ABSTRACT
Metalloenzymes are attractive research targets in fields of chemistry, biology, and medicine. Given that metalloenzymes can manifest conservation of metal-coordination and ligand binding modes, the excavation and expansion of metalloenzyme-specific knowledge is of interest in bridging metalloenzyme-related fields. Building on our previous metalloenzyme-ligand association database, MeLAD, we have expanded the scope of metalloenzyme-specific knowledge and services, by forming a versatile platform, termed the Metalloenzyme Data Bank and Analysis (MeDBA). The MeDBA provides: (i) manual curation of metalloenzymes into different categories, that this M-I, M-II and M-III; (ii) comprehensive information on metalloenzyme activities, expression profiles, family and disease links; (iii) structural information on metalloenzymes, in particular metal binding modes; (iv) metalloenzyme substrates and bioactive molecules acting on metalloenzymes; (v) excavated metal-binding pharmacophores and (vi) analysis tools for structure/metal active site comparison and metalloenzyme profiling. The MeDBA is freely available at https://medba.ddtmlab.org.
Subject(s)
Databases, Protein , Metalloproteins , Catalytic Domain , Ligands , Metalloproteins/metabolism , Metals , EnzymesABSTRACT
BACKGROUND: There is a lack of effective therapeutic strategies for amyotrophic lateral sclerosis (ALS); therefore, drug repurposing might provide a rapid approach to meet the urgent need for treatment. METHODS: To identify therapeutic targets associated with ALS, we conducted Mendelian randomization (MR) analysis and colocalization analysis using cis-eQTL of druggable gene and ALS GWAS data collections to determine annotated druggable gene targets that exhibited significant associations with ALS. By subsequent repurposing drug discovery coupled with inclusion criteria selection, we identified several drug candidates corresponding to their druggable gene targets that have been genetically validated. The pharmacological assays were then conducted to further assess the efficacy of genetics-supported repurposed drugs for potential ALS therapy in various cellular models. RESULTS: Through MR analysis, we identified potential ALS druggable genes in the blood, including TBK1 [OR 1.30, 95%CI (1.19, 1.42)], TNFSF12 [OR 1.36, 95%CI (1.19, 1.56)], GPX3 [OR 1.28, 95%CI (1.15, 1.43)], TNFSF13 [OR 0.45, 95%CI (0.32, 0.64)], and CD68 [OR 0.38, 95%CI (0.24, 0.58)]. Additionally, we identified potential ALS druggable genes in the brain, including RESP18 [OR 1.11, 95%CI (1.07, 1.16)], GPX3 [OR 0.57, 95%CI (0.48, 0.68)], GDF9 [OR 0.77, 95%CI (0.67, 0.88)], and PTPRN [OR 0.17, 95%CI (0.08, 0.34)]. Among them, TBK1, TNFSF12, RESP18, and GPX3 were confirmed in further colocalization analysis. We identified five drugs with repurposing opportunities targeting TBK1, TNFSF12, and GPX3, namely fostamatinib (R788), amlexanox (AMX), BIIB-023, RG-7212, and glutathione as potential repurposing drugs. R788 and AMX were prioritized due to their genetic supports, safety profiles, and cost-effectiveness evaluation. Further pharmacological analysis revealed that R788 and AMX mitigated neuroinflammation in ALS cell models characterized by overly active cGAS/STING signaling that was induced by MSA-2 or ALS-related toxic proteins (TDP-43 and SOD1), through the inhibition of TBK1 phosphorylation. CONCLUSIONS: Our MR analyses provided genetic evidence supporting TBK1, TNFSF12, RESP18, and GPX3 as druggable genes for ALS treatment. Among the drug candidates targeting the above genes with repurposing opportunities, FDA-approved drug-R788 and AMX served as effective TBK1 inhibitors. The subsequent pharmacological studies validated the potential of R788 and AMX for treating specific ALS subtypes through the inhibition of TBK1 phosphorylation.
