scholarly journals Pharmacophore Modeling, Docking, and Molecular Dynamics Simulation of Flavonoids as Inhibitors of Urokinase-type Plasminogen Activator

2022 ◽  
Vol 53 (3) ◽  
pp. 451-465
Author(s):  
Daryono Hadi Tjahjono ◽  
Bina Lohita Sari ◽  
Slamet Ibrahim

The urokinase-type plasminogen activator (uPA) system plays a significant role in the invasion and metastasis of cancer cells. The present study was conducted to investigate natural product compounds as inhibitors and hit molecules of uPA using in-silico analysis. A pharmacophore model was built to screen the Indonesian Herbal Database (HerbalDB) to obtain inhibitors of different scaffolds. Based on the molecular docking score, four ligands were selected as potential uPA inhibitors. Subsequently, the stability of the ligand-uPA complex was analyzed using molecular dynamics (MD) simulation. An RMSD graph of the backbone protein and the RMSF values of the amino acid residues were also determined. In addition, the MM-PBSA method was applied to calculate the free binding energy. According to the results, Model_3, characterized by aromatic rings 23 (F1 and F2), cationic H-bond donor (F3), and metal ligator (F4) features, had an adequate goodness-of-hit score (GH). The four top-ranked ligands, isorhamnetin, rhamnetin, quercetin, and kaempferol, showed higher docking scores compared to the others. This study confirmed that isorhamnetin, rhamnetin, and kaempferol build stable complexes with uPA with lower binding energy than quercetin.

Author(s):  
Sheema Jb ◽  
Waheeta Hopper

  Objective: Glycogen synthase kinase 3 beta (GSK3β) is one of the main targets for wound healing activity. Our objective is to identify novel inhibitors for GSK3β using in silico approach.Methods: Grid-based molecular docking, energy-based pharmacophore (e-pharmacophore) modeling, and molecular dynamics (MD) studies were performed for phytocompounds with GSK3β and compared with standard drugs using Schrodinger software.Results: The glide scores and the molecular interactions of the phytocompounds were well comparable to the standard drugs. The MD was performed for the target bound to the best scoring ligand, entagenic acid. The pharmacophore features of this docked complex were modeled as e-pharmacophore. The constructed e-pharmacophore model was screened against phytocompounds retrieved from literature to identify the ligands with similar pharmacophore features.Conclusion: The glide scores of fukinolic acid, cimicifugic acid, and linarin were −10.99, −8.28, and −7.25 kcal/mol, respectively. The further 50 nanoseconds MD study determined the stability of GSK3β-linarin complex. Nitrofurazone and sulfathiazole drugs can lead to systemic side effects. Hence, it is concluded that linarin could be a potent wound healing compound against GSK3β.


2015 ◽  
Vol 93 (11) ◽  
pp. 1199-1206 ◽  
Author(s):  
Ludi Jiang ◽  
Yong Li ◽  
Liansheng Qiao ◽  
Xi Chen ◽  
Yusu He ◽  
...  

mGluR5, which belongs to the G-protein-coupled receptor superfamily, is believed to be associated with many human diseases, such as a wide range of neurological disorders, gastroesophageal reflux disease, and cancer. Comparing with compounds that target on the orthosteric binding site, significant roles have been established for mGluR5 negative allosteric modulators (NAMs) due to their higher subtype selectivity and more suitable pharmacokinetic profiles. Nevertheless, to date, none of them have come to market for various reasons. In this study, a 3D quantitative pharmacophore model was generated by using the HypoGen module in Discovery Studio 4.0. With several validation methods ultilized, the optimal pharmacophore model Hypo2 was selected to discover potential mGluR5 NAMs from natural products. Two hundred and seventeen potential NAMs were obtained after being filtered by Lipinski’s rule (≥4). Then, molecular docking was used to refine the pharmacophore-based screening results and analyze the binding mode of NAMs and mGluR5. Three compounds, aglaiduline, 5-O-ethyl-hirsutanonol, and yakuchinone A, with good ADMET properties, acceptable Fit value and estimated value, and high docking score, were reserved for a molecular dynamics simulation study. All of them have stability of ligand binding. From our computational results, there might exhibit drug-like negative allosteric moderating effects on mGluR5 in these natural products. This work provides a reliable method for discovering mGluR5 NAMs from natural products.


