Discovery of novel EGFR tyrosine kinase inhibitors by structure-based virtual screening

By using of structure-based virtual screening, 13 novel epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors were discovered from 197,116 compounds in the SPECS database here. Among them, 8 compounds significantly inhibited EGFR kinase activity with IC(50) values lower than 10 μM. 3-{[1-(3-Chloro-4-fluorophenyl)-3,5-dioxo-4-pyrazolidinylidene]methyl}phenyl 2-thiophenecarboxylate (13), particularly, was the most potent inhibitor possessing the IC(50) value of 3.5 μM. The docking studies also provide some useful information that the docking models of the 13 compounds are beneficial to find a new path for designing novel EGFR inhibitors.     

Discovery of potent inhibitors of receptor protein tyrosine phosphatase sigma through the structure-based virtual screening

Receptor protein tyrosine phosphatase sigma (PTPσ) has proved to be a promising target for the development of therapeutics for the treatment of neurological diseases. Here, we report the first example for a successful application of the structure-based virtual screening to identify the novel small-molecule inhibitors of PTPσ. These inhibitors revealed high potencies with the associated IC₅₀ values ranging from 0.1 to 1.3 μM and were also screened for having desirable physicochemical properties as a drug candidate. Therefore, they deserve consideration for further development by structure–activity relationship studies to develop therapeutics for neurological diseases. Structural features relevant to the stabilization of the newly identified inhibitors in the active site of PTPσ are discussed in detail.     

Discovery of VEGFR inhibitors through virtual screening and energy assessment

Vascular endothelial growth factor receptor-2 (VEGFR-2) is crucial in promoting tumor angiogenesis and cancer metastasis. Thus, inhibition of VEGFR-2 has appeared as a good tactic for cancer treatment. To find out novel VEGFR-2 inhibitors, first, the PDB structure of VEGFR-2, 6GQO, was selected based on atomic nonlocal environment assessment (ANOLEA) and PROCHECK assessment. 6GQO was then further used for structure-based virtual screening (SBVS) of different molecular databases, including US-FDA approved drugs, US-FDA withdrawn drugs, may bridge, MDPI, and Specs databases using Glide. Based on SBVS, receptor fit, drug-like filters, and absorption, distribution, metabolism, excretion, and toxicity (ADMET) analysis of 427877 compounds, the best 22 hits were selected. From the 22 hits, hit 5 complex with 6GQO was put through molecular mechanics/generalized born surface area (MM/GBSA) study and hERG binding. The MM/GBSA study revealed that hit 5 possesses lesser binding free energy with more inferior stability in the receptor pocket than the reference compound. The VEGFR-2 inhibition assay of hit 5 disclosed an IC₅₀ of 165.23 nM against VEGFR-2, which can be possibly enhanced through structural modifications.     

Identification of novel inhibitors of BCR-ABL tyrosine kinase via virtual screening

Inhibition of BCR-ABL tyrosine kinase activity has shown to be essential for the treatment of chronic myelogenous leukemia (CML). However, drug resistance has quickly arisen in recent clinical trials for STI571 (Gleevec), which is the first approved drug of CML by inhibiting ABL tyrosine kinase. It is desirable to develop new types of ABL tyrosine kinase inhibitors that may overcome this drug resistance problem. Here we present the discovery of novel inhibitors targeted at the catalytic domain of ABL tyrosine kinase by using three-dimensional database searching techniques. From a database containing 200,000 commercially available compounds, the top 1000 compounds with the best DOCK energy score were selected and subjected to structural diversity and drug likeness analysis, 15 compounds were submitted for biological assay. Eight out of the 15 showed inhibitory activity against K562 cells with IC(50) value ranging from 10 to 200 microM. Two promising compounds showed inhibition in further ABL tyrosine phosphorylation assay. It is anticipated that those two compounds can serve as lead compounds for further drug design and optimization.     

