Application of Molecular Modeling to Development of New Factor Xa Inhibitors

In consequence of the key role of factor Xa in the clotting cascade and absence of its activity in the processes that do not affect coagulation, this protein is an attractive target for development of new blood coagulation inhibitors. Factor Xa is more effective and convenient target for creation of anticoagulants than thrombin, inhibition of which may cause some side effects. This study is aimed at finding new inhibitors of factor Xa by molecular computer modeling including docking SOL and postdocking optimization DISCORE programs. After validation of molecular modeling methods on well-known factor Xa inhibitors the virtual screening of NCI Diversity and Voronezh State University databases of ready-made low molecular weight species has been carried out. Seventeen compounds selected on the basis of modeling results have been tested experimentallyin vitro. It has been found that 12 of them showed activity against factor Xa (IC50= 1.8–40 μM). Based on analysis of the results, the new original compound was synthesized and experimentally verified. It shows activity against factor Xa with IC50value of 0.7 μM.

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An Approach to Assigning In Vitro Potency to Unfractionated and Low Molecular Weight Heparins Based on the Inhibition of Prothrombin Activation and Catalysis of Thrombin Inhibition

Thrombosis and Haemostasis ◽

10.1055/s-0038-1647028 ◽

1988 ◽

Vol 60 (02) ◽

pp. 193-198 ◽

Cited By ~ 12

Author(s):

F A Ofosu ◽

L M Smith ◽

N Anvari ◽

M A Blajchman

Keyword(s):

Molecular Weight ◽

Antithrombin Iii ◽

Factor Xa ◽

Fold Increase ◽

Low Molecular Weight ◽

Thrombin Inhibition ◽

Thrombin Activity ◽

Antithrombotic Efficacy ◽

Prothrombin Activation

SummaryUnfractionated and low molecular weight (LMW) heparins with good antithrombotic activity invariably catalyze thrombin inhibition and inhibit the appearance of thrombin activity in contact-activated plasma. Conversely, the antithrombotic efficacy of LMW heparins decreases as their ability to catalyze thrombin inhibition and to inhibit the appearance of thrombin activity in plasma decrease. The activated partial thromboplastin time (APTT) has proven a reliable test for assaying unfractionated heparin. We therefore compared 2 unfractionated and 3 LMW heparins on the basis of the minimum concentrations required to double the APTT of normal plasma and by then determined how this anticoagulant effect was achieved. The amount of unfractionated and LMW heparin which doubled the APTT was found to be equivalent to —0.25 antithrombin units. This concentration of each glycosaminoglycan completely inhibited prothrombin activation for 45 s after CaCl2 was added to contact-activated plasma; accelerated thrombin inhibition by purified antithrombin III by approximately 50-fold; and accelerated thrombin inhibition equally by anti thrombin III in undiluted plasma. This concentration of the three LMW heparins increased, by approximately 70fold, the rate of factor Xa inhibition by purified antithrombin III compared to the 50-fold increase seen with the two unfractionated heparins. These results thus suggest that tests based on the inhibition of prothrombin activation and/or on the catalysis of thrombin inhibition provide a useful basis for assigning in vitro potency to both unfractionated and LMW heparins.

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The Role of Factor Xa-Independent Pathway and Anticoagulant Therapies in Cancer-Related Stroke

Journal of Clinical Medicine ◽

10.3390/jcm11010123 ◽

2021 ◽

Vol 11 (1) ◽

pp. 123

Author(s):

Hyung Jun Kim ◽

Jong-Won Chung ◽

Oh Young Bang ◽

Yeon Hee Cho ◽

Yun Jeong Lim ◽

...

Keyword(s):