Subject(s)
Aminopyridines , Amyotrophic Lateral Sclerosis , Humans , Amyotrophic Lateral Sclerosis/drug therapy , Amyotrophic Lateral Sclerosis/genetics , Drug Repositioning , Mendelian Randomization Analysis , Protein Serine-Threonine Kinases/geneticsABSTRACT
BACKGROUND: We found a carbapenem-resistant Escherichia coli without known carbapenemase-encoding genes and performed a study to identify the possible new carbapenemase. METHODS: The production of carbapenemase was examined using the modified carbapenem inactivation method. The strain was subjected to short- and long-read genome sequencing and the complete genome was obtained by hybrid assembly. The gene encoding a potential new OXA-type carbapenemase was cloned. The enzyme was purified and was then subjected to kinetic assays. Molecular docking analysis of the enzyme was performed using the MOE software suite. Mating experiments were attempted to obtain the plasmid carrying the corresponding gene. RESULTS: We identified and characterized a novel class D carbapenem-hydrolysing ß-lactamase, OXA-1041, in a carbapenem-resistant E. coli clinical strain. OXA-1041 had 89.77% (237/264) amino acid identity with OXA-427, a known carbapenemase. By cloning in an E. coli laboratory strain, blaOXA-1041 was found to reduce susceptibility to ertapenem by 16 times (MIC 0.25 versus 0.016 mg/L) and meropenem by four times (MIC 0.06 versus 0.016 mg/L) but did not significantly reduce susceptibility to imipenem and doripenem. Enzyme kinetic measurement of purified OXA-1041 showed that OXA-1041 could hydrolyse ertapenem and meropenem with a turnover number (kcat)/Michaelis constant (KM) of 8.57 and 3.63 mM-1s-1, respectively. The complete genome contained a single plasmid (223 341 bp, IncF, containing five replicons), which was self-transmissible. blaOXA-1041 was downstream of insertion sequence ISCR1 and there were three tandem copies of ISCR1-blaOXA-1041-creDΔ (encoding an envelope protein) on this plasmid. CONCLUSIONS: The above findings suggest OXA-1041 is a new plasmid-encoded carbapenemase with preferential activity against ertapenem.
Subject(s)
Carbapenems , Escherichia coli , Carbapenems/pharmacology , Carbapenems/metabolism , Escherichia coli/genetics , Escherichia coli/metabolism , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/metabolism , Meropenem , Ertapenem/pharmacology , Molecular Docking Simulation , beta-Lactamases/metabolism , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Microbial Sensitivity TestsABSTRACT
The organocatalytic asymmetric Morita-Baylis-Hillman (MBH) reaction of isatin derivatives with various vinyl sulfones is disclosed. Chiral sulfone-containing 3-hydroxyoxindoles were produced in good to high yields and with good to high ee's. This report displays an unprecedented example to apply activated alkenes with sulfone moiety other than carbonyl groups in asymmetric MBH reactions and provides an efficient strategy to incorporate the sulfone functional group for the synthesis of chiral 3-hydroxyoxindoles.
ABSTRACT
SIRT5 has been implicated in various physiological processes and human diseases, including cancer. Development of new highly potent, selective SIRT5 inhibitors is still needed to investigate disease-related mechanisms and therapeutic potentials. We here report new ε-N-thioglutaryllysine derivatives, which were designed according to SIRT5-catalysed deacylation reactions. These ε-N-thioglutaryllysine derivatives displayed potent SIRT5 inhibition, of which the potential photo-crosslinking derivative 8 manifested most potent inhibition with an IC50 value of 120 nM to SIRT5, and low inhibition to SIRT1-3 and SIRT6. The enzyme kinetic assays revealed that the ε-N-thioglutaryllysine derivatives inhibit SIRT5 by lysine-substrate competitive manner. Co-crystallographic analyses demonstrated that 8 binds to occupy the lysine-substate binding site by making hydrogen-bonding and electrostatic interactions with SIRT5-specific residues, and is likely positioned to react with NAD+ and form stable thio-intermediates. Compound 8 was observed to have low photo-crosslinking probability to SIRT5, possibly due to inappropriate position of the diazirine group as observed in SIRT5:8 crystal structure. This study provides useful information for developing drug-like inhibitors and cross-linking chemical probes for SIRT5-related studies.
