First steps in the direction of synthetic, allosteric, direct inhibitors of thrombin and factor Xa.
Ontology highlight
ABSTRACT:
SUBMITTER: Verghese J
PROVIDER: S-EPMC2714770 | biostudies-literature | 2009 Aug
REPOSITORIES: biostudies-literature
Publications
First steps in the direction of synthetic, allosteric, direct inhibitors of thrombin and factor Xa.
Verghese Jenson J Liang Aiye A Sidhu Preet Pal Singh PP Hindle Michael M Zhou Qibing Q Desai Umesh R UR
Bioorganic & medicinal chemistry letters 20090606 15
Designing non-saccharide functional mimics of heparin is a major challenge. In this work, a library of small, aromatic molecules based on the sulfated DHP scaffold was synthesized and screened against thrombin and factor Xa. The results reveal that (i) selected monomeric benzofuran derivatives inhibit the two enzymes, albeit weakly; (ii) the two enzymes recognize different structural features in the benzofurans studied suggesting significant selectivity of recognition; and (iii) the mechanism of ...[more]
Similar Datasets
Project description:The absence of an adequate reversal strategy to prevent and stop potential life-threatening bleeding complications is a major drawback to the clinical use of the direct oral inhibitors of blood coagulation factor Xa. Here we show that specific modifications of the substrate-binding aromatic S4 subpocket within the factor Xa active site disrupt high-affinity engagement of the direct factor Xa inhibitors. These modifications either entail amino-acid substitution of S4 subsite residues Tyr99 and/or Phe174 (chymotrypsinogen numbering), or extension of the 99-loop that borders the S4 subsite. The latter modifications led to the engineering of a factor Xa variant that is able to support coagulation in human plasma spiked with (supra-)physiological concentrations of direct factor Xa inhibitors. As such, this factor Xa variant has the potential to be employed to bypass the direct factor Xa inhibitor-mediated anticoagulation in patients that require restoration of blood coagulation.A major drawback in the clinical use of the oral anticoagulants that directly inhibit factor Xa in order to prevent blood clot formation is the potential for life threatening bleeding events. Here the authors describe factor Xa variants that are refractory to inhibition by these anticoagulants and could serve as rescue agents in treated patients.
| S-EPMC5597622 | biostudies-literature
Project description:Thrombin is a key enzyme targeted by the majority of current anticoagulants that are direct inhibitors. Allosteric inhibition of thrombin may offer a major advantage of finely tuned regulation. We present here sulfated benzofurans as the first examples of potent, small allosteric inhibitors of thrombin. A sulfated benzofuran library of 15 sulfated monomers and 13 sulfated dimers with different charged, polar, and hydrophobic substituents was studied in this work. Synthesis of the sulfated benzofurans was achieved through a multiple step, highly branched strategy, which culminated with microwave-assisted chemical sulfation. Of the 28 potential inhibitors, 11 exhibited reasonable inhibition of human ?-thrombin at pH 7.4. Structure-activity relationship analysis indicated that sulfation at the 5-position of the benzofuran scaffold was essential for targeting thrombin. A tert-butyl 5-sulfated benzofuran derivative was found to be the most potent thrombin inhibitor with an IC(50) of 7.3 ?M under physiologically relevant conditions. Michaelis-Menten studies showed an allosteric inhibition phenomenon. Plasma clotting assays indicate that the sulfated benzofurans prolong both the activated partial thromboplastin time and prothrombin time. Overall, this work puts forward sulfated benzofurans as the first small, synthetic molecules as powerful lead compounds for the design of a new class of allosteric inhibitors of thrombin.
