToona microcarpaHarms and investigated the anticoagulant activitiesin vitroby performing activated partial thromboplastin time (APTT), prothrombin time (PT), and thrombin time (TT) assays. Antiplatelet aggregation activity of the extract was examined using adenosine diphosphate (ADP), collagen, and thrombin as agonists, and the inhibitions of factor Xa and thrombin were also investigated. Bleeding and clotting times in mice were used to determine its anticoagulant activitiesin vivo. It is found thatToona microcarpaHarms leaf extract (TMHE) prolonged APTT, PT, and TT clotting times in a dose-dependent manner and significantly inhibited platelet aggregation induced by thrombin, but not ADP or collagen. Clotting time and bleeding time assays showed that TMHE significantly prolonged clotting and bleeding timesin vivo. In addition, at the concentration of 1 mg/mL, TMHE inhibited human thrombin activity by 73.98 ± 2.78%. This is the first report to demonstrate that THME exhibits potent anticoagulant effects, possibly via inhibition of thrombin activity.

scholarly journals Reversal of Factor Xa Inhibitors by Andexanet Alfa May Increase Thrombogenesis Compared to Pretreatment Values

2019 ◽  
Vol 25 ◽  
pp. 107602961986349 ◽  
Author(s):  
Fakiha Siddiqui ◽  
Alfonso Tafur ◽  
Lorenzo Storino Ramacciotti ◽  
Walter Jeske ◽  
Debra Hoppensteadt ◽  
...  
Keyword(s):  
Thrombin Generation ◽  
Area Under The Curve ◽  
Factor Xa ◽  
Coagulation Factor ◽  
Inhibitory Effects ◽  
Clotting Time ◽  
Reversal Effect ◽  
Normal Human ◽  
Fxa Inhibitor ◽  
Andexanet Alfa

Recombinant coagulation factor Xa (FXa), inactivated Zh-zo, also known as andexanet alfa (AA), is a modified version of human FXa that has been developed to neutralize FXa inhibitors. We studied the reversal effect of AA for these inhibitors in various anticoagulant and thrombin generation (TG) assays. Individual aliquots of normal human plasma containing 1 µg/mL of apixaban, betrixaban, edoxaban, and rivaroxaban, were supplemented with saline or AA at a concentration of 100 µg/mL. Clotting profiles include prothrombinase-induced clotting time, activated partial thromboplastin time, and prothrombin time. Factor Xa activity was measured using an amidolytic method. Thrombin generation was measured using a calibrated automated thrombogram. Differential neutralization of all 4 anticoagulants was noted in the activated clotting time and other clotting tests. The FXa activity reversal profile varied with an observed decrease in apixaban (22%), betrixaban (56%), edoxaban (28%), and rivaroxaban (49%). Andexanet alfa also led to an increased TG in comparison to saline. The peak thrombin was higher (40%), area under the curve (AUC) increased (15%), whereas the lag time (LT) decreased (17%). Andexanet alfa added at 100 µg/mL to various FXa supplemented systems resulted in reversal of the inhibitory effects, restoring the TG profile; AUC, LT, and peak thrombin levels were comparable to those of unsupplemented samples. Andexanet alfa is capable of reversing anti-Xa activity of different oral FXa inhibitors but overshoots thrombogenesis in both the saline and FXa inhibitor supplemented systems. The degree of neutralization of Xa inhibitor is specific to each agent.

Prospective assessment of thrombin generation test for dose monitoring of bypassing therapy in hemophilia patients with inhibitors undergoing elective surgery

Blood ◽  
2010 ◽  
Vol 116 (25) ◽  
pp. 5734-5737 ◽  
Author(s):  
Yesim Dargaud ◽  
Anne Lienhart ◽  
Claude Negrier
Keyword(s):  
Clinical Response ◽  
Thrombin Generation ◽  
Elective Surgery ◽  
High Titer ◽  
Surrogate Marker ◽  
Dependent Manner ◽  
Dose Monitoring ◽  
Prospective Assessment ◽  
Generating Capacity ◽  
Generation Test