Subject(s)
Sirtuins , Humans , Sirtuins/metabolism , Lysine/chemistry , Binding SitesABSTRACT
Sorafenib is a clinically useful multiple kinase inhibitor for the treatment of kidney cancer, liver cancer and acute myelocytic leukemia, while it has shown weak efficacy in suppressing breast cancer. Since sirtuin2 (SIRT2) is an important epigenetic regulator and associated with several cancer types including breast cancer, development and evaluation of new SIRT2 inhibitors to probe their therapeutic potentials is currently desirable. A highly selective SIRT2 inhibitor named I was previously developed by us, which showed activity to inhibit non-small cell lung cancer cell lines in vitro. We herein report expanded screening of I and its structurally similar inactive compound II against other cancer cell lines, and found that I had a wide spectrum of anticancer activity while II had no such effects. The I-sorafenib combination treatment exerted obvious synergistic reduction on cell viability of MCF-7 cells. We observed that the combination treatment could suppress cell proliferation, survival and migration, arrest cell cycle at G0/G1 phase, and induce apoptosis in MCF-7 cells, when compared with the single treatment. In vivo studies revealed that the combination treatment showed stronger tumor growth inhibition (87%), comparing with I-(42.8%) or sorafenib-solely-treated groups (61.1%) in MCF-7 xenograft model. In conclusion, this work clearly revealed a potential synthetic lethality effect for I combined with sorafenib, and will probably offer a new strategy at least for breast cancer treatment.
Subject(s)
Antineoplastic Agents , Breast Neoplasms , Carcinoma, Non-Small-Cell Lung , Lung Neoplasms , Antineoplastic Agents/pharmacology , Antineoplastic Agents/therapeutic use , Apoptosis , Breast Neoplasms/drug therapy , Breast Neoplasms/pathology , Carcinoma, Non-Small-Cell Lung/drug therapy , Cell Line, Tumor , Cell Proliferation , Female , Humans , Lung Neoplasms/drug therapy , Niacinamide/pharmacology , Niacinamide/therapeutic use , Phenylurea Compounds/pharmacology , Phenylurea Compounds/therapeutic use , Protein Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/therapeutic use , Sirtuin 2 , Sorafenib/pharmacology , Sorafenib/therapeutic use , Synthetic Lethal Mutations , Xenograft Model Antitumor AssaysABSTRACT
Since metalloenzymes are a large collection of metal ion(s) dependent enzymes, comparison analyses of metalloenzyme active sites are critical for metalloenzyme de novo design, function investigation, and inhibitor development. Here, we report a method named MeCOM for comparing metalloenzyme active sites. It is characterized by metal ion(s) centric active site recognition and three-dimensional superimposition using α-carbon or pharmacophore features. The test results revealed that for the given metalloenzymes, MeCOM could effectively recognize the active sites, extract active site features, and superimpose the active sites; it also could correctly identify similar active sites, differentiate dissimilar active sites, and evaluate the similarity degree. Moreover, MeCOM showed potential to establish new associations between structurally distinct metalloenzymes by active site comparison. MeCOM is freely available at https://mecom.ddtmlab.org.
Subject(s)
Metalloproteins , Catalytic Domain , Metalloproteins/chemistry , Metals , MethyltransferasesABSTRACT
As one of important mechanisms to ß-lactam antimicrobial resistance, metallo-ß-lactamases (MBLs) have been receiving increasing worldwide attentions. Ambler subclass B1 MBLs are most clinically relevant, because they can hydrolyze almost all ß-lactams with the exception of monobactams. However, it is still lacking of clinically useful drugs to combat MBL-medicated resistance. We previously identified 1H-imidazole-2-carboxylic acid as a core metal-binding pharmacophore (MBP) to target multiple B1 MBLs. Herein, we report structural optimization of 1H-imidazole-2-carboxylic acid and substituents. Structure-activity relationship (SAR) analyses revealed that replacement of 1H-imidazole-2-carboxylic acid with other structurally highly similar MBPs excepting thiazole-4-carboxylic acid resulted in decreased MBL inhibition. Further SAR studies identified more potent inhibitors to MBLs, of which 28 manifested IC50 values of 0.018 µM for both VIM-2 and VIM-5. The microbiological tests demonstrated that the most tested compounds showed improved synergistic effects; some compounds at 1 µg/ml were able to reduce meropenem MIC by at least 16-fold, which will be worth further development of new potent inhibitors particularly targeting VIM-type MBLs.
