Antithrombotic Activity Research Articles

Review of Topical Sodium Heparin 1000 IU/g Gel in Symptomatic Uncomplicated Superficial Thrombophlebitis.

Heparin, a mixture of sulfated polymorphic polysaccharides (glycosaminoglycan) chains of variable lengths and weights and a natural anticoagulant, is widely used in medical practice to prevent intravascular blood coagulation. Heparin has demonstrated antithrombotic and anti-inflammatory activity, and it is mostly administered systemically (intravenously or subcutaneously) for primary or secondary prevention of venous thromboembolism after surgical interventions, or immobilized patients, or on short-term antithrombotic therapy of patients with atrial fibrillation who must undergo treatment. However, since systemic administration of heparin could be, in certain cases, linked to an increased risk of bleeding, topical heparin is widely used for the prevention and treatment of local symptoms of peripheral vascular disorders, such as venous insufficiency, varicose veins, or superficial thrombophlebitis. This review summarizes the main safety and efficacy characteristics of the topical formulation of Heparin in Gel form (1000 International Units of Heparin/g Gel) currently in use, which has demonstrated an excellent efficacy and tolerability profile in reducing signs and symptoms of peripheral vascular disease, e.g., varicose syndromes and their complications, phlebothrombosis, thrombophlebitis, superficial periphlebitis, varicose ulcers, for post-operative varicophlebitis, sequelae of saphenectomy, for traumas and contusions, local edemas and infiltrates, subcutaneous hematoma and for traumatic affections of musculotendinous and capsuloligamentous apparatuses.

Tyrosine phosphorylation of recombinant hirudin increases affinity to thrombin and antithrombotic activity.

Thrombin is one of the key enzymes of the blood coagulation system and a promising target for the development of anticoagulants. One of the most specific natural thrombin inhibitors is hirudin, contained in the salivary glands of medicinal leeches. The medicinal use of recombinant hirudin is limited because of the lack of sulfation on Tyr63, resulting in a 10-fold decrease in activity compared to native (sulfated) hirudin. In the present work, a set of hirudin derivatives was tested for affinity to thrombin: phospho-Tyr63, Tyr63(carboxymethyl)Phe, and Tyr63Glu mutants, which mimic Tyr63 sulfation and Gln65Glu mutant and lysine-succinylated hirudin, which enhance the overall negative charge of hirudin, as well as sulfo-hirudin and desulfo-hirudin as references. Using steered molecular dynamics simulations with subsequent umbrella sampling, phospho-hirudin was shown to exhibit the highest affinity to thrombin among all hirudin analogs, including native sulfo-hirudin; succinylated hirudin was also prospective. Phospho-hirudin exhibited the highest antithrombotic activity in in vitro assay in human plasma. Taking into account the modern methods for obtaining phospho-hirudin and succinylated hirudin, they are prospective as anticoagulants in clinical practice.

A pyruvylated and sulfated galactan from the green alga Dictyosphaeria cavernosa: Structure, anticoagulant property and inhibitory effect on zebrafish thrombosis

A pyruvylated and sulfated galactan from the green alga Dictyosphaeria cavernosa, designated PSG, was obtained by dilute alkali extraction, ion-exchange and gel filtration chromatography. The backbone of PSG was composed of 3-linked β-d-Galp units with partial sulfation on C-4 and C-6. Pyruvate ketals were linked to O-3 and O-4 of nonreducing terminal β-d-Galp, as well as O-4 and O-6 of 3-linked β-d-Galp. The branches consisting of 6-linked β-d-Galp(4SO4) and β-d-Galp(3,4-Pyr)-(1→ units were located at C-6 of 3-linked β-d-Galp unit. PSG possessed obvious anticoagulant effect in vitro as assessed by the tests of activated partial thromboplastin time and thrombin time. The assay of anticoagulant mechanism showed that PSG promoted thrombin inactivation mediated by heparin cofactor-II and antithrombin-III (ATIII), and could effectively potentiate factor Xa inactivation by ATIII. The antithrombotic activity of PSG in vivo was assessed by phenylhydrazine (PHZ)-induced zebrafish thrombotic model. The results indicated that PSG obviously reduced peripheral erythrocytes aggregation, enhanced cardiac blood flow and improved peripheral platelet circulation, and PSG possessed a marked inhibitory effect on the PHZ-induced zebrafish thrombosis. Thus, PSG is a hopeful anticoagulant and antithrombotic polysaccharide.