Abstract Clinical response to bypassing agents (BPAs) may vary between patients. Surgery is a particular situation, requiring effective hemostasis during the procedure and for several days postoperatively to obtain satisfactory wound healing. However, the optimal dose of BPA in different surgical situations has not been clearly established. We report here a prospective assessment of thrombin generation test (TGT) in monitoring the effectiveness of BPA during 10 elective invasive procedures performed in 6 patients with severe hemophilia and high-titer inhibitors. A standardized 3-step protocol was used in all cases to individually tailor BPA. Thrombin-generating capacity of patients increased after in vitro and ex vivo addition of BPA in a dose-dependent manner. Our results also showed a correlation between in vivo clinical response to BPA and thrombin-generating capacity. These data suggest that TGT may represent a surrogate marker for monitoring bypassing therapies in surgical situations.

Comparison of Branded Enoxaparin (Lovenox) and a Biosimilar Version of Enoxaparin (Fibrinox)

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 4385-4385
Author(s):  
Walter Jeske ◽  
Elizabeth McGeehan ◽  
Omer Iqbal ◽  
Debra Hoppensteadt ◽  
Jeanine M. Walenga ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Thrombin Generation ◽  
Conflicts Of Interest ◽  
Average Molecular Weight ◽  
Molecular Profile ◽  
Distribution Analysis ◽  
Thrombin Time ◽  
Plasma Samples ◽  
Branded Product ◽  
Inhibition Of Thrombin

Abstract Abstract 4385 Several biosimilar versions of branded enoxaparin (Lovenox, Sanofi-Aventis, Paris, France) have recently become available throughout the world. These biosimilar enoxaparin preparations are distributed by multiple suppliers in Asia and in North and South America. Enoxaparin represents a complex mixture of oligosaccharides obtained by alkaline depolymerization of porcine mucosal heparin. It is the most widely used low molecular weight heparin which has been validated for clinical use in multiple indications. While the molecular profile and anti-Xa potencies of some of the biosimilar versions of enoxaparin are comparable, product based differences have been reported amongst some of the biosimilar versions of enoxaparin. The purpose of this study was to compare the biochemical and pharmacologic profile of one biosimilar version of enoxaparin, namely Fibrinox (Sandoz SA, Buenos Aires, Argentina) with the branded product Lovenox. The products were compared in equigravimetric amounts, assuming equivalent potency (100 AXa U/mg). Both products exhibited comparable molecular weight profiles in terms of average molecular weight and oligosachharide distribution. Analysis of the antithrombin binding hexasaccharide fractions of Fibrinox and Lovenox indicated the presence of eight distinct hexasaccharides. The relative proportions these hexasaccharides differed between Fibrinox and Lovenox. The anti-Xa and anti-IIa activities were comparable. In the whole blood clot-based assays such as TEG and ACT, both agents produced similar anticoagulant effects. In the plasma based assays such as the APTT, Heptest and thrombin time, both products showed comparable anticoagulant effects in the normal human pooled plasma samples. However, in plasma samples collected from patients with liver disease who were apparently anticoagulant free, the two products showed differences in their anticoagulant effects in the APTT assay (p<0.05). In the TF mediated thrombin generation assay, Fibrinox produced a stronger inhibition of thrombin generation compared to Lovenox (IC50; Fibrinox, 1.6 μ g/ml, Lovenox 2.2 μ g/ml). No differences were observed between the two products in the agonist induced platelet aggregation assays. However in the 14C serotonin release study, Fibrinox produced a stronger HIT serum mediated 14C release (p<0.05). Differences in the fibrinokinetic profile and the inhibition of thrombin activatable fibrinolytic inhibitor activation were observed with these LMWHs. These studies suggest while both the molecular profile and the pharmacopoeial potency of Fibrinox is similar to the branded product, these drugs can be differentiated in some of the other assays and should be evaluated in terms of additional pharmacologic mechanisims to demonstrate bioequivalence. Disclosures: No relevant conflicts of interest to declare.