Subject(s)
beta-Lactamase Inhibitors , beta-Lactamases , Anti-Bacterial Agents/chemistry , Anti-Bacterial Agents/pharmacology , Carboxylic Acids/pharmacology , Imidazoles , Meropenem , Monobactams , Thiazoles , beta-Lactamase Inhibitors/chemistry , beta-Lactamase Inhibitors/pharmacology , beta-Lactamases/chemistry , beta-LactamsABSTRACT
MOTIVATION: Metalloenzymes are attractive targets for therapeutic intervention owing to their central roles in various biological processes and pathological situations. The fast-growing body of structural data on metalloenzyme-ligand interactions is facilitating efficient drug discovery targeting metalloenzymes. However, there remains a shortage of specific databases that can provide centralized, interconnected information exclusive to metalloenzyme-ligand associations. RESULTS: We created a Metalloenzyme-Ligand Association Database (MeLAD), which is designed to provide curated structural data and information exclusive to metalloenzyme-ligand interactions, and more uniquely, present expanded associations that are represented by metal-binding pharmacophores (MBPs), metalloenzyme structural similarity (MeSIM) and ligand chemical similarity (LigSIM). MeLAD currently contains 6086 structurally resolved interactions of 1416 metalloenzymes with 3564 ligands, of which classical metal-binding, non-classical metal-binding, non-metal-binding and metal water-bridging interactions account for 63.0%, 2.3%, 34.4% and 0.3%, respectively. A total of 263 monodentate, 191 bidentate and 15 tridentate MBP chemotypes were included in MeLAD, which are linked to different active site metal ions and coordination modes. 3726 and 52 740 deductive metalloenzyme-ligand associations by MeSIM and LigSIM analyses, respectively, were included in MeLAD. An online server is provided for users to conduct metalloenzyme profiling prediction for small molecules of interest. MeLAD is searchable by multiple criteria, e.g. metalloenzyme name, ligand identifier, functional class, bioinorganic class, metal ion and metal-containing cofactor, which will serve as a valuable, integrative data source to foster metalloenzyme related research, particularly involved in drug discovery targeting metalloenzymes. AVAILABILITY AND IMPLEMENTATION: MeLAD is accessible at https://melad.ddtmlab.org. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
Subject(s)
Metalloproteins , Catalytic Domain , Drug Discovery , Ligands , MetalsABSTRACT
The production of ß-lactamases represents the main cause of resistance to clinically important ß-lactam antibiotics. Boron containing compounds have been demonstrated as promising broad-spectrum ß-lactamase inhibitors to combat ß-lactam resistance. Here we report a series of 3-aryl substituted benzoxaborole derivatives, which manifested broad-spectrum inhibition to representative serine-ß-lactamases (SBLs) and metallo-ß-lactamases (MBLs). The most potent inhibitor 9f displayed an IC50 value of 86 nM to KPC-2 SBL and micromolar inhibitory activity towards other tested enzymes. Cell-based assays further revealed that 9f was able to significantly reduce the MICs of meropenem in clinically isolated KPC-2-producing bacterial strains and it showed no apparent toxicity in HEK293T cells.
Subject(s)
Boron Compounds/pharmacology , beta-Lactamase Inhibitors/chemical synthesis , beta-Lactamase Inhibitors/pharmacology , Binding Sites , Boron Compounds/chemical synthesis , Boron Compounds/chemistry , Escherichia coli/drug effects , Escherichia coli/metabolism , HEK293 Cells , Humans , Inhibitory Concentration 50 , Klebsiella pneumoniae/drug effects , Klebsiella pneumoniae/metabolism , Meropenem/pharmacology , Models, Molecular , Molecular Structure , Protein Conformation , beta-Lactamase Inhibitors/chemistryABSTRACT
Resistance to ß-lactam antibacterials is commonly associated with the production of the serine ß-lactamases (SBLs) and/or metallo-ß-lactamases (MBLs). Although clinically useful inhibitors for the SBLs have been developed, no equivalent inhibitors are available for the MBLs, which can hydrolyze almost all ß-lactam antibiotics, including the so-called "last resort" carbapenems. It is still a challenging task to develop a clinically useful inhibitor that should be broad-spectrum targeting multiple clinically relevant MBL enzymes that differ in their active site features. This review provides a detailed description of interaction modes of substrates and small-molecule inhibitors with various MBL enzymes and highlights the importance of metal- and "anchor residue"-binding features to achieve broad-spectrum MBL inhibition. Recently emerging active site interference strategies include metal ion deprivation, metal ion replacement, and cysteine modification as challenging, but worth experimenting directions for inhibitor development. The metalloenzyme selectivity, metal-binding pharmacophore, and cellular permeability and accumulation should be properly considered in the further development of clinically useful inhibitors to combat MBL-mediated antibacterial resistance.