| S-EPMC3150610 | biostudies-literature
Project description:Heparins and vitamin K antagonists have been the primary agents used for anticoagulation in certain cardiovascular and thromboembolic diseases for over 50 years. However, they can be difficult to administer and are fraught with limitations. In response to the need for new anticoagulants, direct thrombin inhibitors (DTIs) have been developed and investigated for their utility in prophylaxis and treatment of venous thromboembolism (VTE), heparin-induced thrombocytopenia (HIT), acute coronary syndromes (ACS), secondary prevention of coronary events after ACS, and nonvalvular atrial fibrillation. Currently, four parenteral direct inhibitors of thrombin activity are FDA-approved in North America: lepirudin, desirudin, bivalirudin and argatroban. Of the new oral DTIs, dabigatran etexilate is the most studied and promising of these agents. This review discusses the clinical indications and efficacy of these direct thrombin inhibitors as well as future directions in anticoagulant therapy.
| S-EPMC3195735 | biostudies-literature
Project description:Human factor Xa (FXa) is a serine protease of the common coagulation pathway. FXa is known to activate prothrombin to thrombin, which eventually leads to the formation of cross-linked blood clots. While this process is important in maintaining hemostasis, excessive thrombin generation results in a host of thrombotic conditions. FXa has also been linked to inflammation via protease-activated receptors. Together, coagulopathy and inflammation have been implicated in the pathogenesis of viral infections, including the current coronavirus pandemic. Direct FXa inhibitors have been shown to possess anti-inflammatory and antiviral effects, in addition to their established anticoagulant activity. This review summarizes the pharmacological activities of direct FXa inhibitors, their pharmacokinetics, potential drug-drug interactions and adverse effects, and the details of clinical trials involving direct FXa inhibitors in coronavirus disease 2019 (COVID-19) patients.
| S-EPMC7486161 | biostudies-literature
Project description:Direct inhibition of coagulation factor Xa (FXa) carries significant promise for developing effective and safe anticoagulants. Although a large number of FXa inhibitors have been studied, each can be classified as either possessing a highly flexible or a rigid core scaffold. We reasoned that an intermediate level of flexibility will provide high selectivity for FXa considering that its active site is less constrained in comparison to thrombin and more constrained as compared to trypsin. We studied several core scaffolds including 1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid for direct FXa inhibition. Using a genetic algorithm-based docking and scoring approach, a promising candidate 23 was identified, synthesized, and found to inhibit FXa with a K(i) of 28 μM. Optimization of derivative 23 resulted in the design of a potent dicarboxamide 47, which displayed a K(i) of 135 nM. Dicarboxamide 47 displayed at least 1852-fold selectivity for FXa inhibition over other coagulation enzymes and doubled PT and aPTT of human plasma at 17.1 μM and 20.2 μM, respectively, which are comparable to those of clinically relevant agents. Dicarboxamide 47 is expected to serve as an excellent lead for further anticoagulant discovery.
| S-EPMC3408863 | biostudies-literature
Project description:Dabigatran and rivaroxaban are novel oral anticoagulants that specifically inhibit thrombin and factor Xa, respectively. The aim of this study is to elucidate antithrombotic properties of these anticoagulant agents under arterial and venous shear conditions. Whole blood samples treated with dabigatran or rivaroxaban at 250, 500, and 1000 nM, with/without aspirin and AR-C66096, a P2Y12 antagonist, were perfused over a microchip coated with collagen and tissue thromboplastin at shear rates of 240 and 600 s(-1). Fibrin-rich platelet thrombus formation was quantified by monitoring flow pressure changes. Dabigatran at higher concentrations (500 and 1000 nM) potently inhibited thrombus formation at both shear rates, whereas 1000 nM of rivaroxaban delayed, but did not completely inhibit, thrombus formation. Dual antiplatelet agents weakly suppressed thrombus formation at both shear rates, but intensified the anticoagulant effects of dabigatran and rivaroxaban. The anticoagulant effects of dabigatran and rivaroxaban were also evaluated under static conditions using thrombin generation (TG) assay. In platelet-poor plasma, dabigatran at 250 and 500 nM efficiently prolonged the lag time (LT) and moderately reduce peak height (PH) of TG, whereas rivaroxaban at 250 nM efficiently prolonged LT and reduced PH of TG. In platelet-rich plasma, however, both anticoagulants efficiently delayed LT and reduced PH of TG. Our results suggest that dabigatran and rivaroxaban may exert distinct antithrombotic effects under flow conditions, particularly in combination with dual antiplatelet therapy.
