新型乙酰胆碱酯酶抑制剂的分子对接研究

构建了乙酰胆碱酯酶抑制剂药效团模型,并基于此模型进行新型乙酰胆碱酯酶抑制剂的分子设计。用MVD软件进行分子对接,预测设计分子与乙酰胆碱酯酶的结合活性。通过计算发现,基于此药效团设计的小分子9与14乙酰胆碱酯酶具有较好的结合活性。     

Combined Pharmacophore Modeling,Docking, and 3D QSAR Studies of ABCB1 and ABCC1 Transporter Inhibitors

Quinazolinones, indolo‐ and pyrrolopyrimidines with inhibitory effects toward ABCB1 (P‐gp) and ABCC1 (MRP1) transporters were studied by pharmacophore modeling, docking, and 3D QSAR to describe the binding preferences of the proteins. The pharmacophore overlays between dual and/or highly selective inhibitors point to binding sites of different topology and physiochemical properties for MRP1 and P‐gp. Docking of selective inhibitors into the P‐gp binding cavity by the use of a structural model based on the recently resolved P‐gp structure confirms the P‐gp pharmacophore features identified, and reveals the interactions of some functional groups and atoms in the structures with particular protein residues. The 3D QSAR analysis of the dual‐effect inhibitors allows satisfactory prediction of the selectivity index of the compounds and outlines electrostatics as most important for selectivity. The results from the combined modeling approach complement each other and could improve our understanding of the protein–ligand interactions involved, and could aid in the development of highly selective and potent inhibitors of P‐gp and MRP1.     

Carborane-Containing Matrix Metalloprotease (MMP) Ligands as Candidates for Boron Neutron-Capture Therapy (BNCT)

Based on the previously reported potent and selective sulfone hydroxamate inhibitors SC-76276, SC-78080 (SD-2590), and SC-77964, potent MMP inhibitors have been designed and synthesized to append a boron-rich carborane cluster by employing click chemistry to target tumor cells that are known to upregulate gelatinases. Docking against MMP-2 suggests binding involving the hydroxamate zinc-binding group, key H-bonds by the sulfone moiety with the peptide backbone residues Leu82 and Leu83, and a hydrophobic interaction with the deep P1’ pocket. The more potent of the two triazole regioisomers exhibits an IC₅₀ of 3.7 nM versus MMP-2 and IC₅₀ of 46 nM versus MMP-9.     

Probing small-molecule binding to cytochrome P450 2D6 and 2C9: An in silico protocol for generating toxicity alerts

Drug metabolism, toxicity, and their interaction profiles are major issues in the drug-discovery and lead-optimization processes. The cytochromes P450 (CYPs) 2D6 and 2C9 are enzymes involved in the oxidative metabolism of a majority of marketed drugs. Therefore, the prediction of the binding affinity towards CYP2D6 and CYP2C9 would be beneficial for identifying cytochrome-mediated adverse effects triggered by drugs or chemicals (e.g., toxic reactions, drug-drug, and food-drug interactions). By identifying the binding mode by using pharmacophore prealignment, automated flexible docking, and by quantifying the binding affinity by multidimensional QSAR (mQSAR), we validated a model family of 56 compounds (46 training, 10 test) and 85 compounds (68 training, 17 test) for CYP2D6 and CYP2C9, respectively. The correlation with the experimental data (cross-validated r2=0.811 for CYP2D6 and 0.687 for CYP2C9) suggests that our approach is suited for predicting the binding affinity of compounds towards CYP2D6 and CYP2C9. The models were challenged by Y-scrambling and by testing an external dataset of binding compounds (15 compounds for CYP2D6 and 40 for CYP2C9). To assess the probability of false-positive predictions, datasets of nonbinders (64 compounds for CYP2D6 and 56 for CYP2C9) were tested by using the same protocol. The two validated mQSAR models were subsequently added to the VirtualToxLab (VTL, http://www.virtualtoxlab.org).     

Biphenyl sulfonylamino methyl bisphosphonic acids as inhibitors of matrix metalloproteinases and bone resorption

A number of matrix metalloproteinases (MMPs), proteins important in the balance of bone remodeling, play a critical role both in cancer metastasis and in bone matrix turnover associated with the presence of cancer cells in bone. Here, we report the synthesis and biological evaluation of a new class of MMP inhibitors characterized by a bisphosphonate function as the zinc binding group. Since the bisphosphonate group is also implicated in osteoclast inhibition and provides a preferential affinity to biological apatite, the new molecules can be regarded as bone-seeking medicinal agents. Docking experiments were performed to clarify the mode of binding of bisphosphonate inhibitors in the active site of MMP-2. The most promising of the studied bisphosphonates showed nanomolar inhibition against MMP-2 and resulted in potent inhibition of osteoclastic bone resorption in vitro.     