Subject(s)
beta-Lactamase Inhibitors , beta-Lactamases , Anti-Bacterial Agents/pharmacology , Catalytic Domain , Drug Resistance, Bacterial , Humans , beta-Lactamase Inhibitors/pharmacology , beta-Lactamases/metabolismABSTRACT
Rosmarinic acid (RA), a polyphenolic phytochemical, has broad-spectrum biological and pharmacological activity. A virtual target screening method termed IFPTarget combined with enzyme inhibition assays led to the identification of the clinically relevant metallo-ß-lactamase (MBL) VIM-2 as one of unexploited targets of RA. The enzyme kinetic studies indicated that RA is a fully reversible, substrate-competitive VIM-2 inhibitor. The isothermal titration calorimetry (ITC) analyses revealed that the initial binding of RA to VIM-2 is mainly due to enthalpy contribution. Further inhibition assays with RA related compounds revealed that salvianolic acid A, a derivative of RA, manifests potent inhibition to VIM-2, more interestingly, which shows inhibitory activity against the NDM-1, another clinically relevant MBL subtype, and the serine-ß-lactamase TEM-1 that is structurally and mechanistically distinct from the VIM-2 and NDM-1.
Subject(s)
Caffeic Acids/pharmacology , Cinnamates/pharmacology , Depsides/pharmacology , Lactates/pharmacology , beta-Lactamases/metabolism , Caffeic Acids/chemistry , Cinnamates/chemistry , Depsides/chemistry , Dose-Response Relationship, Drug , Drug Evaluation, Preclinical , Humans , Lactates/chemistry , Molecular Structure , Structure-Activity Relationship , Rosmarinic AcidABSTRACT
Maternal embryonic leucine zipper kinase (MELK), a serine/threonine protein kinase, has oncogenic properties and plays a key functional role in various cancer cells. Although MELK may not be a cancer addiction target, the development of specific MELK inhibitors would provide useful chemical tools for synthetic lethal investigation. Herein, we identified several hit compounds using a customized structure-based virtual screening, among which compounds 4 and 16 showed the most potent inhibition to MELK with IC50 values of 3.52 µM and 178.3 nM, respectively. In vitro cell-based assays revealed that 16 has no effect on the growth of various types of cancer cells, but has the potential to inhibit cancer cell migration and invasion. Western blotting analyses revealed that 16 suppresses the phosphorylation of focal adhesion kinase (FAK), a downstream molecule of MELK, which is a key kinase in regulating cancer cell migration and invasion.
Subject(s)
Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Drug Discovery , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/pharmacology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Cell Line, Tumor , Cell Movement/drug effects , Focal Adhesion Kinase 1/metabolism , Humans , Neoplasm Invasiveness , Signal Transduction/drug effectsABSTRACT
Small-molecule target identification is an important and challenging task for chemical biology and drug discovery. Structure-based virtual target identification has been widely used, which infers and prioritizes potential protein targets for the molecule of interest (MOI) principally via a scoring function. However, current "universal" scoring functions may not always accurately identify targets to which the MOI binds from the retrieved target database, in part due to a lack of consideration of the important binding features for an individual target. Here, we present IFPTarget, a customized virtual target identification method, which uses an interaction fingerprinting (IFP) method for target-specific interaction analyses and a comprehensive index (Cvalue) for target ranking. Evaluation results indicate that the IFP method enables substantially improved binding pose prediction, and Cvalue has an excellent performance in target ranking for the test set. When applied to screen against our established target library that contains 11,863 protein structures covering 2842 unique targets, IFPTarget could retrieve known targets within the top-ranked list and identified new potential targets for chemically diverse drugs. IFPTarget prediction led to the identification of the metallo-ß-lactamase VIM-2 as a target for quercetin as validated by enzymatic inhibition assays. This study provides a new in silico target identification tool and will aid future efforts to develop new target-customized methods for target identification.