Cancer Patients ◽

Factor Xa ◽

Factor Xa Inhibitors ◽

Low Molecular Weight ◽

Patients With Cancer ◽

Before And After ◽

Underlying Mechanisms ◽

D Dimer ◽

Active Cancer

Background: The optimal strategy for stroke prevention in cancer patients is unknown. We compared the underlying mechanisms of coagulopathy and the effects of anticoagulants in patients with active cancer and atrial fibrillation (AF). Methods: We retrospectively enrolled 46 consecutive patients with embolic stroke of unknown source and active cancer (cancer stroke). We consecutively screened patients with cancer patients without stroke (n = 29), AF stroke (n = 52), and healthy subjects (n = 28), which served as controls. Patients with cancer stroke were treated with either enoxaparin (a low-molecular-weight heparin) or a factor Xa inhibitor, and those with AF stroke were treated with factor Xa inhibitors. D-dimer, factor Xa, and circulating cell-free DNA (cfDNA), a marker of neutrophil extracellular traposis, were measured at both before and after anticoagulation. Results: In AF stroke, factor Xa activity and cfDNA and D-dimer levels were decreased by treatment with factor Xa inhibitors. In contrast, in cancer stroke, factor Xa activity was decreased, D-dimer levels were unchanged, and cfDNA levels were increased by treatment with factor Xa inhibitors. In cancer stroke patients treated with enoxaparin, D-dimer levels were decreased (p = 0.011) and cfDNA levels were unchanged. Conclusion: The anticoagulation effects of factor Xa inhibitors differed between cancer stroke and AF stroke.

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Comparison of the Anticoagulant and Antithrombotic Effects of Synthetic Thrombin and Factor Xa Inhibitors

Thrombosis and Haemostasis ◽

10.1055/s-0038-1645198 ◽

1990 ◽

Vol 63 (02) ◽

pp. 220-223 ◽

Cited By ~ 19

Author(s):

J Hauptmann ◽

B Kaiser ◽

G Nowak ◽

J Stürzebecher ◽

F Markwardt

Keyword(s):

Factor Xa ◽

Thrombin Inhibitors ◽

Factor Xa Inhibitors ◽

Venous Stasis ◽

Clotting Time ◽

Thrombosis Model ◽

Activity Factor ◽

Antithrombotic Effects

SummaryThe anticoagulant effect of selected synthetic inhibitors of thrombin and factor Xa was studied in vitro in commonly used clotting assays. The concentrations of the compounds doubling the clotting time in the various assays were mainly dependent on their thrombin inhibitory activity. Factor Xa inhibitors were somewhat more effective in prolonging the prothrombin time compared to the activated partial thromboplastin time, whereas the opposite was true of thrombin inhibitors.In vivo, in a venous stasis thrombosis model and a thromboplastin-induced microthrombosis model in rats the thrombin inhibitors were effective antithrombotically whereas factor Xa inhibitors of numerically similar IQ value for the respective enzyme were not effective at equimolar dosageThe results are discussed in the light of the different prelequisiles and conditions for inhibition of thrombin and factor Xa in the course of blood clotting.

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Novel concatameric heparin-binding peptides reverse heparin and low-molecular-weight heparin anticoagulant activities in patient plasma in vitro and in rats in vivo

Blood ◽

10.1182/blood-2003-07-2334 ◽

2004 ◽

Vol 103 (4) ◽

pp. 1356-1363 ◽

Cited By ~ 30

Author(s):

Barbara P. Schick ◽

David Maslow ◽

Adrianna Moshinski ◽

James D. San Antonio

Keyword(s):

Molecular Weight ◽

Low Molecular Weight Heparin ◽

Tandem Repeats ◽

Factor Xa ◽

Low Molecular Weight ◽

Heparin Binding ◽

High Affinity ◽

Binding Peptides

Abstract Patients given unfractionated heparin (UFH) or low-molecular-weight heparin (LMWH) for prophylaxis or treatment of thrombosis sometimes suffer serious bleeding. We showed previously that peptides containing 3 or more tandem repeats of heparin-binding consensus sequences have high affinity for LMWH and neutralize LMWH (enoxaparin) in vivo in rats and in vitro in citrate. We have now modified the (ARKKAAKA)n tandem repeat peptides by cyclization or by inclusion of hydrophobic tails or cysteines to promote multimerization. These peptides exhibit high-affinity binding to LMWH (dissociation constant [Kd], ≈ 50 nM), similar potencies in neutralizing anti–Factor Xa activity of UFH and enoxaparin added to normal plasma in vitro, and efficacy equivalent to or greater than protamine. Peptide (ARKKAAKA)3VLVLVLVL was most effective in all plasmas from enoxaparin-treated patients, and was 4- to 20-fold more effective than protamine. Several other peptide structures were effective in some patients' plasmas. All high-affinity peptides reversed inhibition of thrombin-induced clot formation by UFH. These peptides (1 mg/300 g rat) neutralized 1 U/mL anti–Factor Xa activity of enoxaparin in rats within 1 to 2 minutes. Direct blood pressure and heart rate measurements showed little or no hemodynamic effect. These heparin-binding peptides, singly or in combination, are potential candidates for clinical reversal of UFH and LMWH in humans.