Molecular Recognition of the Catalytic Zinc(II) Ion in MMP-13: Structure-Based Evolution of an Allosteric Inhibitor to Dual Binding Mode Inhibitors with Improved Lipophilic Ligand Efficiencies

Matrix metalloproteinases (MMPs) are a class of zinc dependent endopeptidases which play a crucial role in a multitude of severe diseases such as cancer and osteoarthritis. We employed MMP-13 as the target enzyme for the structure-based design and synthesis of inhibitors able to recognize the catalytic zinc ion in addition to an allosteric binding site in order to increase the affinity of the ligand. Guided by molecular modeling, we optimized an initial allosteric inhibitor by addition of linker fragments and weak zinc binders for recognition of the catalytic center. Furthermore we improved the lipophilic ligand efficiency (LLE) of the initial inhibitor by adding appropriate zinc binding fragments to lower the clogP values of the inhibitors, while maintaining their potency. All synthesized inhibitors showed elevated affinity compared to the initial hit, also most of the novel inhibitors displayed better LLE. Derivatives with carboxylic acids as the zinc binding fragments turned out to be the most potent inhibitors (compound 3 (ZHAWOC5077): IC₅₀ = 134 nM) whereas acyl sulfonamides showed the best lipophilic ligand efficiencies (compound 18 (ZHAWOC5135): LLE = 2.91).     

Computational Investigation Identified Potential Chemical Scaffolds for Heparanase as Anticancer Therapeutics

Heparanase (Hpse) is an endo-β-D-glucuronidase capable of cleaving heparan sulfate side chains. Its upregulated expression is implicated in tumor growth, metastasis and angiogenesis, thus making it an attractive target in cancer therapeutics. Currently, a few small molecule inhibitors have been reported to inhibit Hpse, with promising oral administration and pharmacokinetic (PK) properties. In the present study, a ligand-based pharmacophore model was generated from a dataset of well-known active small molecule Hpse inhibitors which were observed to display favorable PK properties. The compounds from the InterBioScreen database of natural (69,034) and synthetic (195,469) molecules were first filtered for their drug-likeness and the pharmacophore model was used to screen the drug-like database. The compounds acquired from screening were subjected to molecular docking with Heparanase, where two molecules used in pharmacophore generation were used as reference. From the docking analysis, 33 compounds displayed higher docking scores than the reference and favorable interactions with the catalytic residues. Complex interactions were further evaluated by molecular dynamics simulations to assess their stability over a period of 50 ns. Furthermore, the binding free energies of the 33 compounds revealed 2 natural and 2 synthetic compounds, with better binding affinities than reference molecules, and were, therefore, deemed as hits. The hit compounds presented from this in silico investigation could act as potent Heparanase inhibitors and further serve as lead scaffolds to develop compounds targeting Heparanase upregulation in cancer.     

Three-dimensional pharmacophore design and biochemical screening identifies substituted 1,2,4-triazoles as inhibitors of the annexin A2-S100A10 protein interaction

Protein interactions are increasingly appreciated as targets in small‐molecule drug discovery. The interaction between the adapter protein S100A10 and its binding partner annexin A2 is a potentially important drug target. To obtain small‐molecule starting points for inhibitors of this interaction, a three‐dimensional pharmacophore model was constructed from the X‐ray crystal structure of the complex between S100A10 and annexin A2. The pharmacophore model represents the favourable hydrophobic and hydrogen bond interactions between the two partners, as well as spatial and receptor site constraints (excluded volume spheres). Using this pharmacophore model, UNITY flex searches were carried out on a 3D library of 0.7 million commercially available compounds. This resulted in 568 hit compounds. Subsequently, GOLD docking studies were performed on these hits, and a set of 190 compounds were purchased and tested biochemically for inhibition of the protein interaction. Three compounds of similar chemical structure were identified as genuine inhibitors of the binding of annexin A2 to S100A10. The binding modes predicted by GOLD were in good agreement with their UNITY‐generated conformations. We synthesised a series of analogues revealing areas critical for binding. Thus computational predictions and biochemical screening can be used successfully to derive novel chemical classes of protein–protein interaction blockers.     