Pharmacophore modeling study based on known Spleen tyrosine kinase inhibitors together with virtual screening for identifying novel inhibitors

In this investigation, chemical features based 3D pharmacophore models were developed based on the known inhibitors of Spleen tyrosine kinase (Syk) with the aid of hiphop and hyporefine modules within catalyst. The best quantitative pharmacophore model, Hypo1, was used as a 3D structural query for retrieving potential inhibitors from chemical databases including Specs, NCI, MayBridge, and Chinese Nature Product Database (CNPD). The hit compounds were subsequently subjected to filtering by Lipinski’s rule of five and docking studies to refine the retrieved hits. Finally 30 compounds were selected from the top ranked hit compounds and conducted an in vitro kinase inhibitory assay. Six compounds showed a good inhibitory potency against Syk, which have been selected for further investigation.     

Pharmacophore modeling and virtual screening to identify potential RET kinase inhibitors

Chemical features based 3D pharmacophore model for REarranged during Transfection (RET) tyrosine kinase were developed by using a training set of 26 structurally diverse known RET inhibitors. The best pharmacophore hypothesis, which identified inhibitors with an associated correlation coefficient of 0.90 between their experimental and estimated anti-RET values, contained one hydrogen-bond acceptor, one hydrogen-bond donor, one hydrophobic, and one ring aromatic features. The model was further validated by a testing set, Fischer’s randomization test, and goodness of hit (GH) test. We applied this pharmacophore model to screen NCI database for potential RET inhibitors. The hits were docked to RET with GOLD and CDOCKER after filtering by Lipinski’s rules. Ultimately, 24 molecules were selected as potential RET inhibitors for further investigation.     

Ligand-based and e-pharmacophore modeling, 3D-QSAR and hierarchical virtual screening to identify dual inhibitors of spleen tyrosine kinase (Syk) and janus kinase 3 (JAK3)

The clinical efficacy of multiple kinase inhibitors has caught the interest of Pharmaceutical and Biotech researchers to develop potential drugs with multi-kinase inhibitory activity for complex diseases. In the present work, we attempted to identify dual inhibitors of spleen tyrosine kinase (Syk) and janus kinase 3 (JAK3), keys players in immune signaling, by developing ideal pharmacophores integrating Ligand-based pharmacophore models (LBPMs) and Structure-based pharmacophore models (SBPMs), thereby projecting the optimum pharmacophoric required for inhibition of both the kinases. The four point LBPM; ADPR.14 suggested the presence of one hydrogen bond acceptor, one hydrogen bond donor, one positive ionizable, and one ring aromatic feature for Syk inhibitory activity and AADH.54 proposed the necessity of two hydrogen bond acceptor, one hydrogen bond donor, and one hydrophobic feature for JAK3 inhibitory activity. To our interest, SBPMs identified additional ring aromatic features required for inhibition of both the kinases. For Syk inhibitory activity, the hydrogen bond acceptor feature indicated by LBPM was devoid of forming hydrogen bonding interaction with the hinge region amino acid residue (Ala451). Thus merging the information revealed by both LBPMs and SBPMs, ideal pharmacophore models i.e. ADPRR.14 (Syk) and AADHR.54 (JAK3) were generated. These models after rigorous statistical validation were used for screening of Asinex database. The systematic virtual screening protocol, including pharmacophore and docking-based screening, ADME property, and MM-GBSA energy calculations, retrieved final 10 hits as dual inhibitors of Syk and JAK3. Final 10 hits thus obtained can aid in the development of potential therapeutic agents for autoimmune disorders. Also the top two hits were evaluated against both the enzymes.     

Identification of potential Leptospira phosphoheptose isomerase inhibitors through virtual high-throughput screening

The hfe-threatening infections caused by Leptospira serovars demand the need for designing anti-leptospirosis drugs.The present study encompasses exploring inhibitors against phosphoheptose isomerase(GmhA)of Leptospira,which is vital for lipopolysaccharide(LPS)biosynthesis and is identified as a common drug target through the subtractive genomic approach.GmhA model was built in Modeller 9v7.Structural refinement and energy minimization of the predicted model was carried out using Maestro 9.0.The refined model reliability was assessed through Procheck,ProSA,ProQ and Profile 3D.The substrate-based virtual high-throughput screening(VHTS)in Ligand.Info Meta-Database tool generated an in-house library of 354 substrate structural analogs.Furthermore,structure-based VHTS from the in-house library with different conformations of each ligand provided 14 novel competitive inhibitors.The model together with insight gained from the VHTS would be a promising starting point for developing anti-leptospirosis competitive inhibitors targeting LPS biosynthesis pathway.     

