Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.6 (thromboplastin)
 13,278 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To enhance the potency of 1,2-dibenzamidobenzene-derived inhibitors of factor Xa (fXa), an amidine substituent was incorporated on one of the benzoyl side chains to interact with Asp189 in the S1 specificity pocket. Lead molecule 1 was docked into the active site of fXa to facilitate inhibitor design. Subsequently, iterative SAR studies and molecular modeling led to a 1000-fold increase in fXa affinity and a refined model of the new inhibitors in the fXa active site. Strong support for the computational model was achieved through the acquisition of an X-ray crystal structure using thrombin as a surrogate protein. The amidines in this series show high levels of selectivity for the inhibition of fXa relative to other trypsin-like serine proteases. Furthermore, the fXa affinity of compounds in this series (K(ass) = 50-500 x 10(6) L/mol) translates effectively into both anticoagulant activity in vitro and antithrombotic activity in vivo.
 ...
PMID:Structure-based design of potent, amidine-derived inhibitors of factor Xa: evaluation of selectivity, anticoagulant activity, and antithrombotic activity. 1071 55

Thrombin inhibitors are potentially useful in medicine for their anticoagulant and antithrombotic effects. We synthesized and evaluated diverse heterocycle-activated ketones based on the d-Phe-Pro-Arg, and related thrombin active-site recognition motifs, as candidate inhibitors. The peptide-based alpha-ketoheterocycles were typically prepared by either an imidate or a Weinreb amide route (Schemes 1 and 2), the latter of which proved to be more general. Test compounds were generally assayed for inhibition of human alpha-thrombin and bovine trypsin. From a structure-based design standpoint, the heterocycle allows one to explore and adjust interactions within the S1' subsite of thrombin. The preferred alpha-ketoheterocycle is a pi-rich 2-substituted azole with at least two heteroatoms proximal to the carbon bearing the keto group, and a preferred thrombin inhibitor is 2-ketobenzothiazole 3, with a potent K(i) value of 0.2 nM and ca. 15-fold selectivity over trypsin. 2-Ketobenzothiazole 13 exhibited exceedingly potent thrombin inhibition (K(i) = 0.000 65 nM; slow tight binding). Several alpha-ketoheterocycles had thrombin K(i) values in the range 0.1-400 nM. The "Arg" unit in the alpha-ketoheterocycles can be sensitive to stereomutation under mildy basic conditions. For example, 2-ketothiazoles 4 and 59 readily epimerize at pH 7.4, although they are fairly stable stereochemically at pH 3-4; thus, suitable conditions had to be selected for the enzymatic assays. Lead d-Phe-Pro-Arg 2-benzothiazoles 3, 4, and 68 displayed good selectivity for thrombin over other key coagulation enzymes (e.g., factor Xa, plasmin, protein Ca, uPA, tPA, and streptokinase); however, their selectivity for thrombin over trypsin was modest (<25-fold). Compounds 3, 4, and 68 exhibited potent in vitro antithrombotic activity as measured by inhibition of gel-filtered platelet aggregation induced by alpha-thrombin (IC(50) = 30-40 nM). They also proved to be potent anticoagulant/antithrombotic agents in vivo on intravenous administration, as determined in the canine arteriovenous shunt (ED(50) = 0.45-0.65 mg/kg) and the rabbit deep vein thrombosis (ED(50) = 0.1-0.4 mg/kg) models. Intravenous administration of 3, and several analogues, to guinea pigs caused hypotension and electrocardiogram abnormalities. Such cardiovascular side effects were also observed with some nonguanidine inhibitors and inhibitors having recognition motifs other than d-Phe-Pro-Arg. 2-Benzothiazolecarboxylates 4 and 68 exhibited significantly diminished cardiovascular side effects, and benzothiazolecarboxylic acid 4 had the best profile with respect to therapeutic index. The X-ray crystal structures of the ternary complexes 3-thrombin-hirugen and 4-thrombin-hirugen depict novel interactions in the S(1)' region, with the benzothiazole ring forming a hydrogen bond with His-57 and an aromatic stacking interaction with Trp-60D of thrombin's insertion loop. The benzothiazole ring of 3 displaces the Lys-60F side chain into a U-shaped gauche conformation, whereas the benzothiazole carboxylate of 4 forms a salt bridge with the side chain of Lys-60F such that it adopts an extended anti conformation. Since 3 has a 10-fold greater affinity for thrombin than does 4, any increase in binding energy resulting from this salt bridge is apparently offset by perturbations across the enzyme (viz. Figure 4). The increased affinity and selectivity of 2-ketobenzothiazole inhibitors, such as 3, may be primarily due to the aromatic stacking interaction with Trp-60D. However, energy contour calculations with the computer program GRID also indicate a favorable interaction between the benzothiazole sulfur atom and a hydrophobic patch on the surface of thrombin.
 ...
PMID:In-depth study of tripeptide-based alpha-ketoheterocycles as inhibitors of thrombin. Effective utilization of the S1' subsite and its implications to structure-based drug design. 1577 42

