Liansheng Qiao

Virtual Screening and Molecular Dynamics Study of Potential Negative Allosteric Modulators of mGluR1 from Chinese Herbs

The metabotropic glutamate subtype 1 (mGluR1), a member of the metabotropic glutamate receptors, is a therapeutic target for neurological disorders. However, due to the lower subtype selectivity of mGluR1 orthosteric compounds, a new targeted strategy, known as allosteric modulators research, is needed for the treatment of mGluR1-related diseases. Recently, the structure of the seven-transmembrane domain (7TMD) of mGluR1 has been solved, which reveals the binding site of allosteric modulators and provides an opportunity for future subtype-selectivity drug design. In this study, a series of computer-aided drug design methods were utilized to discover potential mGluR1 negative allosteric modulators (NAMs). Pharmacophore models were constructed based on three different structure types of mGluR1 NAMs. After validation using the built-in parameters and test set, the optimal pharmacophore model of each structure type was selected and utilized as a query to screen the Traditional Chinese Medicine Database (TCMD). Then, three different hit lists of compounds were obtained. Molecular docking was used based on the latest crystal structure of mGluR1-7TMD to further filter these hits. As a compound with high QFIT and LibDock Score was preferred, a total of 30 compounds were retained. MD simulation was utilized to confirm the stability of potential compounds binding. From the computational results, thesinine-4ʹ-O-β-d-glucoside, nigrolineaxanthone-P and nodakenin might exhibit negative allosteric moderating effects on mGluR1. This paper indicates the applicability of molecular simulation technologies for discovering potential natural mGluR1 NAMs from Chinese herbs.

Virtual Screening for Potential Allosteric Inhibitors of Cyclin-Dependent Kinase 2 from Traditional Chinese Medicine

Cyclin-dependent kinase 2 (CDK2), a member of Cyclin-dependent kinases (CDKs), plays an important role in cell division and DNA replication. It is regarded as a desired target to treat cancer and tumor by interrupting aberrant cell proliferation. Compared to lower subtype selectivity of CDK2 ATP-competitive inhibitors, CDK2 allosteric inhibitor with higher subtype selectivity has been used to treat CDK2-related diseases. Recently, the first crystal structure of CDK2 with allosteric inhibitor has been reported, which provides new opportunities to design pure allosteric inhibitors of CDK2. The binding site of the ATP-competition inhibitors and the allosteric inhibitors are partially overlapped in space position, so the same compound might interact with the two binding sites. Thus a novel screening strategy was essential for the discovery of pure CDK2 allosteric inhibitors. In this study, pharmacophore and molecular docking were used to screen potential CDK2 allosteric inhibitors and ATP-competition inhibitors from Traditional Chinese Medicine (TCM). In the docking result of the allosteric site, the compounds which can act with the CDK2 ATP site were discarded, and the remaining compounds were regarded as the potential pure allosteric inhibitors. Among the results, prostaglandin E1 and nordihydroguaiaretic acid (NDGA) were available and their growth inhibitory effect on human HepG2 cell lines was determined by MTT assay. The two compounds could substantially inhibit the growth of HepG2 cell lines with an estimated IC50 of 41.223 μmol/L and 45.646 μmol/L. This study provides virtual screening strategy of allosteric compounds and a reliable method to discover potential pure CDK2 allosteric inhibitors from TCM. Prostaglandin E1 and NDGA could be regarded as promising candidates for CDK2 allosteric inhibitors.