Subject(s)
Models, Molecular , Proteins/metabolism , Amino Acid Sequence , Binding Sites , Hydrogen Bonding , Hydrophobic and Hydrophilic Interactions , Ligands , Protein Binding , Protein Conformation , Proteins/chemistry , Substrate Specificity , beta-Lactamases/chemistry , beta-Lactamases/metabolismABSTRACT
Resistance to ß-lactam antibiotics mediated by metallo-ß-lactamases (MBLs) is a growing problem. We describe the use of protein-observe 19 F-NMR (PrOFâ NMR) to study the dynamics of the São Paulo MBL (SPM-1) from ß-lactam-resistant Pseudomonas aeruginosa. Cysteinyl variants on the α3 and L3 regions, which flank the di-ZnII active site, were selectively 19 F-labeled using 3-bromo-1,1,1-trifluoroacetone. The PrOFâ NMR results reveal roles for the mobile α3 and L3 regions in the binding of both inhibitors and hydrolyzed ß-lactam products to SPM-1. These results have implications for the mechanisms and inhibition of MBLs by ß-lactams and non-ß-lactams and illustrate the utility of PrOFâ NMR for efficiently analyzing metal chelation, identifying new binding modes, and studying protein binding from a mixture of equilibrating isomers.
Subject(s)
Fluorine-19 Magnetic Resonance Imaging , beta-Lactamase Inhibitors/pharmacology , beta-Lactamases/metabolism , Binding Sites/drug effects , Models, Molecular , Molecular Conformation , beta-Lactamase Inhibitors/chemical synthesis , beta-Lactamase Inhibitors/chemistryABSTRACT
The development of amyloid-specific fluorophores allows the visualization of cerebral ß-amyloid deposits using optical imaging technology. In the present study, a series of smart styrylpyran fluorophores with compact donor-acceptor architecture were designed and evaluated for noninvasive detection of cerebral ß-amyloid deposits. Spectral behavior of the fluorophores changed significantly (optical turn-on) upon binding to ß-amyloid aggregates. Computational studies were conducted to correlate the experimental Kd values with calculated binding energies, speculating the relationship between fluorophore structure and ß-amyloid affinity. In vivo studies demonstrated that PAD-2 could discriminate APP/PS1 transgenic mice from wild type controls, with specific labeling of cerebral ß-amyloid deposits confirmed by ex vivo observation. Collectively, these styrylpyran fluorophores could provide a new scaffold for the development of optical imaging probes targeting cerebral ß-amyloid deposits.
Subject(s)
Alzheimer Disease/pathology , Amyloid/ultrastructure , Fluorescent Dyes/chemical synthesis , Plaque, Amyloid/pathology , Pyrans/chemical synthesis , Styrenes/chemical synthesis , Alzheimer Disease/diagnosis , Alzheimer Disease/metabolism , Amino Acid Motifs , Amyloid/metabolism , Animals , Blood-Brain Barrier/metabolism , Brain/metabolism , Brain/pathology , Brain/ultrastructure , Disease Models, Animal , Fluorescent Dyes/administration & dosage , Fluorescent Dyes/metabolism , Male , Mice , Mice, Transgenic , Molecular Docking Simulation , Molecular Sequence Data , Optical Imaging , Plaque, Amyloid/diagnosis , Plaque, Amyloid/metabolism , Protein Binding , Protein Structure, Secondary , Pyrans/administration & dosage , Pyrans/metabolism , Structure-Activity Relationship , Styrenes/administration & dosage , Styrenes/metabolismABSTRACT
Epigenetic modifications are critical mechanisms that regulate many biological processes and establish normal cellular phenotypes. Aberrant epigenetic modifications are frequently linked to the development and maintenance of several diseases including cancer, inflammation and metabolic diseases and so on. The key proteins that mediate epigenetic modifications have been thus recognized as potential therapeutic targets for these diseases. Consequently, discovery of small molecule inhibitors for epigenetic targets has received considerable attention in recent years. Here, virtual screening methods and their applications in the discovery of epigenetic target inhibitors are the focus of this review. Newly emerging approaches or strategies including rescoring methods, docking pose filtering methods, machine learning methods and 3D molecular similarity methods were also underlined. They are expected to be employed for identifying novel inhibitors targeting epigenetic regulation more efficiently.