Halting Tumor Progression via Novel Non-Hydroxamate Triazole-Based Mannich Bases MMP-2/9 Inhibitors; Design,Microwave-Assisted Synthesis,and Biological Evaluation

Matrix metalloproteinases (MMPs) are key signaling modulators in the tumor microenvironment. Among MMPs, MMP-2 and MMP-9 are receiving renewed interest as validated druggable targets for halting different tumor progression events. Over the last decades, a diverse range of MMP-2/9 inhibitors has been identified starting from the early hydroxamic acid-based peptidomimetics to the next generation non-hydroxamates. Herein, focused 1,2,4-triazole-1,2,3-triazole molecular hybrids with varying lengths and decorations, mimicking the thematic features of non-hydroxamate inhibitors, were designed and synthesized using efficient protocols and were alkylated with pharmacophoric amines to develop new Mannich bases. After full spectroscopic characterization the newly synthesized triazoles tethering Mannich bases were subjected to safety assessment via MTT assay against normal human fibroblasts, then evaluated for their potential anticancer activities against colon (Caco-2) and breast (MDA-MB 231) cancers. The relatively lengthy bis-Mannich bases 15 and 16 were safer and more potent than 5-fluorouracil with sub-micromolar IC₅₀ and promising selectivity to the screened cancer cell lines rather than normal cells. Both compounds upregulated p53 (2–5.6-fold) and suppressed cyclin D expression (0.8–0.2-fold) in the studied cancers, and thus, induced apoptosis. 15 was superior to 16 in terms of cytotoxic activities, p53 induction, and cyclin D suppression. Mechanistically, both were efficient MMP-2/9 inhibitors with comparable potencies to the reference prototype hydroxamate-based MMP inhibitor NNGH at their anticancer IC₅₀ concentrations. 15 (IC₅₀ = 0.143 µM) was 4-fold more potent than NNGH against MMP-9 with promising selectivity (3.27-fold) over MMP-2, whereas 16 was comparable to NNGH. Concerning MMP-2, 16 (IC₅₀ = 0.376 µM) was 1.2-fold more active than 15. Docking simulations predicted their possible binding modes and highlighted the possible structural determinants of MMP-2/9 inhibitory activities. Computational prediction of their physicochemical properties, ADMET, and drug-likeness metrics revealed acceptable drug-like criteria.     

Discovery of Tyrosine Kinase Inhibitors by Docking into an Inactive Kinase Conformation Generated by Molecular Dynamics

Several small molecules that bind to the inactive DFG‐out conformation of tyrosine kinases (called type II inhibitors) have shown a good selectivity profile over other kinase targets. To obtain a set of DFG‐out structures, we performed an explicit solvent molecular dynamics (MD) simulation of the complex of the catalytic domain of a tyrosine kinase receptor, ephrin type‐A receptor 3 (EphA3), and a manually docked type II inhibitor. Automatic docking of four previously reported type II inhibitors was used to select a single snapshot from the MD trajectory for virtual screening. High‐throughput docking of a pharmacophore‐tailored library of 175 000 molecules resulted in about 4 million poses, which were further filtered by van der Waals efficiency and ranked according to a force‐field‐based energy function. Notably, around 20 % of the compounds with predicted binding energy smaller than ?10 kcal mol^?1 are known type II inhibitors. Moreover, a series of 5‐(piperazine‐1‐yl)isoquinoline derivatives was identified as a novel class of low‐micromolar inhibitors of EphA3 and unphosphorylated Abelson tyrosine kinase (Abl1). The in silico predicted binding mode of the new inhibitors suggested a similar affinity to the gatekeeper mutant T315I of Abl1, which was verified in vitro by using a competition binding assay. Additional evidence for the type II binding mode was obtained by two 300 ns MD simulations of the complex between N‐(3‐chloro‐4‐(difluoromethoxy)phenyl)‐2‐(4‐(8‐nitroisoquinolin‐5‐yl)piperazin‐1‐yl)acetamide and EphA3.     

Structural Investigation of the Binding of 5‐Substituted Swainsonine Analogues to Golgi α‐Mannosidase II

Golgi α‐mannosidase II (GMII) is a key enzyme in the N‐glycosylation pathway and is a potential target for cancer chemotherapy. The natural product swainsonine is a potent inhibitor of GMII. In this paper we characterize the binding of 5α‐substituted swainsonine analogues to the soluble catalytic domain of Drosophila GMII by X‐ray crystallography. These inhibitors enjoy an advantage over previously reported GMII inhibitors in that they did not significantly decrease the inhibitory potential of the swainsonine head‐group. The phenyl groups of these analogues occupy a portion of the binding site not previously seen to be populated with either substrate analogues or other inhibitors and they form novel hydrophobic interactions. They displace a well‐organized water cluster, but the presence of a C(10) carbonyl allows the reestablishment of important hydrogen bonds. Already approximately tenfold more active against the Golgi enzyme than the lysosomal enzyme, these inhibitors offer the potential of being extended into the N‐acetylglucosamine binding site of GMII for the creation of even more potent and selective GMII inhibitors.     