| S-EPMC3908954 | biostudies-literature
Project description:BackgroundThe dry root and rhizome of Ligusticum chuanxiong Hort., or Chuanxiong, has been used as a blood-activating and stasis-removing traditional Chinese medicine for 1000 years. Our previous studies have shown the inhibitory activity on platelet and thrombin (THR) of Chuanxiong. THR and factor Xa (FXa) play significant roles in the coagulation cascade and their inhibitors are of valuable in the treatment of thromboembolic diseases. The aim of the present study is to screen THR and FXa inhibitors from Chuanxiong.MethodsFour extracts [ethyl acetate (EA), butanol (BA) and remained extract (RE) from 75% ethanol extract, and water extract (WE)] of Chuanxiong were prepared, and their THR/FXa inhibitory activities were assessed in vitro. Following silica-gel column chromatography (SC), the active EA extract and BA extract was further partitioned, respectively. Their active fractions (EA-SC1 to EA-SC5; BA-SC1 to BA-SC5) were obtained and analyzed by LC-MS. After modeling by the principal component analysis (PCA) and orthogonal partial least squares discriminate analysis (OPLS-DA), the specific marker compounds were predicted and identified. Their enzyme inhibitory was assessed in vitro and interactions with THR/FXa were investigated by molecular docking analysis.ResultsChuanxiong EA extract showed strong activity against THR and BA extract was more effective in inhibiting FXa activity, and their fractions exhibited obvious difference in enzyme inhibitory activity. Furthermore, marker compounds a-h were predicted by PCA and OPLS-DA, and their chemical structures were identified. Among them, senkyunolide A, Z-ligustilide, ferulic acid and senkyunolide I (IC50 was determined as 0.77 mM) with potential THR inhibitory activity, as well as isochlorogenic acid A with FXa inhibitory activity were screened out. It was found that the four components could interact with the active site of THR, and the binding energy was lower than -?5 kcal/mol. Isochlorogenic acid A were bound to the active site of FXa, and the binding energy was -?9.39 kcal/mol. The IC50 was determined as 0.56 mM.ConclusionsTHR/FXa inhibitory components in different extracts of Chuanxiong were successfully characterized by the method of enzyme inhibition activity assays with ultra performance liquid chromatography-quadrupole time of flight mass spectrometry-based multivariate statistical analysis.
| S-EPMC7457533 | biostudies-literature
Project description:INTRODUCTION:In recent years, the traditional treatments for thrombotic diseases, heparin and warfarin, are increasingly being replaced by novel oral anticoagulants offering convenient dosing regimens, more predictable anticoagulant responses, and less frequent monitoring. However, these drugs can be contraindicated for some patients and, in particular, their bleeding liability remains high. METHODS:We have developed a new class of direct thrombin inhibitors (VE-DTIs) and have utilized kinetics, biochemical, and X-ray structural studies to characterize the mechanism of action and in vitro pharmacology of an exemplary compound from this class, Compound 1. RESULTS:We demonstrate that Compound 1, an exemplary VE-DTI, acts through reversible covalent inhibition. Compound 1 inhibits thrombin by transiently acylating the active site S195 with high potency and significant selectivity over other trypsin-like serine proteases. The compound inhibits the binding of a peptide substrate with both clot-bound and free thrombin with nanomolar potency. Compound 1 is a low micromolar inhibitor of thrombin activity against endogenous substrates such as fibrinogen and a nanomolar inhibitor of the activation of protein C and thrombin-activatable fibrinolysis inhibitor. In the thrombin generation assay, Compound 1 inhibits thrombin generation with low micromolar potency but does not increase the lag time for thrombin formation. In addition, Compound 1 showed weak inhibition of clotting in PT and aPTT assays consistent with its distinctive profile in the thrombin generation assay. CONCLUSION:Compound 1, while maintaining strong potency comparable to the current DTIs, has a distinct mechanism of action which produces a differentiating pharmacological profile. Acting through reversible covalent inhibition, these direct thrombin inhibitors could lead to new anticoagulants with better combined efficacy and bleeding profiles.