Subject(s)
Drug Discovery/methods , Epigenesis, Genetic , Artificial Intelligence , Binding Sites , Computer Simulation , DNA-Cytosine Methylases/antagonists & inhibitors , DNA-Cytosine Methylases/chemistry , Drug Evaluation, Preclinical/methods , Histone Deacetylase Inhibitors/chemistry , Histone Deacetylases/chemistry , Models, Molecular , Protein Structure, Tertiary , Small Molecule Libraries , SoftwareABSTRACT
BACKGROUND: The enzymatic chemistry method is currently the most widely used method for the rapid detection of organophosphorus (OP) pesticides, but the enzymes used, such as cholinesterases, lack sufficient sensitivity to detect low concentrations of OP pesticides present in given samples. Serine hydrolase is considered an ideal enzyme source in seeking high-sensitivity enzymes used for OP pesticide detection. However, it is difficult to systematically evaluate sensitivities of various serine hydrolases to OP pesticides by in vitro experiments. This study aimed to establish an in silico method to predict the sensitivity spectrum of various serine hydrolases to OP pesticides. RESULTS: A serine hydrolase database containing 219 representative serine hydrolases was constructed. Based on this database, an integrated molecular docking and rescoring method was established, in which the AutoDock Vina program was used to produce the binding poses of OP pesticides to various serine hydrolases and the ID-Score method developed recently by us was adopted as a rescoring method to predict their binding affinities. In retrospective case studies, this method showed good performance in predicting the sensitivities of known serine hydrolases to two OP pesticides: paraoxon and diisopropyl fluorophosphate. The sensitivity spectrum of the 219 collected serine hydrolases to 37 commonly used OP pesticides was finally obtained using this method. CONCLUSION: Overall, this study presented a promising in silico tool to predict the sensitivity spectrum of various serine hydrolases to OP pesticides, which will help in finding high-sensitivity serine hydrolases for OP pesticide detection.
Subject(s)
Computer Simulation , Models, Chemical , Organophosphorus Compounds/pharmacology , Pesticides/pharmacology , Serine Endopeptidases/metabolism , Serine Proteinase Inhibitors/pharmacology , Binding Sites , Databases, Factual , Models, Molecular , Organophosphorus Compounds/chemistry , Pesticides/chemistry , Protein Conformation , Serine Endopeptidases/chemistry , Serine Proteinase Inhibitors/chemistryABSTRACT
A potential fluorescence probe for in vivo detection of cerebral ß-amyloid fibrils, (E)-2-(2-(2-(5-(dimethylamino)thiophen-2-yl)vinyl)-6-methyl-4H-pyran-4-ylidene)malononitrile (PT-1), was synthesized and evaluated. In experiments in vitro, PT-1 exhibited clear labeling of ß-amyloid fibrils and significant fluorescence changes upon binding to aggregated ß-amyloid fibrils. It also showed favorite kinetics in the brain, which is critical for cerebral imaging. In vivo fluorescence imaging with PT-1 and semi-quantitative analysis of the images further confirmed noninvasive visualization of cerebral ß-amyloid fibrils in vivo and obvious distinction between APP/PS1 transgenic mice and wild-type controls. The results demonstrate the potential of PT-1 as a novel fluorescence probe for noninvasive prediction of cerebral ß-amyloid fibrils.
Subject(s)
Amyloid beta-Peptides/analysis , Fluorescent Dyes/chemistry , Nitriles/chemistry , Thiophenes/chemistry , Animals , Fluorescence , Fluorescent Dyes/administration & dosage , Fluorescent Dyes/chemical synthesis , Injections, Intravenous , Mice , Mice, Inbred C57BL , Mice, Transgenic , Models, Molecular , Molecular Structure , Nitriles/chemical synthesis , Thiophenes/chemical synthesisABSTRACT
COP9 signalosome catalytic subunit CSN5 plays a key role in tumorigenesis and tumor immunity, showing potential as an anticancer target. Currently, only a few CSN5 inhibitors have been reported, at least partially, due to the challenges in establishing assays for CSN5 deubiquitinase activity. Here, we present the establishment and validation of a simple and reliable non-catalytic activity assay platform for identifying CSN5 inhibitors utilizing a new fluorescent probe, CFP-1, that exhibits enhanced fluorescence and fluorescence polarization features upon binding to CSN5. By using this platform, we identified 2-aminothiazole-4-carboxylic acids as new CSN5 inhibitors, which inhibited CSN5 but slightly downregulated PD-L1 in cancer cells. Furthermore, through the integration of deep learning-enabled virtual screening, we discovered that shikonins are nanomolar CSN5 inhibitors, which can upregulate PD-L1 in HCT116 cells. The binding modes of these structurally distinct inhibitors with CSN5 were explored by using microsecond-scale molecular dynamics simulations and tryptophan quenching assays.