Relieving Inhibition of Prothrombinase By TFPIα: a Procoagulant Activity of Unfractionated, Low Molecular Weight, and Nonanticoagulant Heparins

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 1478-1478
Author(s):  
Jeremy P Wood ◽  
Lisa M Baumann Kreuziger ◽  
Rodney M. Camire ◽  
Umesh R Desai ◽  
Alan E. Mast
Keyword(s):  
Molecular Weight ◽  
Unfractionated Heparin ◽  
Research Funding ◽  
Thrombin Generation ◽  
Size Dependence ◽  
Procoagulant Activity ◽  
Low Molecular Weight ◽  
Direct Inhibition ◽  
Acidic Region ◽  
Inhibition Of Thrombin

Abstract Introduction: Prothrombinase, the complex of factor Xa (FXa) and factor Va (FVa), is inhibited by tissue factor pathway inhibitor (TFPI)α during the initiation of coagulation (Wood JP et al, PNAS 2013). Efficient inhibition of thrombin generation by prothrombinase requires an interaction between the TFPIα basic C-terminus and an acidic region of the FVa B-domain. This acidic region is present in FXa-activated FVa and FVa released from activated platelets, but is rapidly removed by thrombin. Thus, prothrombinase inhibition only occurs during the initiation phase of thrombin generation. As the exosite interaction is charge-dependent, large negatively charged molecules, including unfractionated heparin (UFH), block it, prevent prothrombinase inhibition, and promote thrombin generation. Studies using the negatively charged molecule polyphosphate have suggested a size requirement for blocking this TFPIα activity (Smith SA et al, Blood2010). A similar size-dependence may exist with heparins and could have clinical implications, as currently-used heparins range from long (unfractionated heparin; UFH) to medium (low molecular weight heparins; LMWHs) to short (the antithrombin-binding pentasaccharide fondaparinux). Studies were performed to assess the ability of the LMWHs enoxaparin and dalteparin, fondaparinux, and the nonanticoagulant heparin 2-O, 3-O desulfated heparin (ODSH) to block TFPIα and promote thrombin generation through this mechanism. Methods: TFPIα inhibition of thrombin generation by prothrombinase, assembled with a form of FVa containing the acidic region of the B domain, was measured in the absence or presence of UFH, enoxaparin, dalteparin, fondaparinux, and ODSH. The effect of these compounds on the direct inhibition of FXa by TFPIα was measured using a FXa chromogenic substrate. The effect of these compounds on thrombin generation in plasma was measured by calibrated automated thrombography using human plasma immunodepleted of antithrombin III and heparin cofactor II (AT3/HCII-depleted plasma). Results: TFPIα inhibited prothrombinase activity (IC50 = 6.8 nM), and UFH blocked this inhibition (IC50 = 12.5 nM or 14.9 nM at 0.5 or 1 U/mL, respectively). Enoxaparin (0.8 U/mL; IC50 = 30.3 nM) and dalteparin (1 U/mL; IC50 = 29.7 nM) appeared to be more effective at reversing TFPIα inhibition. The reason for this apparent enhanced effect of LMWHs compared to UFH is not clear, as UFH and the LMWHs similarly enhanced the direct inhibition of FXa by TFPIα, and the differential activity was also observed when heparins were normalized to saccharide concentration. The same pattern was observed when measuring thrombin generation in AT3/HCII-depleted plasma, with LMWHs being more procoagulant than UFH. Consistent with TFPIα inhibition being charge-dependent, ODSH promoted thrombin generation similarly to LMWHs in both purified systems and AT3/HCII-depleted plasma. In contrast, clinical doses of fondaparinux had no effect in any assay. In a purified system, ~1000 times the clinical dose of fondaparinux was required to promote thrombin generation. Conclusion: There is a size-dependence for blocking TFPIα inhibition of prothrombinase using heparins, as the pentasaccharide has no effect. However, both LMWHs and UFH are sufficiently long to express this procoagulant activity at therapeutic doses. In addition, the nonanticoagulant heparin ODSH blocks prothrombinase inhibition by TFPIα. This procoagulant activity is likely most clinically relevant under conditions of antithrombin deficiency, which may result from sepsis, liver failure, or administration of L-asparaginase. Under any of these conditions, UFH, LMWHs, and ODSH may have unanticipated procoagulant activity mediated by blocking TFPIα. Disclosures Camire: Pfizer: Consultancy, Patents & Royalties, Research Funding. Mast:Novo Nordisk: Research Funding.