Marine Drugs ◽  
2021 ◽  
Vol 20 (1) ◽  
pp. 29
Author(s):  
Lianxiang Luo ◽  
Ai Zhong ◽  
Qu Wang ◽  
Tongyu Zheng

Background: In the past decade, several antibodies directed against the PD-1/PD-L1 interaction have been approved. However, therapeutic antibodies also exhibit some shortcomings. Using small molecules to regulate the PD-1/PD-L1 pathway may be another way to mobilize the immune system to fight cancer. Method: 52,765 marine natural products were screened against PD-L1(PDBID: 6R3K). To identify natural compounds, a structure-based pharmacophore model was generated, following by virtual screening and molecular docking. Then, the absorption, distribution, metabolism, and excretion (ADME) test was carried out to select the most suitable compounds. Finally, molecular dynamics simulation was also performed to validate the binding property of the top compound. Results: Initially, 13 small marine molecules were screened based on the pharmacophore model. Then, two compounds were selected for further evaluation based on the molecular docking scores. After ADME and toxicity studies, molecule 51320 was selected for further verification. By molecular dynamics analysis, molecule 51320 maintains a stable conformation with the target protein, so it has the chance to become an inhibitor of PD-L1. Conclusions: Through structure-based pharmacophore modeling, virtual screening, molecular docking, ADMET approaches, and molecular dynamics (MD) simulation, the marine natural compound 51320 can be used as a small molecule inhibitor of PD-L1.


2001 ◽  
Vol 120 (5) ◽  
pp. A599-A600 ◽  
Author(s):  
L HERSZENYI ◽  
F FARINATI ◽  
G ISTVAN ◽  
M PAOLI ◽  
G ROVERONI ◽  
...  

1994 ◽  
Vol 71 (01) ◽  
pp. 134-140 ◽  
Author(s):  
S Ueshima ◽  
P Holvoet ◽  
H R Lijnen ◽  
L Nelles ◽  
V Seghers ◽  
...  

SummaryIn an effort to modify the fibrinolytic and/or pharmacokinetic properties of recombinant low M r single-chain urokinase-type plasminogen activator (rscu-PA-32k), mutants were prepared by site-directed mutagenesis of clusters of charged amino acids with the highest solvent accessibility. The following mutants of rscu-PA-32k were prepared: LUK-2 (Lys 212, Glu 213 and Asp 214 to Ala), LUK-3 (Lys 243 and Asp 244 to Ala), LUK-4 (Arg 262, Lys 264, Glu 265 and Arg 267 to Ala), LUK-5 (Lys 300, Glu 301 and Asp 305 to Ala) and LUK-6 (Arg 400, Lys 404, Glu 405 and Glu 406 to Ala).The rscu-PA 32k moictic3 were expressed in High Five Ttichoplasiani cells, and purified to humugciicily from the conditioned cell culture medium, with recoveries of 0.8 to 3.7 mg/1. The specific fibrinolytic activities (220,000 to 300,000 IU/mg), the rates of plasminogen activation by the single-chain moieties and the rates of conversion In lwo chain moieties by plasmin were comparable for mutant and wild-type rscu PA 32k moieties, with the exception of LUK-5 which was virtually inactive. Equi-effective lysis (50% in 2 h) of 60 pi 125I-fibrin labeled plasma clots submerged in 0.5 ml normal human plasma was obtained with 0.7 to 0.8 μg/ml of wild-type or mutant rscu-PA-3?.k, except with LUK-5 (no significant lysis with 16 pg/ml). Following bolus injection in hamsters, all rscu-PA-32k moieties had a comparably rapid plasma clearance (1.3 to 2.7 ml/min), as a result of a short initial half-life (1.4 to 2.5 min). In hamsters with pulmonary embolism, continuous intravenous infusion over 60 min at a dose of 1 mg/kg, resulted in 53 to 72% clot lysis with the mutants, but only 23% with LUK-5, as compared to 36% for wild-type rscu-PA-32k.These data indicate that clustered charge-to-alanine mutants of rscu-PA-32k, designed to eliminate charged regions with the highest solvent accessibility, do not have significantly improved functional, fibrinolytic or pharmacokinetic properties.