Docking study and molecular dynamic approach to predicting the activity of 4-(4-methoxy)benzoyloxy-3-methoxycinnamic acid against COX-1 enzyme

Background: Ferulic acid is a phenolic acid compound that has anti-inflammatory and anti-thrombosis activity. However, ferulic acid has the disadvantage of poor absorption. Structural modifications can be made to increase the biological activity of the compound. In this research, the structure of ferulic acid was modified into 4-(4-methoxy)benzoyloxy-3-methoxycinnamic acid. Objective: The purpose of this research is to predict the activity of 4-(4-methoxy)benzoyloxy-3-methoxycinnamic acid. Method: It was carried out using the Pass Online Prediction, Autodock 1.5.7, Discovery Studio and Maestro Schrödinger 2020-1 software. Result: The activity prediction results showed that 4-(4-methoxy)benzoyloxy-3-methoxycinnamic acid had anti-inflammatory and anti-thrombotic activity. The molecular docking results showed that 4-(4-methoxy)benzoyloxy-3-methoxycinnamic acid had lower free bond energy values than ferulic acid, hence, predicted to have greater activity. Molecular dynamics also reveal better stability interactions of the 4-(4-methoxy)benzoyloxy-3-methoxycinnamic acid with COX-1 protein than the ferulic acid. Conclusion: The compund, 4-(4-methoxy)benzoyloxy-3-methoxycinnamic acid is feasible to be synthesised.

In-Silico Screening of Phytocompounds of Justicia Adhatoda for Thrombolytic Activity

Background: The risk of thrombosis and its associated mortality is becoming a greater public health concern in both industrialized and developing nations. Clinical condition that is myocardial infarction, ischemic stroke, unstable angina, peripheral vascular diseases like deep vein thrombosis prioritise top in the list of thrombotic disorders. Conventional thrombolytic medications including alteplase, anistreplase, streptokinase and urokinase offer life risking adverse effects like haemorrhage, stroke, vascular dysfunction, hypertension, internal bleeding and so on. In order to overcome the undesirable side effects caused by the conventional thrombolytic therapy, there is ominous need of alternate complementary treatment from traditional medicine. Majority of the Siddha formulations comprise of some potential medicinal herbs which are found to be biocompatible and also have a wide safety window. Justicia adhatoda is one such novel herb which is investigated broadly by researchers till date, aspired by its unique pharmacological property rendered by the bioactive components present in it. Methods: Hence, the present research is aimed at in-silico screening of certain classes of alkaloids and flavonoids retrieved from J. adhatoda to explore the possible anti-thrombotic activity against the target human plasminogen activation loop peptide using AutoDock screening tool. Results and Conclusion: The computational analysis’s findings led to the conclusion that the bioactive compounds present in Justicia adhatoda like Astragalin, Kaempferol, Vitexin, Vasicolinone and Adhatodine reveals significant binding affinity against target plasminogen. Thereby it was determined that these compounds may have promising anti-thrombotic efficacy due to their considerable binding affinity towards the target plasminogen.

Anti-thrombotic Effects Mediated by a Novel Dual-Target Peptide Inhibiting Both Platelet Aggregation and Thrombin Activity without Causing Bleeding.