| S-EPMC6072017 | biostudies-literature
Project description:Background:The dry root and rhizome of Salvia miltiorrhiza Bunge, or Danshen, is a well-known traditional Chinese medicine with anticoagulant activity. Taking into account that thrombin (THR) and factor Xa (FXa) play crucial roles in the coagulation cascade, it is reasonable and meaningful to screening THR and/or FXa inhibitors from Danshen. Methods:Four extracts [butanol (BA), ethyl acetate (EA) and remained extract (RE) from 75% ethanol extract, and water extract (WE)] of Danshen were prepared, and their THR/FXa inhibitory activities were assessed in vitro. Then, the active EA extract was further separated by silica-gel column chromatography (SC), and its fractions (SC1-SC5) were analyzed by LC-MS. The principal component analysis (PCA) and orthogonal partial least squares discriminate analysis (OPLS-DA) were employed for predicting the specific marker compounds. The chemical structures of targeted compounds were identified by LC-MS/MS and their interactions with THR/FXa were analyzed by the molecular docking analysis. Results:Danshen EA extract showed strong activity against THR and FXa, and its fractions (SC1-SC5) exhibited obvious difference in inhibitory activity against these two enzymes. Furthermore, four marker compounds with potential THR/FXa inhibitory activity were screened by PCA and OPLS-DA, and were identified as cryptotanshinone, tanshinone I, dihydrotanshinone I and tanshinone IIA. The molecular docking study showed that all these four tanshinones can interact with some key amino acid residues of the THR/FXa active cavities, such as HIS57 and SER195, which were considered to be promising candidates targeting THR and/or FXa with low binding energy (<?-?7 kcal mol-1). Conclusions:LC-MS combined with multivariate statistical analysis can effectively screen potential THR/FXa inhibitory components in Danshen.
| S-EPMC7183602 | biostudies-literature
Project description:BackgroundIn premenopausal women, treatment with direct oral factor Xa inhibitors is associated with an increased risk of heavy menstrual bleeding (HMB) compared with vitamin K antagonists (VKA). Treatment with the direct oral thrombin inhibitor dabigatran appears to be associated with a reduced risk of HMB compared with VKA. These findings come from small observational studies or post hoc analyses of trials in which HMB was not a primary outcome. Use of tranexamic acid during the menstrual period may be effective in patients with HMB, but prospective data regarding efficacy and safety in patients on anticoagulant treatment are lacking.Rationale and designA direct comparison of a factor Xa inhibitor and a thrombin inhibitor with HMB as primary outcome, as well as an evaluation of the effects of adding tranexamic acid in women with anticoagulant-associated HMB is highly relevant for clinical practice. The MEDEA study is a randomized, open-label, pragmatic clinical trial to evaluate management strategies in premenopausal women with HMB associated with factor Xa inhibitor therapy.OutcomesWomen using factor Xa inhibitors with proven HMB, as assessed by a pictorial blood loss assessment chart (PBAC) score of >150, will be randomized to one of three study arms: (i) switch to dabigatran; (ii) continue factor Xa inhibitor with addition of tranexamic acid during the menstrual period; or (iii) continue factor Xa inhibitor without intervention. The primary outcome is the difference in PBAC score before and after randomization. Here, we present the rationale and highlight several unique features in the design of the study.
| S-EPMC7845056 | biostudies-literature