scholarly journals Effects of Andexanet Alfa on Thrombin Generation in Bleeding Associated with Factor Xa Inhibitors

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 711-711
Author(s):  
Michiel Coppens ◽  
Lizhen Xu ◽  
Roisin Bavalia ◽  
Saskia Middeldorp ◽  
Peter Verhamme ◽  
...  
Keyword(s):  
Board Of Directors ◽  
Research Funding ◽  
Thrombin Generation ◽  
Factor Xa ◽  
Educational Material ◽  
Factor Xa Inhibitor ◽  
Employment Equity ◽  
Advisory Committees ◽  
Equity Ownership ◽  
Andexanet Alfa

Introduction Andexanet alfa is a modified recombinant inactive form of human factor Xa developed for reversal of factor Xa inhibitors. In the ANNEXA-4 study, patients with acute major bleeding within 18 h after administration of a factor Xa inhibitor were enrolled and received a bolus of andexanet, followed by a 2-h infusion (Connolly, NEJM 2019;380:1326). In this study, 82% of patients achieved effective hemostasis at 12 h and 10% developed a thrombotic event within 30 days. Anti-Xa activity decreased by 92% after the andexanet bolus but partially recovered after the end of the 2 h infusion. In the present analysis, we evaluated the effect of andexanet alfa on thrombin generation (TG) in patients enrolled in the ANNEXA-4 study and we explored whether TG predicts effective hemostasis or thrombotic events. Methods We included all patients who received andexanet alfa. TG was expressed as the endogenous thrombin potential (ETP) which is the area under the thrombin generation curve. We plotted mean TG at different timepoints between baseline and 30 days after andexanet alfa in patients treated with apixaban and rivaroxaban. We compared the absolute ETP level at 8 h (ETP-8H) after andexanet bolus as this was the first timepoint after the 2 h infusion for which an ETP level was available for most patients. We compared ETP-8H levels between patients with and without effective hemostasis and between those with and without thrombotic events, respectively. ETP-8H was evaluated as a predictor of effective hemostasis and thrombotic events by logistic regression analysis in all patients, and in subgroups of patients with intracranial hemorrhage (ICH) and non-ICH separately. In the ICH subgroups, ETP-8H was also evaluated as a predictor of absolute change in hematoma volume. Results The study population comprised 352 patients (mean age 77.4 years; 47% female) with acute major bleeding (64% ICH, 26% gastrointestinal, 10% other) treated with apixaban (55%), rivaroxaban (36%), enoxaparin (6%), or edoxaban (3%). ETP-8H was available for 327 patients (93%). In patients treated with apixaban or rivaroxaban, andexanet bolus promptly increased mean ETP and this was maintained during infusion. After end of infusion ETP fell but remained in the reference range for at least 18 hours (Figure 1). ETP-8H was similar in patients with or without effective hemostasis (Fig 2a, p = 0.544) and in patients with or without thrombotic complications (Fig 2b, p = 0.610). In the logistic regression analysis, ETP-8H did not predict effective hemostasis (p=0.491) or thrombotic events (p=0.743) (Table), and these results were consistent in ICH and non-ICH patients. ETP-8H did not predict hematoma growth in patients with ICH (p = 0.349). Conclusion A bolus of andexanet alfa, followed by a 2-h infusion in patients with factor Xa inhibitor associated major bleeding promptly restores thrombin generation and this effect is sustained for