Classical anticoagulants and antiplatelets are associated with high frequencies of bleeding complications or treatment failure when used as single agents. Thrombin plays an important role in the blood coagulation system. GP IIb/IIIa is the central receptor of platelets, which can recognize the Arg-Gly-Asp (RGD) sequence and activate platelets. Molecular simulation and homology modeling were performed to design a novel dual-target anticoagulant short peptide (PTIP ). The activities of PTIP on coagulation and platelet in vitro were analyzed. The antithrombotic activity of PTIP was determined by pulmonary thromboembolism model, ferric chloride injury model and arteriovenous bypass thrombosis model. Bleeding effect and toxicity of PTIP were evaluated. We have constructed a novel dual-target peptide (PTIP) based on the direct thrombin inhibitor peptide (DTIP). PTIP was expressed at high levels in Pichia pastoris. PTIP interfered with thrombin-mediated coagulation and ADP-induced platelet aggregation in vitro. When injected intravenously or subcutaneously, PTIP showed potent and dose-dependent extension of aPTT and PT which were similar to DTIP; but only PTIP was capable of inhibiting platelet aggregation. PTIP (1.0 mg/kg) decelerated thrombosis formation in venous and arterial vessels induced by FeCl3 injury. PTIP (1.0 mg/kg) also prevented deep venous thrombosis and increased the survival rate associated with pulmonary thromboembolism. And PTIP effectively reduced thrombus length in arteriovenous bypass thrombosis model. Moreover, the antithrombotic dose of PTIP could not induce bleeding. These data establish that PTIP represents a novel antithrombotic agent whose effects involve both inhibition of platelet activation and reduction of fibrin generation. And PTIP not only can be used in venous thrombosis and arterial thrombosis, it can also replace the combined treatment of antiplatelet and anticoagulant drugs in thrombotic diseases.

Polyester urethane urea (PEUU) functionalization for enhanced anti-thrombotic performance: advancing regenerative cardiovascular devices through innovative surface modifications.

Introduction: Thrombogenesis, a major cause of implantable cardiovascular device failure, can be addressed through the use of biodegradable polymers modified with anticoagulating moieties. This study introduces a novel polyester urethane urea (PEUU) functionalized with various anti-platelet deposition molecules for enhanced antiplatelet performance in regenerative cardiovascular devices. Methods: PEUU, synthesized from poly-caprolactone, 1,4-diisocyanatobutane, and putrescine, was chemically oxidized to introduce carboxyl groups, creating PEUU-COOH. This polymer was functionalized in situ with polyethyleneimine, 4-arm polyethylene glycol, seleno-L-cystine, heparin sodium, and fondaparinux. Functionalization was confirmed using Fourier-transformed infrared spectroscopy and X-ray photoelectron spectroscopy. Bio-compatibility and hemocompatibility were validated through metabolic activity and hemolysis assays. The anti-thrombotic activity was assessed using platelet aggregation, lactate dehydrogenase activation assays, and scanning electron microscopy surface imaging. The whole-blood clotting time quantification assay was employed to evaluate anticoagulation properties. Results: Results demonstrated high biocompatibility and hemocompatibility, with the most potent anti-thrombotic activity observed on pegylated surfaces. However, seleno-L-cystine and fondaparinux exhibited no anti-platelet activity. Discussion: The findings highlight the importance of balancing various factors and addressing challenges associated with different approaches when developing innovative surface modifications for cardiovascular devices.

AEE alleviates ox-LDL-induced lipid accumulation and inflammation in macrophages

Atherosclerosis is a chronic immune inflammatory disease. Aspirin eugenol ester (AEE) is a novel safe and non-toxic compound with many pharmacological effects such as anti-inflammatory, anti-hyperlipidemic and anti-thrombotic action. In order to investigate the effect of AEE on the inhibition of aortic lipid plaque formation and macrophage-derived foam cell formation induced by oxidized low density lipoprotein (ox-LDL), in vivo atherosclerosis model by feeding ApoE-/- mice with a high-fat diet and foam cells formation in vitro model by ox-LDL-induced RAW264.7 macrophages were established. It was found that AEE decreased the levels of TC and LDL-C in serum, and the plaque formation area and lipid accumulation in the aortic intima of ApoE-/- mice. In vitro studies showed that AEE could prevent the uptake of ox-LDL and reduce the contents of TC and FC in cells. AEE enhanced the cholesterol efflux by increasing the expression of ABCA1, ABCG1 and PPARγ, which effectively alleviated excess cholesterol accumulated in the cells. Meanwhile, AEE also reduced the secretion and expression of inflammatory factors in the cells. In addition, AEE could reverse the action of PPARγ inhibitor T0070907 and/or ox-LDL. Therefore, AEE may become an effective candidate drug for the prevention of atherosclerosis.

Exploiting Chemical Protein Synthesis to Study the Role of Tyrosine Sulfation on Anticoagulants from Hematophagous Organisms.