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Heparin induced thrombocytopenia

Hämostaseologie ◽

10.5482/hamo-15-04-0013 ◽

2015 ◽

Vol 35 (04) ◽

pp. 372-375 ◽

Cited By ~ 1

Author(s):

N. A. Viniou ◽

P. Diamantopoulos ◽

J. Barbetseas ◽

E. A. Sanidas

Keyword(s):

Oral Anticoagulants ◽

Direct Oral Anticoagulants ◽

Treatment Options ◽

Factor Xa ◽

Additional Treatment ◽

Thrombin Inhibitors ◽

Factor Xa Inhibitors ◽

Clinical Aspects ◽

Heparin Induced Thrombocytopenia

SummaryHeparin induced thrombocytopenia (HIT) is a prothrombotic syndrome initiated by platelet-activating auto-antibodies with potentially devastating complications. Once the diagnosis of HIT is suspected, discontinuation of heparin and treatment with an alternative anticoagulant are mandatory. While established drugs for HIT are no longer available, parenteral factor Xa inhibitors, thrombin inhibitors and perhaps the direct oral anticoagulants provide additional treatment options. The aim of this review was to highlight the current clinical aspects regarding HIT focusing on the role of novel medications.

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Role of Factor Xa Inhibitors in Cancer-Associated Thrombosis: Any New Data?

Advances in Hematology ◽

10.1155/2011/196135 ◽

2011 ◽

Vol 2011 ◽

pp. 1-12 ◽

Cited By ~ 5

Author(s):

Ali Zalpour ◽

Michael H. Kroll ◽

Vahid Afshar-Kharghan ◽

Syed Wamique Yusuf ◽

Carmen Escalante

Keyword(s):

Clinical Trials ◽

Cancer Patients ◽

Anticoagulation Therapy ◽

Factor Xa ◽

Factor Xa Inhibitors ◽

The Us ◽

Prevention And Treatment ◽

Cancer Associated Thrombosis ◽

Current Guidelines

The association between cancer and venous thromboembolism (VTE) has been well documented in the literature. Prevention and treatment of VTE in cancer patients is imperative. Typically, the mainstay regimen for VTE prevention and treatment has been anticoagulation therapy, unless contraindicated. This therapy consists of unfractionated heparin (UFH), low-molecular-weight heparin (LMWH), factor Xa inhibitor, or vitamin K antagonist (VKA). Current guidelines recommend LMWH over VKA for the treatment of VTE in cancer patients. Factor-specific anticoagulants have been proven safe and effective, and recently factor Xa inhibitors have emerged as a treatment alternative to heparins and VKA. Currently, three factor Xa inhibitors have been identified: fondaparinux (the only one approved so far by the US Food and Drug Administration), idraparinux (in clinical trials), and idrabiotaparinux (in clinical trials). This paper will examine the role of these agents, focusing on fondaparinux, for the prevention and treatment of VTE in cancer patients.

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THE ROLE OF A 3-0 SULFO GROUP IN THE GLUCOSAMINE RESIDUE OF A SYNTHETIC OLIGOSACCHARIDE FOR THE DETERMINATION OF ANTITHROMBOTIC ACTIONS

10.1055/s-0038-1644181 ◽

1987 ◽

Author(s):

J M Walenga ◽

J Fareed ◽

M Petitou ◽

J C Lormeau ◽

M Samama ◽

...

Keyword(s):

Sulfo Group ◽

Factor Xa ◽

Antithrombotic Effect ◽

Antithrombotic Activity ◽

Thrombosis Model ◽

Factor Xa Inhibition

We have previously reported on the antithromboticaction of a chemically synthesized heparin pentasaccharide which exhibits high affinity to anti thrombinIII and sole anti-factor Xa activity. In order to investigate the relative importance of the 3-0 sulfo group of this pentasaccharide, we evaluated the in vitro and in vivo antithrombotic activity of a synthetic pentasccharide devoid of the sulfo group at the third position of the glucosamine residue. In amidolytic and clot-based assays the 3-0 de- sulfated pentasaccharide (3-0-DP) failed to exhibit any antifactor Xa actions at concentrations <100 ug/ml in humanor rabbit plasmas, whereas pentasaccharide showed strong factor Xa inhibition at 1.0 ug/ml IK-=3.2x10 M)and at 10.0 ug/ml in rabbit plasma (K.=9.0×10™7 M). Using a rabbit stasis thrombosis model in which thrombosis was induce by human serum or an activated pro-thrombin complex concentrate, 3-0-DP failed to produce any antithrombotic action in acute intravenous regimens at dosages up to 200 ug/kg. In these two models, pentasaccharide produced >80% inhibition of induced thrombosis. These studies demonstrate the critical importance of the 3-0 sulfo group in this heparin pentasaccharide for the determination of antithrombotic activity, and that in this type of oligosaccharide, anti-factor Xa activity is responsible for producing the antithrombotic effect.