at least 18 hours. Thrombin generation at 8 h after andexanet bolus did not predict effective hemostasis, intracranial hematoma growth, or thrombotic events. This may be explained by the andexanet dose which was chosen to ensure full reversal of the factor Xa inhibitor in all patients. Disclosures Coppens: Bayer: Honoraria, Research Funding; Bristol-Myers Squibb: Honoraria; Daiichi Sankyo: Honoraria, Research Funding; Sanquin Blood Supply: Research Funding; Pfizer: Honoraria; Uniqure: Research Funding; CSL Behring: Honoraria, Research Funding; Portola Pharmaceuticals, Inc: Honoraria; Boehringer Ingelheim: Research Funding. Middeldorp:Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees, Other: honoraria for advisory activities; Aspen: Research Funding; Portola Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Membership on an entity's Board of Directors or advisory committees, Other: honoraria for advisory activities; Boehringer Ingelheim: Membership on an entity's Board of Directors or advisory committees, Other: honoraria for advisory activities; Bayer: Membership on an entity's Board of Directors or advisory committees, Other: honoraria for advisory activities, Research Funding; Sanofi: Speakers Bureau; Daiichi Sankyo: Other: honoraria for advisory activities, Research Funding. Verhamme:Portola Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bayer Healthcare: Consultancy, Research Funding, Speakers Bureau; Boehringer Ingelheim: Consultancy, Research Funding, Speakers Bureau; Daiichi Sankyo: Consultancy, Research Funding, Speakers Bureau; Pfizer: Consultancy, Research Funding, Speakers Bureau; Bristol-Myers Squibb: Consultancy, Research Funding, Speakers Bureau; Leo Pharma: Consultancy, Research Funding, Speakers Bureau; Janssen: Consultancy. Eikelboom:Heart and Stroke Foundation: Research Funding; Sanofi Aventis: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; Glaxo Smith Kline: Honoraria, Research Funding; Eli Lilly: Honoraria, Research Funding; Daiichi Sankyo: Honoraria, Research Funding; Bristol-Myers Squibb: Honoraria, Research Funding; Boehringer Ingelheim: Honoraria, Research Funding; Bayer: Honoraria, Research Funding; AstraZeneca: Honoraria, Research Funding. Crowther:Bayer: Other: Data and Safety Monitoring Board, Research Funding, Speakers Bureau; BMS Canada: Membership on an entity's Board of Directors or advisory committees, Research Funding; Servier Canada: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Pfizer: Other: preparing educational material and/or providing educational presentations; CSL Behring: Other: preparing educational material and/or providing educational presentations; Diagnostica Stago: Other: preparing educational material and/or providing educational presentations, Research Funding; Alnylam: Equity Ownership; Asahi Kasei: Membership on an entity's Board of Directors or advisory committees; Alexion: Speakers Bureau; Shionogi: Membership on an entity's Board of Directors or advisory committees; Octapharma: Membership on an entity's Board of Directors or advisory committees. Lu:Portola Pharmaceuticals: Employment, Equity Ownership. Yue:Portola Pharmaceuticals: Employment, Equity Ownership. Conley:Portola Pharmaceuticals, Inc.: Employment, Equity Ownership. Connolly:Portola Pharmaceuticals: Consultancy, Research Funding; Bayer Healthcare: Consultancy, Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; Daiichi Sankyo: Consultancy, Research Funding.