ConspectusTyrosine sulfation is a post-translational modification (PTM) that modulates function by mediating key protein-protein interactions. One of the early proteins shown to possess this PTM was hirudin, produced in the salivary glands of the medicinal leech Hirudo medicinalis, whereby tyrosine sulfation led to a ∼10-fold improvement in α-thrombin inhibitory activity. Outside of this pioneering discovery, the involvement of tyrosine sulfation in modulating the activity of salivary proteins from other hematophagous organisms was unknown. We hypothesized that the intrinsic instability of the tyrosine sulfate functionality, particularly under the acidic conditions used to isolate and analyze peptides and proteins, has led to poor detection during the isolation and/or expression of these molecules.Herein, we summarize our efforts to interrogate the functional role of tyrosine sulfation in the thrombin inhibitory and anticoagulant activity of salivary peptides and proteins from a range of different blood feeding organisms, including leeches, ticks, mosquitoes, and flies. Specifically, we have harnessed synthetic chemistry to efficiently generate homogeneously sulfated peptides and proteins for detailed structure-function studies both in vitro and in vivo.Our studies began with the leech protein hirudin P6 (from Hirudinaria manillensis), which is both sulfated on tyrosine and O-glycosylated at a nearby threonine residue. Synthetically, this was achieved through solid-phase peptide synthesis (SPPS) with a late-stage on-resin sulfation, followed by native chemical ligation and a folding step to generate six differentially modified variants of hirudin P6 to assess the functional interplay between O-glycosylation and tyrosine sulfation. A one-pot, kinetically controlled ligation of three peptide fragments was used to assemble homogeneously sulfoforms of madanin-1 and chimadanin from the tick Haemaphysalis longicornis. Dual tyrosine sulfation at two distinct sites was shown to increase the thrombin inhibitory activity by up to 3 orders of magnitude through a novel interaction with exosite II of thrombin. The diselenide-selenoester ligation developed by our lab provided us with a means to rapidly assemble a library of different sulfated tick anticoagulant proteins: the andersonins, hyalomins, madanin-like proteins, and hemeathrins, thus enabling the generation of key structure-activity data on this family of proteins. We have also confirmed the presence of tyrosine sulfation in the anticoagulant proteins of Anopheles mosquitoes (anophelins) and the Tsetse fly (TTI) via insect expression and mass spectrometric analysis. These molecules were subsequently synthesized and assessed for thrombin inhibitory and anticoagulant activity. Activity was significantly improved by the addition of tyrosine sulfate modifications and led to molecules with potent antithrombotic activity in an in vivo murine thrombosis model.The Account concludes with our most recent work on the design of trivalent hybrids that tandemly occupy the active site and both exosites (I and II) of α-thrombin, with a TTI-anophelin hybrid (Ki = 20 fM against α-thrombin) being one of the most potent protease inhibitors and anticoagulants ever generated. Taken together, this Account highlights the importance of the tyrosine sulfate post-translational modification within salivary proteins from blood feeding organisms for enhancing anticoagulant activity. This work lays the foundation for exploiting native or engineered variants as therapeutic leads for thrombotic disorders in the future.

Sarcoeleganolides H–K, new anti-thrombotic cembranes from soft coral Sarcophyton elegans

Chemical investigation of soft coral the Sarcophyton elegans collected from the South China Sea led to the identification of four new cembranes, namely sarcoeleganolides H–K (1–4). Their structures and absolute configurations were established by 1D and 2D NMR spectroscopy, quantum chemical calculations of NMR chemical shifts and electronic circular dichroism (ECD) data analysis. Moreover, compound 2 exhibited moderate anti-thrombotic activity in zebrafish.