Computationally Empowered Workflow Identifies Novel Covalent Allosteric Binders for KRASG12C

Due to its frequent mutations in multiple lethal cancers, KRAS is one of the most-studied anticancer targets nowadays. Since the discovery of the druggable allosteric binding site containing a G12C mutation, KRAS^G12C has been the focus of attention in oncology research. We report here a computationally driven approach aimed at identifying novel and selective KRAS^G12C covalent inhibitors. The workflow involved initial enumeration of virtual molecules tailored for the KRAS allosteric binding site. Tools such as pharmacophore modeling, docking, and free-energy perturbations were deployed to prioritize the compounds with the best profiles. The synthesized naphthyridinone scaffold showed the ability to react with G12C and inhibit KRAS^G12C. Analogues were prepared to establish structure-activity relationships, while molecular dynamics simulations and crystallization of the inhibitor-KRAS^G12C complex highlighted an unprecedented binding mode.     

The Discovery of Potentially Selective Human Neuronal Nitric Oxide Synthase (nNOS) Inhibitors: A Combination of Pharmacophore Modelling,CoMFA, Virtual Screening and Molecular Docking Studies

Neuronal nitric oxide synthase (nNOS) plays an important role in neurotransmission and smooth muscle relaxation. Selective inhibition of nNOS over its other isozymes is highly desirable for the treatment of neurodegenerative diseases to avoid undesirable effects. In this study, we present a workflow for the identification and prioritization of compounds as potentially selective human nNOS inhibitors. Three-dimensional pharmacophore models were constructed based on a set of known nNOS inhibitors. The pharmacophore models were evaluated by Pareto surface and CoMFA (Comparative Molecular Field Analysis) analyses. The best pharmacophore model, which included 7 pharmacophore features, was used as a search query in the SPECS database (SPECS^®, Delft, The Netherlands). The hit compounds were further filtered by scoring and docking. Ten hits were identified as potential selective nNOS inhibitors.     

A Multistage In Silico Study of Natural Potential Inhibitors Targeting SARS-CoV-2 Main Protease

Among a group of 310 natural antiviral natural metabolites, our team identified three compounds as the most potent natural inhibitors against the SARS-CoV-2 main protease (PDB ID: 5R84), M^pro. The identified compounds are sattazolin and caprolactin A and B. A validated multistage in silico study was conducted using several techniques. First, the molecular structures of the selected metabolites were compared with that of GWS, the co-crystallized ligand of M^pro, in a structural similarity study. The aim of this study was to determine the thirty most similar metabolites (10%) that may bind to the M^pro similar to GWS. Then, molecular docking against M^pro and pharmacophore studies led to the choice of five metabolites that exhibited good binding modes against the M^pro and good fit values against the generated pharmacophore model. Among them, three metabolites were chosen according to ADMET studies. The most promising M^pro inhibitor was determined by toxicity and DFT studies to be caprolactin A (292). Finally, molecular dynamics (MD) simulation studies were performed for caprolactin A to confirm the obtained results and understand the thermodynamic characteristics of the binding. It is hoped that the accomplished results could represent a positive step in the battle against COVID-19 through further in vitro and in vivo studies on the selected compounds.     

Acetyl Coenzyme A Analogues as Rationally Designed Inhibitors of Citrate Synthase

In this study, we probed the inhibition of pig heart citrate synthase (E.C. 4.1.3.7) by synthesising seven analogues either designed to mimic the proposed enolate intermediate in this enzyme reaction or developed from historical inhibitors. The most potent inhibitor was fluorovinyl thioether 9 (K_i=4.3 μm ), in which a fluorine replaces the oxygen atom of the enolate. A comparison of the potency of 9 with that of its non-fluorinated vinyl thioether analogue 10 (K_i=68.3 μm ) revealed a clear “fluorine effect” favouring 9 by an order of magnitude. The dethia analogues of 9 and 10 proved to be poor inhibitors. A methyl sulfoxide analogue was a moderate inhibitor (K_i=11.1 μm ), thus suggesting hydrogen bonding interactions in the enolate site. Finally, E and Z propenoate thioether isomers were explored as conformationally constrained carboxylates, but these were not inhibitors. All compounds were prepared by the synthesis of the appropriate pantetheinyl diol and then assembly of the coenzyme A structure according to a three-enzyme biotransformation protocol. A quantum mechanical study, modelling both inhibitors 9 and 10 into the active site indicated short CF ⋅⋅⋅ H contacts of ≈2.0 Å, consistent with fluorine making two stabilising hydrogen bonds, and mimicking an enolate rather than an enol intermediate. Computation also indicated that binding of 9 to citrate synthase increases the basicity of a key aspartic acid carboxylate, which becomes protonated.     