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EFFECT OF VARIOUS HEPARIN(OID)S ON HEPARIN COFACTOR II MEDIATED ANTI-THROMBIN ACTIVITY AND INHIBITION OF THROMBIN GENERATION IN VITRO

10.1055/s-0038-1644353 ◽

1987 ◽

Author(s):

T G van Dinther ◽

F Hol ◽

D G Meuleman

Keyword(s):

Molecular Weight ◽

Thrombin Generation ◽

Factor Xa ◽

Weight Fraction ◽

Blood Platelet ◽

Low Molecular Weight ◽

Heparin Cofactor Ii ◽

Thrombin Activity ◽

At Iii

The effects of various heparin(oid)s, standard heparin VII (SH), dermatan sulphate (DS), a low molecular weight fraction of heparin (UMW-H), FragminR (FRA), Org 10172 = low molecular weight heparinoid, the fraction of Org 10172 with high affinity for AT-III (HA-10172) and the low affinity fraction (LA-10172) respectively were examined on in vitro thrombin generation and inactivation.Thrombin inactivation in the presence of either heparin cofactor II (HC-II) or anti-thrombin III (AT-III) was assessed with two newly developed assays using the purified cofactors, thrombin and chromogenic substrate S2238 on microtiterplates. Thrombin generation in the presence of HC-II and AT-III was studied using purified factor Xa, prothrombin and blood platelet lysate and the residual thrombin activity was assessed amidolytically.The inhibition of the compounds on thrombin activity are summarized in the tableThe following conclusions can be drawn:- SH, LMW-H, HA-10172 and FRA potentiate the AT-III mediated inactivation of Ha more strongly than the HC-II mediated inactivation.- DS and LA-10172 show the reverse pattern of inactivation, while Org 10172 potentiates both inactivaton pathways to a similar extent.Thrombin generation in the presence of HC-II is inhibited by mw-heparin(oid)s at approx. 2-5 times lower concentrations than the HC-II mediated thrombin inactivation, while the inhibiting effect of SH in both assays is comparable.AT-III mediated thrombin generation inhibition and AT-III mediated thrombin inactivation is comparable as well for SH, LMW-H and FRA. In contrast, Org 10172 and its subfractions are approx. 10 times more potent on AT-III mediated thrombin generation inhibition than on AT-III mediated thrombin inactivation.Org 10172 shows low anti-thrombin activity and this activity is mainly mediated via FC-II.

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Novel Design of Peptides to Reverse the Anticoagulant Activities of Heparin and other Glycosaminoglycans

Thrombosis and Haemostasis ◽

10.1055/s-0037-1615609 ◽

2001 ◽

Vol 85 (03) ◽

pp. 482-487 ◽

Cited By ~ 15

Author(s):

Joel Gradowski ◽

James San Antonio ◽

Jose Martinez ◽

Barbara Schick

Keyword(s):

Molecular Weight ◽

Unfractionated Heparin ◽

Low Molecular Weight Heparin ◽

Factor Xa ◽

Low Molecular Weight ◽

Heparin Binding ◽

Antifactor Xa ◽

Novel Design

SummaryPatients undergoing anticoagulation with unfractionated heparin, low molecular weight heparin, or danaparoid may experience excess bleeding which requires reversal of the anticoagulant agent. Protamine is at present the only agent available for reversal of unfractionated heparin. Protamine is not effective in patients who have received low molecular weight heparin or danaparoid. We have developed a series of peptides based on consensus heparin binding sequences (Verrecchio et al., J Biol Chem 2000; 275: 7701-7707) that are capable of neutralizing the anti-thrombin activity of unfractionated heparin in vitro, the antifactor Xa activity of unfractionated heparin, Enoxaparin (Lovenox) and danaparoid (Orgaran) in vitro and the anti-Factor Xa activity of Enoxaparin in vivo in rats. These peptides may serve as alternatives for Protamine reversal of UFH and may be useful for neutralization of enoxaparin and danaparoid in humans.

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