Evaluation of the Antithrombotic Activity of Acmella oleracea L. Flower Ethanol Extract

Acmella oleracea L. (hereinafter abbreviated as AOE) is one of the plants with the potential for antithrombosis, one of the deadliest cardiovascular diseases in Indonesia. The antithrombotic activity test on AOE leaf extract revealed that it could lyse blood clots. However, no information regarding the AOE flower as an antithrombosis is provided. This study, thus, aims to determine the chromatography profile and the antithrombotic activity of the AOE flower ethanol extract. To identify the AOE chemical profile, thin-layer chromatography was carried out. Antithrombotic testing was performed on male rats of the Sprague-Dawley strain. Then, the antithrombotic activity was tested using the FeCl3-induced rat method, with the observed parameter being total occlusion time. The test animals were also divided into six groups: normal, solvent (CMC-Na 0.9%), comparator drug (clopidogrel 8.67 mg/kg), and AOE (doses 125, 250, and 375 mg/kg). The data obtained were then analyzed statistically using Kruskal-Wallis, followed by Tukey's. The TLC profile results confirmed the presence of the alkaloid compound in AOE. The authors also found that AOE at doses of 125, 250, and 375 mg/kg significantly prolonged the occlusion time comparable to that of clopidogrel at 8.67 mg/kg (p 0.05). This finding indicates that AOE has antithrombotic activity in FeCl3-induced rats.

Nemopilema nomurai jellyfish venom attenuates phenotypic modulation of PDGF BB-induced vascular smooth muscle cells and κ-carrageenan-induced rat tail thrombosis

Jellyfish venoms have long been recognized as a potentially rich source of natural bioactive compounds with pharmacological potential for the creation of innovative drugs. Our previous study demonstrated that Nemopilema nomurai jellyfish venom (NnV) has a chymotrypsin-like serine protease with fibrinolytic activity in vitro. Therefore, the present study aims to investigate the potential effect of NnV on cell migration, proliferation, and differentiation of vascular smooth muscle cells (VSMC; A7r5 cells) involved in the probable mechanism pathways. We also determined its anti-thrombotic effect through κ-carrageenan-induced Sprague–Dawley (SD) rat tail thrombus model. NnV inhibits on Platelet-derived growth factor (PDGF)-BB-stimulated A7r5 cells migration and proliferation by decreasing matrix metalloproteinase 2 (MMP-2) level and phosphorylation of ERK and Akt in a dose-dependent manner, but not p38. Furthermore, NnV regulates the phenotype transition of differentiation in PDGF-BB-stimulated A7r5 cells via ɑ-SMA and calponin in a dose-dependent manner. In an in vivo study, NnV treatment demonstrated clear anti-thrombotic activity in a dose-dependent manner, which was associated with decreased thrombus formation and length in κ-carrageenan-induced SD rat tail. These findings suggested that NnV has a novel fibrinolytic enzyme that can be used to prevent and/or treat thrombosis-related cardiovascular disorders.

Antiplatelet and thrombolytic activity of phenolic-insistent fractions from the new-fangled stem of Ficus benghalensis with concurrent GC-MS analysis

Ficus benghalensis is the ingredient of a variety of Ayurvedic herbal formulations for the management of blood-related illnesses. In the current study, the new-fangled stem ethanol extract fractions in chloroform and methanol (CFFB & MFFB) were assessed for antiplatelet, thrombolytic and toxicity studies, as well as for phytoconstituent identification GC/MS was performed. The dried powdered stem bud was extracted with 80% ethanol and successively fractionated by chloroform and methanol (CFFB & MFFB). The anti-platelet, anti-thrombotic, and thrombolytic, activity of CFFB & MFFB were tested in ex vivo mode and toxicity of methanol fraction (MFFB) was tested in in vivo. The chief feasible marker components for antiplatelet activity recognized by GC-MS in the MFFB are Diethyl phthalate, (E)-4-(3-Hydroxyprop-1-en-1-yl)-2-methoxyphenol, 7,9-Di-tert-butyl-1-oxaspiro (4,5) deca-6,9-diene-2,8-dione and trans-Sinapyl alcohol might act as irreversible cyclooxygenase inhibitors like Aspirin. In the study, at 50 ​μg/mL, the antiplatelet activity of CFFB, MFFB, and aspirin was 50.41, 82.19, and 86.34%, and a substantial adjournment in clot development was observed whereas CFFB at different dosages did not exhibit significant outcome on the adjournment of clot formation, antiplatelet, and antioxidant activity. The toxicity examination of MFFB did not confirm any substantial signs of toxicity and mortality up to 1.5 ​g/kg, b.w and non-toxic up to 1.0 ​g/kg, b.w which is capable of the comportment of atherothrombotic ailments. The MFFB exhibited anti-platelet, anti-thrombotic, thrombolytic, and anti-oxidant activity, and capacity to prevent cardiovascular disorders without causing toxicity.