1988 ◽  
Vol 60 (02) ◽  
pp. 247-250 ◽  
Author(s):  
H R Lijnen ◽  
L Nelles ◽  
B Van Hoef ◽  
F De Cock ◽  
D Collen

SummaryRecombinant chimaeric molecules between tissue-type plasminogen activator (t-PA) and single chain urokinase-type plasminogen activator (scu-PA) or two chain urokinase-type plasminogen activator (tcu-PA) have intact enzymatic properties of scu-PA or tcu-PA towards natural and synthetic substrates (Nelles et al., J Biol Chem 1987; 262: 10855-10862). In the present study, we have compared the reactivity with inhibitors of both the single chain and two chain variants of recombinant u-PA and two recombinant chimaeric molecules between t-PA and scu-PA (t-PA/u-PA-s: amino acids 1-263 of t-PA and 144-411 of u-PA; t-PA/u-PA-e: amino acids 1-274 of t-PA and 138-411 of u-PA). Incubation with human plasma in the absence of a fibrin clot for 3 h at 37° C at equipotent concentrations (50% clot lysis in 2 h), resulted in significant fibrinogen breakdown (to about 40% of the normal value) for all two chain molecules, but not for their single chain counterparts. Preincubation of the plasminogen activators with plasma for 3 h at 37° C, resulted in complete inhibition of the fibrinolytic potency of the two chain molecules but did not alter the potency of the single chain molecules. Inhibition of the two chain molecules occurred with a t½ of approximately 45 min. The two chain variants were inhibited by the synthetic urokinase inhibitor Glu-Gly-Arg-CH2CCl with apparent second-order rate constants of 8,000-10,000 M−1s−1, by purified α2-antiplasmin with second-order rate constants of about 300 M−1s−1, and by plasminogen activator inhibitor-1 (PAI-1) with second-order rate constants of approximately 2 × 107 M−1s−1.It is concluded that the reactivity of single chain and two chain forms of t-PA/u-PA chimaers with inhibitors is very similar to that of the single and two chain forms of intact u-PA.


1991 ◽  
Vol 65 (01) ◽  
pp. 082-086 ◽  
Author(s):  
G Dooijewaard ◽  
A de Boer ◽  
P N C Turion ◽  
A F Cohen ◽  
D D Breimer ◽  
...  

SummaryThe enhancement of the blood fibrinolytic potential by physical exercise is generally attributed to the release of tissue-type plasminogen activator (t-PA) from the vessel wall. In this study we have investigated the possible contribution of urokinase-type plasminogen activator (u-PA).Six healthy male volunteers (age 21–25 years) were screened for their ability to perform maximal exercise for their age-group for 12 min on a bicycle ergometer. Subsequently, on one occasion they were required to remain supine for 2 h (from 8.30 a. m. onwards) and on another they performed maximal exercise (from 9.00 a.m. onwards). During exercise an increase in u-PA antigen and plasmin-activatable pro-urokinase (proUK) activity, concurrent with t-PA antigen and euglobulin t-PA activity, was observed in all six volunteers, while at rest these parameters remained unaffected. Mean u-PA- and t-PA antigen increased, respectively, from 4.2 ± 1.0 ng/ml and 5.8 ± 2.1 ng/ml before exercise to 9.8 ± 3.0 ng/ml and 18.3 ± 3.8 ng/ml (peak). Mean plasminactivatable proUK activity and t-PA activity increased, respectively, from 2.1 ± 0.4 ng/ml and 0.3 ± 0.2 ng/ml before exercise to 4.3 ± 1.7 ng/ml and 7.2 ± 4.0 ng/ml (peak). The increases were statistically significant throughout (paired t-test, pre vs post, antigen P <0.005 and activity P <0.02). After cessation of exercise u-PA and t-PA declined concurrently to normal values with a 50"/" decay in about 5 min. In conclusion, we found that both u-PA antigen and plasmin-activatable proUK activity are, concurrently with t-PA, enhanced upon exercise and, therefore, we consider that u-PA also contributes to – and co-operates in – the enhancement of the blood fibrinolytic potential and activity under these conditions.