scholarly journals Differential Neutralization of Apixaban, Betrixaban, Edoxaban, and Rivaroxaban By Andexanet Alfa As Measured By Whole Blood Thromboelastographic Analysis

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1155-1155
Author(s):  
Siddharth Mehrotra ◽  
Debra Hoppensteadt ◽  
Walter Jeske ◽  
Omer Iqbal ◽  
Alfonso J Tafur ◽  
...  
Keyword(s):  
Whole Blood ◽  
Research Funding ◽  
Surrogate Marker ◽  
Factor Xa ◽  
Coagulation Factor ◽  
Saline Control ◽  
Unequal Variances ◽  
Healthy Donors ◽  
Andexanet Alfa ◽  
The Individual

Introduction/Background: Recombinant coagulation factor Xa (FXa), inactivated Zh-zo, also known as andexanet alfa (AA), is a modified version of human FXa that has been developed as an antidote to neutralize the bleeding effects of oral FXa inhibitors such as, Apixaban and Rivaroxaban. The relative biological effect of these drugs have been investigated using various clot based and amidolytic methods for FXa inhibition. This Factor Xa inhibitory activity acts as a surrogate marker for the circulating level of these agents. We have recently reported that the FXa activity of these Anti-Xa agents does not fully reflect their biologic spectrum (JCath 25,1-11,2019). Whole blood assays such as thromboelastographic analysis represent a global assay which takes into account both the plasmatic and cellular components of blood and provides a more physiologic endpoint to study the anticoagulant effects of these drugs. The purpose of this study was to investigate the anticoagulant effects of currently available oral Anti-Xa agents such as Apixaban, Betrixaban, Edoxaban, and Rivaroxaban and their relative neutralization by AA in terms of such thromboelastographic parameters as R, K, Angle and MA. Materials and Methods: Analysis was carried out in whole blood using thromboelastography (TEG) using the TEG 5000 Hemostasis System (Haemonetics Corp, Massachusetts). Blood was drawn from healthy donors in individual groups (n=5-10) into 3.2% citrated tubes. In the TEG cup for testing, 0.2 M CaCl2, saline (with a filler and control), each of the individual FXa Inhibitors at a final concentration(FC) of 1 ug/mL, AA at FC of both 100 ug/mL and 50 ug/mL were tested for the relative neutralization of the anticoagulant effects. TEG parameters such as R-time, K-time, angle and MA were measured. All results were compiled individually for the saline control, 100 ug/mL AA and 50 ug/mL AA supplemented systems. Statistical analysis was carried out via an F test for equality of variances followed by the appropriate t test for equal or unequal variances. Results: When comparing the anticoagulants directly to one another, it was observed that Edoxaban shows the strongest anticoagulant effects in both R and K time followed by Betrixaban, then Rivaroxaban which were very similar in their anticoagulative effects, with Apixaban showing the weakest anticoagulant effect as shown in Table 1A. In the reversal studies as shown in Table 1B, as measured by various TEG parameters, R-Time, AA (FC=100 ug/mL) showed full neutralization effects in Apixaban (p=.027), Betrixaban(p=<.01), Edoxaban(p<.01), and Rivaroxaban(p<.01). In K-time, Betrixaban and Edoxaban were fully neutralized (respectfully p=.049 and p=.035) with partial neutralizations of Apixaban and Rivaroxaban. No significant neutralization was noted in the Angle and MA. AA at 50 ug/mL showed full neutralizations, in R-time, in Betrixaban and Rivaroxaban (Betrixaban[p<.01] and Rivaroxaban[p=.0287]), AA at this concentration showed partial neutralizations of Apixaban and Edoxaban. In K-time, AA showed full neutralization of Betrixaban and Edoxaban (Betrixaban[p=<.01] and Edoxaban[p<.01]). Apixaban and Rivaroxaban saw no neutralization effect by AA at FC=50 ug/mL in K-time. AA did not exhibit any significant neutralization effects in the Angle or MA parameters. Summary and Conclusion: All of the 4 Agents produced measurable anticoagulative activities at 1 ug/mL as measured by the TEG parameters. Edoxaban exhibited the strongest anticoagulative effect followed by Betrixaban and Rivaroxaban whereas Apixaban showed much weaker anticoagulant effects. AA FC=100 ug/mL showed much stronger, consistent, and complete neutralization effects of all of the 4 FXa Inhibitors when compared to AA at FC=50 ug/mL. These results strongly suggest that regardless of the variable anticoagulative effect exhibited by the FXa Inhibitors, AA at FC=100ug/mL fully neutralized the anticoagulant effects of this agent as measured by the TEG parameters. AA is shown to be the most effective in neutralizing Betrixaban in R-Time and K-Time at both concentrations of AA. AA was seen to neutralize Apixaban the least. It can be concluded that the effect of AA as a neutralizing agent is both drug and donor dependent and therefore dosage adjustment may be needed for the optimal clinical outcome with this antidote. Disclosures Tafur: Recovery Force: Consultancy; Janssen: Other: Educational Grants, Research Funding; BMS: Research Funding; Idorsia: Research Funding; Daichi Sanyo: Research Funding; Stago: Research Funding; Doasense: Research Funding.

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