Aspirin 15cH has Different Effects on Morphology and Function of Lipopolysaccharide-Challenged RAW 264.7 Macrophages In Vitro Compared to a Pharmacological Dose of Aspirin.

Aspirin is one of the most commonly used drugs worldwide. It is known to present antipyretic, anti-inflammatory and anti-thrombotic actions, making it extremely useful in a wide range of clinical contexts. Interestingly, homeopathically prepared Aspirin 15cH has been found to have a pro-thrombotic effect in rats, raising the hypothesis that Aspirin 15cH could also modulate the activity of inflammatory cells in different pathological processes. Our objective was to assess what effect Aspirin 15cH has on RAW 264.7 macrophages in vitro. The effects of Aspirin 15cH on biochemical and morphological activities of lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages were evaluated. These effects were compared with unchallenged macrophages (negative control), untreated LPS-stimulated macrophages, macrophages treated with succussed water (vehicle control), or aspirin 200 µg/mL (pharmacological inhibitor of LPS activity). Cell morphology (adhered cell area and cytoskeleton arrangements), cell viability, toll-like receptor-4 (TLR-4) expression, and the production of nitric oxide, cytokines and intracellular reactive oxygen species were assessed. Aspirin 15cH reduced the number of cells expressing TLR-4 on the surface (p = 0.03) and induced a "columnar" morphology of macrophage pseudopods, indicating changes in cytoskeleton arrangement. When cells were treated with both Aspirin 15cH and LPS, cell morphology became heterogeneous, suggesting that sub-populations of cells had differing sensitivities to LPS or Aspirin 15cH. Exposure of the cells to LPS alone, succussed water or aspirin 200 µg/mL produced effects consistent with the literature. Aspirin 15cH, aspirin 200 µg/mL, LPS and succussed water appear to act as independent stimuli able to induce different patterns of macrophage response. Aspirin 15cH induced changes suggestive of M2 polarization of the macrophages (i.e., toward a wound healing or tissue repair, rather than inflammatory, phenotype). These preliminary findings need to be confirmed in further specific studies.

Integrated omics approach for the identification of HDL structure-function relationships in PCSK9-related familial hypercholesterolemia

The role of proprotein convertase subtilisin/kexin type 9 (PCSK9) in dyslipidemia may go beyond its immediate effects on low-density lipoprotein receptor (LDL-R) activity. This study aimed to assess PCSK9-derived alterations of high-density lipoprotein (HDL) physiology, which bear a potential to contribute to cardiovascular risk profile. HDL was isolated from 33 patients with familial autosomal dominant hypercholesterolemia (FH), including those carrying PCSK9 gain-of-function (GOF) genetic variants (FH-PCSK9, n=11), together with two groups of dyslipidemic patients employed as controls and carrying genetic variants in the LDL-R not treated (ntFH-LDLR, n=11) and treated (tFH-LDLR, n=11) with statins, and 11 normolipidemic controls. Biological evaluations paralleled by proteomic, lipidomic and glycomic analyses were applied to characterize functional and compositional properties of HDL. Multiple deficiencies in the HDL function were identified in the FH-PCSK9 group relative to dyslipidemic FH-LDLR patients and normolipidemic controls, which involved reduced antioxidative, antiapoptotic, anti-thrombotic and anti-inflammatory activities. By contrast, cellular cholesterol efflux capacity of HDL was unchanged. In addition, multiple alterations of the proteomic, lipidomic and glycomic composition of HDL were found in the FH-PCSK9 group. Remarkably, HDLs from FH-PCSK9 patients were systematically enriched in several lysophospholipids as well as in A2G2S2 (GP13) glycan and apolipoprotein A-IV. Based on network analysis of functional and compositional data, a novel mosaic structure-function model of HDL biology involving FH was developed. Several metrics of anti-atherogenic HDL functionality are altered in FH-PCSK9 patients paralleled by distinct compositional alterations. These data provide a first-ever overview of the impact of GOF PCSK9 genetic variants on structure-function relationships in HDL.