Discovery of small molecule inhibitors of integrin alphavbeta3 through structure-based virtual screening

Inhibitors of integrin alphavbeta3 have been implicated in the treatment of a variety of diseases, including tumor metastasis, neovascularization, osteoporosis, and rheumatoid arthritis. It is therefore desirable to develop new types of small molecule inhibitors of integrin alphavbeta3. Here we describe the discovery of novel classes of small molecule inhibitors, via structure-based virtual screening, that target the ligand binding site of integrin alphavbeta3. Application of the docking procedure for screening of a commercially available compound database resulted in a 1774-fold reduction in the size of the screening set (88695 to 50 compounds) and gave a hit-rate of 14% upon biological evaluation (IC50 value ranging from 30 to 200 microM). The best hit, compound 37, 3,4-dichloro-phenylbiguanide, showed inhibitory activity, in a time- and dose-dependent manner, in both cell motility and angiogenesis assays. Based on the best hit, compound 37, a more effective derivative compound 62 has been identified. Furthermore, molecular graphics analyses of a series of substituted phenylbiguanides were carried out to predict the binding mode between the active compounds and integrin alphavbeta3. Our results indicate that the substituted phenylbiguanides might be involved in the inhibition of bivalent cation-mediated ligand binding of integrin alphavbeta3.     

Discovery of 9-phenylacridinediones as highly selective butyrylcholinesterase inhibitors through structure-based virtual screening

Butyrylcholinesterase (BuChE) is considered a promising drug target as it plays an important role in the progression of late stage Alzheimer’s disease (AD). Two compound libraries were selected and 64 124 amine containing moieties were screened using a hierarchical virtual screening protocol to discover new selective BuChE inhibitors. From these and subsequent docking experiments, 9-phenylacridinedione (9-PAD) was identified as a promising scaffold for selective inhibition of BuChE. Selected top dock scored 9-PADs were assayed and compounds 3 and 6 exhibited potent and highly selective human BuChE inhibition (IC₅₀: 98 nM and 142 nM, respectively). Both molecules were also predicted to show sufficient brain permeability, not have any substantial toxicities, especially hepatotoxicity, and no significant in vitro cytotoxicity against SH-SY5Y neuroblastoma cells at concentrations up to 100 µM. These findings indicate that 9-PAD is a promising lead structure for the development of agents able to treat late stage AD.     

Discovery of Entamoeba histolytica hexokinase 1 inhibitors through homology modeling and virtual screening

Abstract

Entamoeba histolytica, the parasite which causes amebiasis is responsible for 110?000 deaths a year. Entamoeba histolytica depends on glycolysis to obtain ATP for cellular work. According to metabolic flux studies, hexokinase exerts the highest flux control of this metabolic pathway; therefore, it is an excellent target in the search of new antiamebic drugs. To this end, a tridimensional model of E. histolytica hexokinase 1 (EhHK1) was constructed and validated by homology modeling. After virtual screening of 14?400 small molecules, the 100 with the best docking scores were selected, purchased and assessed in their inhibitory capacity. The results showed that three molecules (compounds 2921, 11275 and 2755) inhibited EhHK1 with an I₅₀ of 48, 91 and 96?µM, respectively. Thus, we found the first inhibitors of EhHK1 that can be used in the search of new chemotherapeutic agents against amebiasis.     

Discovery of novel PRL-3 inhibitors based on the structure-based virtual screening

The inhibitors of phosphatase of regenerating liver-3 (PRL-3) have been shown to be useful as therapeutics for the treatment of cancer. We have been able to identify 12 novel PRL-3 inhibitors by means of the virtual screening with docking simulations under the consideration of the effects of ligand solvation in the scoring function. Because the newly identified inhibitors are structurally diverse and reveal a significant potency with IC(50) values ranging from 10 to 50muM, all of them can be considered for further development by structure-activity relationship or de novo design methods. Structural features relevant to the interactions of the newly identified inhibitors with the amino acid residues in the active site and the peripheral binding site of PRL-3 are discussed in detail.     