A Novel Antihypertensive Derived from Adlay (Coix larchryma-jobi L. var. ma-yuen Stapf) Glutelin

Our previous studies have shown that Coix glutelin pepsin hydrolysate can effectively inhibit angiotensin converting enzyme (ACE) activity in vitro. The main purpose of this study was to obtain potent anti-hypertensive peptides from Coix glutelin. The Coix glutelin hydrolysates (CGH) were prepared by pepsin catalysis and further separated by an ultrafitration (UF) system, gel filtration chromatography (GFC) and reversed-phase high performance liquid chromatography (RP-HPLC). As a result, the sub-fraction F5-3 had the highest ACE-inhibitory activity. Six ACE inhibitory peptides were identified using nano-liquid chromatography coupled to tandem mass spectrometry. The most potent peptide GAAGGAF (IC50 = 14.19 μmol·L−1) was finally obtained by further molecular simulation screening and a series of division and optimization. Single oral administration of synthesized GAAGGAF at 15 mg/kg body weight (BW) in spontaneously hypertensive rats (SHR) could reduce the systolic blood pressure (SBP) around 27.50 mmHg and the effect lasted for at least 8 h. The study demonstrated for the first time that the ACE inhibitory peptide GAAGGAF from Coix glutelin has a significant antihypertensive effect, and it could be a good natural ingredient for pharmaceuticals against hypertension and the related diseases.

Discovery of Anti-Hypertensive Oligopeptides from Adlay Based on In Silico Proteolysis and Virtual Screening

Adlay (Coix larchryma-jobi L.) was the commonly used Traditional Chinese Medicine (TCM) with high content of seed storage protein. The hydrolyzed bioactive oligopeptides of adlay have been proven to be anti-hypertensive effective components. However, the structures and anti-hypertensive mechanism of bioactive oligopeptides from adlay were not clear. To discover the definite anti-hypertensive oligopeptides from adlay, in silico proteolysis and virtual screening were implemented to obtain potential oligopeptides, which were further identified by biochemistry assay and molecular dynamics simulation. In this paper, ten sequences of adlay prolamins were collected and in silico hydrolyzed to construct the oligopeptide library with 134 oligopeptides. This library was reverse screened by anti-hypertensive pharmacophore database, which was constructed by our research team and contained ten anti-hypertensive targets. Angiotensin-I converting enzyme (ACE) was identified as the main potential target for the anti-hypertensive activity of adlay oligopeptides. Three crystal structures of ACE were utilized for docking studies and 19 oligopeptides were finally identified with potential ACE inhibitory activity. According to mapping features and evaluation indexes of pharmacophore and docking, three oligopeptides were selected for biochemistry assay. An oligopeptide sequence, NPATY (IC50 = 61.88 ± 2.77 µM), was identified as the ACE inhibitor by reverse-phase high performance liquid chromatography (RP-HPLC) assay. Molecular dynamics simulation of NPATY was further utilized to analyze interactive bonds and key residues. ALA354 was identified as a key residue of ACE inhibitors. Hydrophobic effect of VAL518 and electrostatic effects of HIS383, HIS387, HIS513 and Zn2+ were also regarded as playing a key role in inhibiting ACE activities. This study provides a research strategy to explore the pharmacological mechanism of Traditional Chinese Medicine (TCM) proteins based on in silico proteolysis and virtual screening, which could be beneficial to reveal the pharmacological action of TCM proteins and provide new lead compounds for peptides-based drug design.

Discovery of Potential Orthosteric and Allosteric Antagonists of P2Y1R from Chinese Herbs by Molecular Simulation Methods

P2Y1 receptor (P2Y1R), which belongs to G protein-coupled receptors (GPCRs), is an important target in ADP-induced platelet aggregation. The crystal structure of P2Y1R has been solved recently, which revealed orthosteric and allosteric ligand-binding sites with the details of ligand-protein binding modes. And it suggests that P2Y1R antagonists, which recognize two distinct sites, could potentially provide an efficacious and safe antithrombotic profile. In present paper, 2D similarity search, pharmacophore based screening, and molecular docking were used to explore the potential natural P2Y1R antagonists. 2D similarity search was used to classify orthosteric and allosteric antagonists of P2Y1R. Based on the result, pharmacophore models were constructed and validated by the test set. Optimal models were selected to discover potential P2Y1R antagonists of orthosteric and allosteric sites from Traditional Chinese Medicine (TCM). And the hits were filtered by Lipinskis rule. Then molecular docking was used to refine the results of pharmacophore based screening and analyze the binding mode of the hits and P2Y1R. Finally, two orthosteric and one allosteric potential compounds were obtained, which might be used in future P2Y1R antagonists design. This work provides a reliable guide for discovering natural P2Y1R antagonists acting on two distinct sites from TCM.