The choice of a modern neuroprotective drug as a basic strategy in the treatment of chronic cerebrovascular diseases

The topicality of the issues of effective treatment and prevention of cerebrovascular diseases (CVD) is not doubtful in view of their leadership in the top ranked causes of high mortality and disability in the general population almost all over the world, including Russia. The pathogenetic aspects of chronic CVD caused by hypoperfusion, which triggers a chain of successively developing metabolic disorders, represent a potential therapeutic target for the development of new dosage forms. The enormous clinical experience has been accumulated and evidence of therapeutic benefits in patients with various types of vascular and metabolic disorders accompanied by ischemia, including cerebrovascular disease, has been obtained over almost thirty years of active clinical use of ethylmethylhydroxypyridine succinate (6-methyl-2-ethylpyridin-3-ol butanedioate, EMHPS). It proved that EMHPS has such important effects as antioxidant, membrane-protective, antihypoxic, and anti-ischemic properties, as well as confirmed its hypoglycemic, hypolipidemic, antiplatelet and antithrombotic actions, which makes it an important component of the complex therapy of CVD. Progress in the development of innovative drugs is the result, on the one hand, of the availability of highly active pharmacological substances and, on the other hand, of the creation or selection of new dosage forms. The use of matrix tablets as a modern dosage form allowed us to efficiently optimize the pharmacokinetics of EMHPS, which led to the creation of a drug with a high predictive efficacy and safety. Thus, the developed new prolonged dosage form of EMHPS enables a constant-rate daily release of the active substance for 20 hours and reduces the likelihood of not only side effects, but also skipped intake during the day. The new dosage form of EMHPS is available at 375 and 750 mg doses, which makes it possible to take just 1 tablet per day and get all the therapeutic benefits while staying compliant.

Study on Chemical Constituents of the Enriched Extracts of Erigeron multiradiatus and Anti-thrombotic Effect on Zebrafish

Background Erigeron multiradiatus (Lindl.) Benth. has been used for years to treat various diseases in Traditional Tibetan Medicine (TTM). Our previous studies have shown that E. multiradiatus extract has important pharmacological effects, such as anti-myocardial ischemia–reperfusion (I/R) injury, anti-inflammatory, and anti-diabetic effects. To date, there have been no reports on using this extract to treat thrombosis. Objectives The aim of this study was to enrich the active components of E. multiradiatus and to further investigate its anti-thrombotic activity. Materials and Methods We used D101 macroporous adsorption resin to enrich the main active substances of E. multiradiatus and performed quantitative determination using ultra-high-performance liquid chromatography with a photodiode array detector (UHPLC-PDA). Furthermore, we investigated the anti-thrombotic effects of E. multiradiatus using three models: an arachidonic acid-induced platelet aggregation thrombotic zebrafish model, a phenylhydrazine-induced red blood cell (RBC)-injuring thrombotic zebrafish model, and a ponatinib-induced vascular endothelial cell (VEC)-injuring thrombotic zebrafish model. Results The yield of active component enrichment of E. multiradiatus was 4.95% using the D101 resin. We accurately determined eight compounds in the extract. Our results showed that these extracts had significant anti-thrombotic effects in the three different zebrafish models. Conclusion The enriched extract of E. multiradiatus could be used as a natural anti-thrombotic agent. Our results will provide a reference for reasonable clinical applications of E. multiradiatus.

Isolation and Characterization of Poeciguamerin, a Peptide with Dual Analgesic and Anti-Thrombotic Activity from the Poecilobdella manillensis Leech

When Poecilobdella manillensis attacks its prey, the prey bleeds profusely but feels little pain. We and other research teams have identified several anticoagulant molecules in the saliva of P. manillensis, but the substance that produces the paralyzing effect in P. manillensis is not known. In this study, we successfully isolated, purified, and identified a serine protease inhibitor containing an antistasin-like domain from the salivary secretions of P. manillensis. This peptide (named poeciguamerin) significantly inhibited elastase activity and slightly inhibited FXIIa and kallikrein activity, but had no effect on FXa, trypsin, or thrombin activity. Furthermore, poeciguamerin exhibited analgesic activity in the foot-licking and tail-withdrawal mouse models and anticoagulant activity in the FeCl3-induced carotid artery thrombosis mouse model. In this study, poeciguamerin was found to be a promising elastase inhibitor with potent analgesic and antithrombotic activity for the inhibition of pain and thrombosis after surgery or in inflammatory conditions.