Structure-based virtual screening of Src kinase inhibitors

Src is an important target in multiple processes associated with tumor growth and development, including proliferation, neovascularization, and metastasis. In this study, hit identification was performed by virtual screening of commercial and in-house compound libraries. Docking studies for the hits were performed, and scoring functions were used to evaluate the docking results and to rank ligand-binding affinities. Subsequently, hit optimization for potent and selective candidate Src inhibitors was performed through focused library design and docking analyses. Consequently, we report that a novel compound ‘43’ with an IC₅₀ value of 89 nM, representing (S)-N-(4-(5-chlorobenzo[d][1,3]dioxol-4-ylamino)-7-(2-methoxyethoxy)quinazolin-6-yl)pyrrolidine-2-carboxamide, is highly selective for Src in comparison to EGFR (IC₅₀ ratio > 80-fold) and VEGFR-2 (IC₅₀ ratio > 110-fold). Compound 43 exerted anti-proliferative effects on Src-expressing PC3 human prostate cancer and A431 human epidermoid carcinoma cells, with calculated IC₅₀ values of 1.52 and 0.78 μM, respectively. Moreover, compound 43 (0.1 μM) suppressed the phosphorylation of extracellular signal-regulated kinases and p90 ribosomal S6 kinase, downstream molecules of Src, in a time-dependent manner, in both PC3 and A431 cell lines. The docking structure of compound 43 with Src disclosed that the chlorobenzodioxole moiety and pyrrolidine ring of C-6 quinazoline appeared to fit tightly into the hydrophobic pocket of Src. Additionally, the pyrrolidine NH forms a hydrogen bond with the carboxyl group of Asp348. These results confirm the successful application of virtual screening studies in the lead discovery process, and suggest that our novel compound 43 can be an effective Src inhibitor candidate for further lead optimization.     

非受体酪氨酸激酶ARG与过氧化氢酶相互作用机制的初步研究

通过体外结合实验,发现Arg蛋白与过氧化氢酶之间有相互作用,且这种作用是通过Arg蛋白的SH3结构域和过氧化氢酶的P290FNP介导的。利用体外激酶分析和Western印迹证明Arg蛋白可以使过氧化氢酶磷酸化。本研究为过氧化氢酶的活性调控研究奠定了基础。     

Discovery of indenopyrazoles as EGFR and VEGFR-2 tyrosine kinase inhibitors by in silico high-throughput screening

A series of indenopyrazoles 8 and 9 were designed and synthesized as EGFR tyrosine kinase inhibitors by in silico high-throughput screening. Compounds 8b and 8d showed significant inhibition of A431 cell growth (GI50 = 0.062 and 0.057 microM, respectively). Compounds 8b and 9a showed inhibitory activity toward both EGFR and VEGFR-2 (KDR) tyrosine kinases, whereas 8d inhibited VEGFR-2 tyrosine kinase, exclusively.     

Discovery of novel small-molecule Src kinase inhibitors via a kinase-focused druglikeness rule and structure-based virtual screening

Overexpression of the non-receptor tyrosine kinase Src is implicated in the development and progression of various human cancers. Blocking signalling pathways mediated by Src is believed to be a promising anticancer strategy. We report herein the discovery of novel small-molecule Src inhibitors by crystal structure-based virtual screening. A kinase-focused druglikeness rule was proposed and used in the design of compound library. Combination of large-scale docking with DOCK and rescoring with GOLD resulted in 6 hits with moderate to potent inhibitory activity against Src. Among them, compound 1 with an IC₅₀ of 1.2 μM shows the most potent inhibitory activity. By using molecular docking, binding models of the top 3 hits (ranked by potency and ligand efficiency) with Src were constructed to provide a rational strategy that simultaneously exploits hydrogen bonding interaction in the hinge region and hydrophobic stacking in the back pocket. This approach is instructive and meaningful to further structure-based drug design of Src inhibitors.     