Identification of potential ACAT-2 selective inhibitors using pharmacophore, SVM and SVR from Chinese herbs.

Acyl-coenzyme A cholesterol acyltransferase (ACAT) plays an important role in maintaining cellular and organismal cholesterol homeostasis. Two types of ACAT isozymes with different functions exist in mammals, named ACAT-1 and ACAT-2. Numerous studies showed that ACAT-2 selective inhibitors are effective for the treatment of hypercholesterolemia and atherosclerosis. However, as a typical endoplasmic reticulum protein, ACAT-2 protein has not been purified and revealed, so combinatorial ligand-based methods might be the optimal strategy for discovering the ACAT-2 selective inhibitors. In this study, selective pharmacophore models of ACAT-1 inhibitors and ACAT-2 inhibitors were built, respectively. The optimal pharmacophore model for each subtype was identified and utilized as queries for the Traditional Chinese Medicine Database screening. A total of 180 potential ACAT-2 selective inhibitors were obtained, which were identified using an ACAT-2 pharmacophore and not by our ACAT-1 model. Selective SVM model and bioactive SVR model were generated for further identification of the obtained ACAT-2 inhibitors. Ten compounds were finally obtained with predicted inhibitory activities toward ACAT-2. Hydrogen bond acceptor, 2D autocorrelations, GETAWAY descriptors, and BCUT descriptors were identified as key structural features for selectivity and activity of ACAT-2 inhibitors. This study provides a reasonable ligand-based approach to discover potential ACAT-2 selective inhibitors from Chinese herbs, which could help in further screening and development of ACAT-2 selective inhibitors.

Discovery of Potential Inhibitors of Aldosterone Synthase from Chinese Herbs Using Pharmacophore Modeling, Molecular Docking, and Molecular Dynamics Simulation Studies

Aldosterone synthase (CYP11B2) is a key enzyme for the biosynthesis of aldosterone, which plays a significant role for the regulation of blood pressure. Excess aldosterone can cause the dysregulation of the renin-angiotensin-aldosterone system (RAAS) and lead to hypertension. Therefore, research and development of CYP11B2 inhibitor are regarded as a novel approach for the treatment of hypertension. In this study, the pharmacophore models of CYP11B2 inhibitors were generated and the optimal model was used to identify potential CYP11B2 inhibitors from the Traditional Chinese Medicine Database (TCMD, Version 2009). The hits were further refined by molecular docking and the interactions between compounds and CYP11B2 were analyzed. Compounds with high Fitvalue, high docking score, and expected interactions with key residues were selected as potential CYP11B2 inhibitors. Two most promising compounds, ethyl caffeate and labiatenic acid, with high Fitvalue and docking score were reserved for molecular dynamics (MD) study. All of them have stability of ligand binding which suggested that they might perform the inhibitory effect on CYP11B2. This study provided candidates for novel drug-like CYP11B2 inhibitors by molecular simulation methods for the hypertension treatment.