1992 ◽  
Vol 67 (01) ◽  
pp. 095-100 ◽  
Author(s):  
Paul J Declerck ◽  
Leen Van Keer ◽  
Maria Verstreken ◽  
Désiré Collen

SummaryAn enzyme-linked immunosorbent assay (ELISA) for quantitation of natural and recombinant plasminogen activators containing the serine protease domain (B-chain) of urokinase-type plasminogen activator (u-PA) was developed, based on two murine monoclonal antibodies, MA-4D1E8 and MA-2L3, raised against u-PA and reacting with non-overlapping epitopes in the B-chain. MA-4D1E8 was coated on microtiter plates and bound antigen was quantitated with MA-2L3 conjugated with horseradish peroxidase. The intra-assay, inter-assay and inter-dilution coefficients of variation of the assay were 6%, 15% and 9%, respectively. Using recombinant single-chain u-PA (rscu-PA) as a standard, the u-PA-related antigen level in normal human plasma was 1.4 ± 0.6 ng/ml (mean ± SD, n = 27).The ELISA recognized the following compounds with comparable sensitivity: intact scu-PA (amino acids, AA, 1 to 411), scu-PA-32k (AA 144 to 411), a truncated (thrombin-derived) scu-PA comprising A A 157 to 411, and chimeric t-PA/u-PA molecules including t-PA(AA1-263)/scu-PA(AA144-411), t-PA(AA1-274)/scu-PA(AA138-411) and t-PA(AA87-274)/scu-PA(AA138-411). Conversion of single-chain to two-chain forms of u-PA or inhibition of active two-chain forms with plasminogen activator inhibitor-1 or with the active site serine inhibitor phenyl-methyl-sulfonyl fluoride, did not alter the reactivity in the assay. In contrast, inactivation with α2-antiplasmin or with the active site histidine inhibitor Glu-Gly-Arg-CH2Cl resulted in a 3- to 5-fold reduction of the reactivity. When purified scu-PA-32k was added to pooled normal human plasma at final concentrations ranging from 20 to 1,000 ng/ml, recoveries in the ELISA were between 84 and 110%.The assay was successfully applied for the quantitation of pharmacological levels of scu-PA and t-PA(AA87_274)/scu-PA(AA138-411) in plasma during experimental thrombolysis in baboons.Thus the present ELISA, which is specifically dependent on the presence of the serine protease part of u-PA, is useful for measurement of a wide variety of variants and chimeras of u-PA which are presently being developed for improved thrombolytic therapy.


1996 ◽  
Vol 75 (06) ◽  
pp. 933-938 ◽  
Author(s):  
Marten Fålkenberg ◽  
Johan Tjärnstrom ◽  
Per Örtenwall ◽  
Michael Olausson ◽  
Bo Risberg

SummaryLocal fibrinolytic changes in atherosclerotic arteries have been suggested to influence plaque growth and promote mural thrombosis on ruptured or ulcerated plaques. Increased levels of plasminogen activator inhibitor (PAI-1) have been found in atherosclerotic arteries. In this study tissue plasminogen activator (t-PA), urokinase-type plasminogen activator (u-PA) and PAI-1 were localized in arterial biopsies of healthy and atherosclerotic vessels by immunohistochemis-try. The expression of fibrinolytic regulators was related to the distribution of endothelial cells (EC) and macrophages. Results: t-PA was expressed in vasa vasorum. PAI-1 was positive in endothelial cells, in the media and in the adventitia. Increased expression of t-PA, u-PA and PAI-1 was found in atherosclerotic vessels. t-PA, u-PA, PAI-1 and macrophages were co-localized in plaques. These results support the concept that macrophages can be important in the local regulation of fibrinolysis in atherosclerotic vessels.


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