Identification of potential glycogen kinase-3 inhibitors by structure based virtual screening

Glycogen synthase kinase-3 (GSK3) is a serine/threonine kinase that has attracted much drug discovery attention in recent years. Structural crystallography of the kinase has produced several high resolution inhibitor-GSK3 complexes and this is offering valuable information about the important pharmacophoric features present in the inhibitor, the protein target and the bioactive conformation. The availability of several GSK3-inhibitor co-crystals was successfully exploited to derive a pharmacophore query which retains the all important inhibitor-GSK3 interaction chemistry. A hypothesis containing three features: two hydrogen bond donors and one hydrogen acceptor was found to explain much of the inhibitor-GSK3 interaction. Subsequently, the query has been submitted to three databases for electronic screening. The hits obtained were docked into glycogen synthase kinase-3beta active site. A total of 21 novel potential leads were proposed after thorough examination by a combination of methods: (i) visual examination of how well they dock into the glycogen synthase kinase-3beta binding site, (ii) detailed analysis of their FlexX, G_Score, PMF_Score, ChemScore and D_Score values, (iii) comparative investigation of the docking scores of the hits with that of the thus far reported inhibitors (iv) determination of the binding mode and examination of how the hits retain interactions with the important amino acid residues of the kinase binding site. The hydrophobic heterocycles identified in this investigation are expected to be important additions to the armamentarium of GSK3 hyperactivity antagonism. Further more, the present work may further our current knowledge of the molecular basis of activation, inhibition and regulation of this pharmaceutically important kinase.     

Discovery of novel HCV polymerase inhibitors using pharmacophore-based virtual screening

We report the use of pharmacophore-based virtual screening as an efficient tool for the discovery of novel HCV polymerase inhibitors. A three-dimensional pharmacophore model for the HCV-796 binding site, NNI site IV inhibitor, to the enzyme was built by means of the structure-based focusing module in Cerius2 program. Using these models as a query for virtual screening, we produced a successful example of using pharmacophore-based virtual screening to identify novel compounds with HCV replicon assay through inhibition of HCV polymerization. Among the hit compounds, compounds 1 and 2 showed 56% and 48% inhibition of NS5B polymerization activity at 20 μM, respectively. In addition, compound 1 also exhibited replicon activity with EC₅₀ value of 2.16 μM. Following up the initial hit, we obtained derivatives of compound 1 and evaluated polymerization inhibition activity and HCV replicon assay. These results provide information necessary for the development of more potent NS5B inhibitors.     

Molecular docking of 1H-pyrazole derivatives to receptor tyrosine kinase and protein kinase for screening potential inhibitors

Tyrosine kinase receptor and protein kinases drawn much attention for the scientific fraternity in drug discovery due to its important role in different cancer, cardiovascular diseases and other hyper-proliferative disorders. Docking studies of pyrazole derivatives with tyrosine kinase and different serine/threonine protein kinases were employed by using flexible ligand docking approach of AutoDock 4.2. Among the molecules tested for docking study, 2-(4-chlorophenyl)-5-(3-(4-chlorophenyl)-5-methyl-1- phenyl-1H-pyrazol-4-yl)-1,3,4-thiadiazole (1b), 2-(4-methoxyphenyl)-5-(3-(4-methoxyphenyl)-5-methyl-1-phenyl-1H-pyrazol-4-yl)- 1,3,4-thiadiazole (1d) and 2-(4-chlorophenyl)-5-(3-(4-chlorophenyl)-5-methyl-1-phenyl-1H-pyrazol-4-yl)-1,3,4-thiadiazole (2b) revealed minimum binding energy of -10.09, -8.57 and -10.35 kJ/mol with VEGFR-2 (2QU5), Aurora A (2W1G) and CDK2 (2VTO) protein targets, respectively. These proteins are representatives of plausible models of interactions with different anticancer agents. All the ligands were docked deeply within the binding pocket region of all the three proteins, showing reasonable hydrogen bonds. The docking study results showed that these pyrazole derivatives are potential inhibitor of all the three protein targets; and also all these docked compounds have good inhibition constant, vdW + Hbond + desolv energy with best RMSD value.     

Discovery of the first SecA inhibitors using structure-based virtual screening

Bacterial protein secretion is a critical and complex process. The Sec machinery provides a major pathway for protein translocation across and integration into the cellular membrane in bacteria. Small molecule probes that perturb the functions of individual member proteins within the Sec machinery will be very important research tools as well as leads for future antimicrobial agent development. Herein we describe the discovery of inhibitors, through virtual screening, that specifically act on SecA ATPase, which is a critical member of the Sec system. These are the very first inhibitors reported for intrinsic SecA ATPase.