Discovery of Potential Inhibitors of Squalene Synthase from Traditional Chinese Medicine Based on Virtual Screening and In Vitro Evaluation of Lipid-Lowering Effect

Squalene synthase (SQS), a key downstream enzyme involved in the cholesterol biosynthetic pathway, plays an important role in treating hyperlipidemia. Compared to statins, SQS inhibitors have shown a very significant lipid-lowering effect and do not cause myotoxicity. Thus, the paper aims to discover potential SQS inhibitors from Traditional Chinese Medicine (TCM) by the combination of molecular modeling methods and biological assays. In this study, cynarin was selected as a potential SQS inhibitor candidate compound based on its pharmacophoric properties, molecular docking studies and molecular dynamics (MD) simulations. Cynarin could form hydrophobic interactions with PHE54, LEU211, LEU183 and PRO292, which are regarded as important interactions for the SQS inhibitors. In addition, the lipid-lowering effect of cynarin was tested in sodium oleate-induced HepG2 cells by decreasing the lipidemic parameter triglyceride (TG) level by 22.50%. Finally. cynarin was reversely screened against other anti-hyperlipidemia targets which existed in HepG2 cells and cynarin was unable to map with the pharmacophore of these targets, which indicated that the lipid-lowering effects of cynarin might be due to the inhibition of SQS. This study discovered cynarin is a potential SQS inhibitor from TCM, which could be further clinically explored for the treatment of hyperlipidemia.

In Silico Analysis of the Association Relationship between Neuroprotection and Flavors of Traditional Chinese Medicine Based on the mGluRs

The metabotropic glutamate receptors (mGluRs) are known as both synaptic receptors and taste receptors. This feature is highly similar to the Property and Flavor theory of Traditional Chinese medicine (TCM), which has the pharmacological effect and flavor. In this study, six ligand based pharmacophore (LBP) models, seven homology modeling models, and fourteen molecular docking models of mGluRs were built based on orthosteric and allosteric sites to screening potential compounds from Traditional Chinese Medicine Database (TCMD). Based on the Pharmacopoeia of the Peoples Republic of China, TCMs of compounds and their flavors were traced and listed. According to the tracing result, we found that the TCMs of the compounds which bound to orthosteric sites of mGluRs are highly correlated to a sweet flavor, while the allosteric site corresponds to a bitter flavor. Meanwhile, the pharmacological effects of TCMs with highly frequent flavors were further analyzed. We found that those TCMs play a neuroprotective role through the efficiencies of detumescence, promoting blood circulation, analgesic effect, and so on. This study provides a guide for developing new neuroprotective drugs from TCMs which target mGluRs. Moreover, it is the first study to present a novel approach to discuss the association relationship between flavor and the neuroprotective mechanism of TCM based on mGluRs.

A Component Formula of Chinese Medicine for Hypercholesterolemia Based on Virtual Screening and Biology Network

Hypercholesterolemia is a risk factor to atherosclerosis and coronary heart disease II. The abnormal rise of cholesterol in plasma is the main symptom. Cholesterol synthesis pathway is an important pathway of the origin of cholesterol, which is an essential pathway for the therapy of hypercholesterolemia. The 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMG-CoA reductase), squalene synthase (SQS), and sterol regulatory element binding protein-2 (SREBP-2) are closely connected with the synthesis of cholesterol. The inhibition of these targets can reduce the cholesterol in plasma. This study aimed to build a component formula including three Traditional Chinese Medicines (TCM) components with the inhibition activity of these targets by using virtual screening and biological network. Structure-based pharmacophore models of HMG-CoA reductase and SQS and ligand-based pharmacophore model of SREBP-2 were constructed to screen the Traditional Chinese Medicine Database (TCMD). Molecular docking was used for further screening of components of HMG-CoA reductase and SQS. Then, metabolic network was constructed to elucidate the comprehensive interaction of three targets for lipid metabolism. Finally, three potential active compounds were obtained, which are poncimarin, hexahydrocurcumin, and forsythoside C. The source plants of the compounds were also taken into account, which should have known action of lowering hyperlipidemia. The lipid-lowering effect of hexahydrocurcumin was verified by experiment in vitro. The components that originated from TCMs with lipid-lowering efficacy made up a formula with a synergistic effect through the computer aid drug design methods. The research provides a fast and efficient method to build TCM component formula and it may inspire the study of the explanation of TCM formula mechanism.