3D QSAR pharmacophore modeling, in silico screening, and density functional theory (DFT) approaches for identification of human chymase inhibitors

Human chymase is a very important target for the treatment of cardiovascular diseases. Using a series of theoretical methods like pharmacophore modeling, database screening, molecular docking and Density Functional Theory (DFT) calculations, an investigation for identification of novel chymase inhibitors, and to specify the key factors crucial for the binding and interaction between chymase and inhibitors is performed. A highly correlating (r = 0.942) pharmacophore model (Hypo1) with two hydrogen bond acceptors, and three hydrophobic aromatic features is generated. After successfully validating "Hypo1", it is further applied in database screening. Hit compounds are subjected to various drug-like filtrations and molecular docking studies. Finally, three structurally diverse compounds with high GOLD fitness scores and interactions with key active site amino acids are identified as potent chymase hits. Moreover, DFT study is performed which confirms very clear trends between electronic properties and inhibitory activity (IC(50)) data thus successfully validating "Hypo1" by DFT method. Therefore, this research exertion can be helpful in the development of new potent hits for chymase. In addition, the combinational use of docking, orbital energies and molecular electrostatic potential analysis is also demonstrated as a good endeavor to gain an insight into the interaction between chymase and inhibitors.     

From On‐Target to Off‐Target Activity: Identification and Optimisation of Trypanosoma brucei GSK3 Inhibitors and Their Characterisation as Anti‐Trypanosoma brucei Drug Discovery Lead Molecules

Human African trypanosomiasis (HAT) is a life‐threatening disease with approximately 30 000–40 000 new cases each year. Trypanosoma brucei protein kinase GSK3 short (TbGSK3) is required for parasite growth and survival. Herein we report a screen of a focused kinase library against T. brucei GSK3. From this we identified a series of several highly ligand‐efficient TbGSK3 inhibitors. Following the hit validation process, we optimised a series of diaminothiazoles, identifying low‐nanomolar inhibitors of TbGSK3 that are potent in vitro inhibitors of T. brucei proliferation. We show that the TbGSK3 pharmacophore overlaps with that of one or more additional molecular targets.     

Inhibition of HSP90 with Pochoximes: SAR and Structure‐Based Insights

The pochoximes, based on the radicicol pharmacophore, are potent inhibitors of heat shock protein 90 (HSP90) that retain their activity in vivo. Herein we report an extended library that broadly explores the structure–activity relationship (SAR) of the pochoximes with four points of diversity. Several modifications were identified that afford improved cellular efficacy, new opportunities for conjugation, and further diversifications. Cocrystal structures of pochoximes A and B with HSP90 show that pochoximes bind to a different conformation of HSP90 than radicicol and provide a rationale for the enhanced affinity of the pochoximes relative to radicicol and the pochonins.     

Hybrid Pharmacophore- and Structure-Based Virtual Screening Pipeline to Identify Novel EGFR Inhibitors That Suppress Non-Small Cell Lung Cancer Cell Growth

Dysregulated epidermal growth factor receptor (EGFR) expression is frequently observed in non-small cell lung cancer (NSCLC) growth and metastasis. Despite recent successes in the development of tyrosine kinase inhibitors (TKIs), inevitable resistance to TKIs has led to urgent calls for novel EGFR inhibitors. Herein, we report a rational workflow used to identify novel EGFR-TKIs by combining hybrid ligand- and structure-based pharmacophore models. Three types of models were developed in this workflow, including 3D QSAR-, common feature-, and structure-based EGFR-TK domain-containing pharmacophores. A National Cancer Institute (NCI) compound dataset was adopted for multiple-stage pharmacophore-based virtual screening (PBVS) of various pharmacophore models. The six top-scoring compounds were identified through the PBVS pipeline coupled with molecular docking. Among these compounds, NSC609077 exerted a significant inhibitory effect on EGFR activity in gefitinib-resistant H1975 cells, as determined by an enzyme-linked immunosorbent assay (ELISA). Further investigations showed that NSC609077 inhibited the anchorage-dependent growth and migration of lung cancer cells. Furthermore, NSC609077 exerted a suppressive effect on the EGFR/PI3K/AKT pathway in H1975 cells. In conclusion, these findings suggest that hybrid virtual screening may accelerate the development of targeted drugs for lung cancer treatment.