First-in-human study to assess the safety, pharmacokinetics, and pharmacodynamics of BMS-986141, a novel, reversible, small-molecule, PAR4 agonist in non-Japanese and Japanese healthy participants

BMS-986141 is a novel, oral, protease-activated, receptor 4 (PAR4)-antagonist that exhibited robust antithrombotic activity and low bleeding risk in preclinical studies. The pharmacokinetic, pharmacodynamic, and tolerability profiles of BMS-986141 in healthy participants were assessed in a randomized, double-blind, placebo-controlled, single-ascending-dose (SAD; N = 60) study; a multiple-ascending-dose (MAD; N = 32) study; and a Japanese MAD (JMAD; N = 32) study. Exposure was dose-proportional for BMS-986141 2.5 mg and 150 mg; maximum concentrations were 17.6 ng/mL and 958 ng/mL; and areas under the curve (AUC) to infinity were 183 h* × ng/mL and 9207 h* × ng/mL, respectively. Mean half-life ranged from 33.7 to 44.7 hours across dose panels. The accumulation index following once-daily administration for 7 days suggested a 1.3- to 2-fold AUC increase at steady state. In the SAD study, BMS-986141 75 and 150 mg produced ≥80% inhibition of 25–100 µM PAR4 agonist peptide (AP)-induced platelet aggregation, without affecting PAR1-AP-induced platelet aggregation, through ≥24 hours postdose. In the MAD and JMAD studies, BMS-986141 doses ≥10 mg completely inhibited 12.5 μM and 25 μM PAR4-AP-induced platelet aggregation through 24 hours. This study found BMS-986141 was safe and well tolerated, with dose-proportional pharmacokinetics and concentration-dependent pharmacodynamics in healthy participants over a wide dose range. ClinicalTrials.gov ID: NCT02341638.

In Vitro Antioxidant, Anti-inflammatory, Antimicrobial, and Antidiabetic Activities of Synthesized Chitosan-loaded p-Coumaric Acid Nanoparticles.

p-Coumaric acid is a phenolic compound widely distributed in fruits and vegetables that displays an array of therapeutic properties, including antidiabetic effects. Prominent application in diabetes is limited due to its suboptimal pharmacokinetics, poor aqueous solubility, and poor bioavailability. Nanotechnology-based delivery methods have been developed to address these limitations and improve the therapeutic uses of p-coumaric acid, and the nanoencapsulation method is emerging as a feasible alternative. The objective of this study is to synthesize p-coumaric acid nanoparticles (PCNPs) and to evaluate their In Vitro activities. The PCNPs were synthesized by the nanoprecipitation method and characterized by UV-visible spectroscopy, zeta potential, Fourier-transform infrared (FTIR) spectroscopy, and scanning electron microscopy (SEM) with dispersive energy X-ray (EDX) analysis. In addition, the PCNPs were analyzed for In Vitro antioxidant activity using six different free radical scavenging assays and were also analyzed for antimicrobial, anti-inflammatory, antithrombotic, and antidiabetic effects. The formation of PCNPs was confirmed by UV-visible spectra at 283 nm, and FTIR analysis revealed the reduction and capping of the chitosan nanoparticles. SEM was used to assess the size and shape of the PCNPs, and the high absorption property of the PCNPs was investigated using EDX analysis. The PCNPs had significant antioxidant, hydrogen peroxide (H2O2), lipid peroxidation (LPO), superoxide and nitric oxide (NO) radical scavenging power activities, and showed potent antimicrobial, anti-inflammatory, antithrombotic, and antidiabetic activities. The present study suggests that PCNPs can be used as a potential medication delivery approach to provide a greater nephroprotective effect in the treatment of diabetic nephropathy. To the best of our knowledge, this is the first attempt at the synthesis of chitosan-loaded PCNPs.