Lead (Pb) is a ubiquitous heavy metal pollutant in various environmental media, especially in food and drinking water. In human blood, about 95% of lead is associated with erythrocytes, suggesting that erythrocytes could be an important target of lead toxicity in the cardiovascular system. Recent studies suggested that erythrocytes could contribute to blood coagulation via phosphatidylserine (PS) exposure and resultant procoagulant activation. We investigated the effects of lead on the procoagulant activity of erythrocytes using in vitro human erythrocyte and in vivo rat models. In a flow cytometric analysis, lead (Pb2+) enhanced PS exposure on human erythrocytes in a concentration- and time-dependent manner. The concentration of lead (1-5 microM) used in the current investigation is well within the ranges observed in blood from lead-exposed populations. PS exposure by lead appeared to be mediated by increased intracellular calcium levels as shown by 19F-NMR and intracellular ATP depletion. Consistent with these findings, the activity of scramblase, which is important in the induction of PS exposure, was enhanced, whereas the activity of flippase, which translocates exposed PS to inner membrane, was inhibited by lead treatment. Furthermore, lead-exposed erythrocytes increased thrombin generation as determined by a prothrombinase assay and accelerated the coagulation process initiated by tissue factor in plasma. These procoagulant activations by lead were also confirmed in vivo. Administration of lead significantly enhanced PS exposure on erythrocytes and, more importantly, elevated thrombus formation in a rat venous thrombosis model. These results suggest that lead exposure can provoke procoagulant activity in erythrocytes by PS exposure, contributing to enhanced clot formation. These data will provide new insights into the mechanism of lead-induced cardiovascular diseases.
 ...
PMID:Lead-induced procoagulant activation of erythrocytes through phosphatidylserine exposure may lead to thrombotic diseases. 1722 25

There remains a high unmet medical need for a safe oral therapy for thrombotic disorders. The serine protease factor Xa (fXa), with its central role in the coagulation cascade, is among the more promising targets for anticoagulant therapy and has been the subject of intensive drug discovery efforts. Investigation of a hit from high-throughput screening identified a series of thiophene-substituted anthranilamides as potent nonamidine fXa inhibitors. Lead optimization by incorporation of hydrophilic groups led to the discovery of compounds with picomolar inhibitory potency and micromolar in vitro anticoagulant activity. Based on their high potency, selectivity, oral pharmacokinetics, and efficacy in a rat venous stasis model of thrombosis, compounds ZK 814048 (10b), ZK 810388 (13a), and ZK 813039 (17m) were advanced into development.
 ...
PMID:Thiophene-anthranilamides as highly potent and orally available factor Xa inhibitors. 1753 95

West Nile virus and dengue virus are mosquito-borne flaviviruses that cause a large number of human infections each year. No vaccines or chemotherapeutics are currently available. These viruses encode a serine protease that is essential for polyprotein processing, a required step in the viral replication cycle. In this study, a high-throughput screening assay for the West Nile virus protease was employed to screen approximately 32,000 small-molecule compounds for identification of inhibitors. Lead inhibitor compounds with three distinct core chemical structures (1 to 3) were identified. In a secondary screening of selected compounds, two compounds, belonging to the 8-hydroxyquinoline family (compounds A and B) and containing core structure 1, were identified as potent inhibitors of the West Nile virus protease, with K(i) values of 3.2 +/- 0.3 microM and 3.4 +/- 0.6 microM, respectively. These compounds inhibited the dengue virus type 2 protease with K(i) values of 28.6 +/- 5.1 microM and 30.2 +/- 8.6 microM, respectively, showing some selectivity in the inhibition of these viral proteases. However, the compounds show no inhibition of cellular serine proteases, trypsin, or factor Xa. Kinetic analysis and molecular docking of compound B onto the known crystal structure of the West Nile virus protease indicate that the inhibitor binds in the substrate-binding cleft. Furthermore, compound B was capable of inhibiting West Nile virus RNA replication in cultured Vero cells (50% effective concentration, 1.4 +/- 0.4 microM; selectivity index, 100), presumably by inhibition of polyprotein processing.
 ...
PMID:Identification and biochemical characterization of small-molecule inhibitors of west nile virus serine protease by a high-throughput screen. 1860 44

A novel series of potent and efficacious factor Xa inhibitors which possesses sulfoximine moiety as novel S4 binding element in anthranilamide chemotype has been identified. Lead optimization at this novel P4 group led to many potent factor Xa inhibitors with excellent anticoagulant activity in human plasma. Selected compounds were dosed orally in rats and checked for their ex vivo prothrombin time prolonging activity, which resulted in identification of compound 5-chloro-N-(5-chloropyridin-2-yl)-2-(4-(N-(2-(diethylamino)acetyl)-S-methylsulfonimidoyl)benzamido)benzamide (18f). The detailed pharmacokinetic evaluation and subsequent metabolism study of 18f suggested the presence of an active metabolite. The compound 18f and its active metabolite 18b demonstrated excellent in vivo efficacy in both arterial and venous thrombosis model in rats and were found to be highly selective against related serine proteases. Based on this promising profile, compound 18f was selected for further evaluation.
 ...
PMID:Synthesis and structure-activity relationship of potent, selective and orally active anthranilamide-based factor Xa inhibitors: application of weakly basic sulfoximine group as novel S4 binding element. 2312 11