ABSTRACT
Acute lung injury (ALI) is a life-threatening disease characterized by severe lung inflammation and intestinal microbiota disorder. The GPR18 receptor has been demonstrated to be a potential therapeutic target against ALI. Extracting Naringin dihydrochalcone (NDC) from the life-sustaining orange peel is known for its diverse anti-inflammatory properties, yet the specific action target remains uncertain. In the present study, we identified NDC as a potential agonist of the GPR18 receptor using virtual screening and investigated the pharmacological effects of NDC on sepsis-induced acute lung injury in rats and explored underlying mechanisms. In in vivo experiments, CLP-induced ALI model was established by cecum puncture and treated with NDC gavage one hour prior to drug administration, lung histopathology and inflammatory cytokines were evaluated, and feces were subjected to 16s rRNA sequencing and untargeted metabolomics analysis. In in vitro experiments, the anti-inflammatory properties were exerted by evaluating NDC targeting the GPR18 receptor to inhibit lipopolysaccharide (LPS)-induced secretion of TNF-α, IL-6, IL-1ß and activation of inflammatory signaling pathways in MH-S cells. Our findings showed that NDC significantly ameliorated lung damage and pro-inflammatory cytokine levels (TNF-α, IL-6, IL-1ß) in both cells and lung tissues via inhibiting the activation of STAT3, NF-κB, and NLRP3 inflammatory signaling pathways through GRP18 receptor activation. In addition, NDC can also partly reverse the imbalance of gut microbiota composition caused by CLP via increasing the proportion of Firmicutes/Bacteroidetes and Lactobacillus and decreasing the relative abundance of Proteobacteria. Meanwhile, the fecal metabolites in the NDC treatment group also significantly were changed, including decreased secretion of Phenylalanin, Glycine, and bile secretion, and increased secretion of Lysine. In conclusion, these findings suggest that NDC can alleviate sepsis-induced ALI via improving gut microbial homeostasis and metabolism and mitigate inflammation via activating GPR18 receptor. In conclusion, the results indicate that NDC, derived from the typical orange peel of food, could significantly contribute to development by enhancing intestinal microbial balance and metabolic processes, and reducing inflammation by activating the GPR18 receptor, thus mitigating sepsis-induced ALI and expanding the range of functional foods.
Subject(s)
Acute Lung Injury , Anti-Inflammatory Agents , Chalcones , Cytokines , Gastrointestinal Microbiome , Receptors, G-Protein-Coupled , Sepsis , Animals , Receptors, G-Protein-Coupled/metabolism , Gastrointestinal Microbiome/drug effects , Acute Lung Injury/drug therapy , Acute Lung Injury/pathology , Acute Lung Injury/etiology , Acute Lung Injury/microbiology , Acute Lung Injury/metabolism , Male , Sepsis/drug therapy , Sepsis/complications , Cytokines/metabolism , Rats , Chalcones/pharmacology , Chalcones/therapeutic use , Anti-Inflammatory Agents/therapeutic use , Anti-Inflammatory Agents/pharmacology , Rats, Sprague-Dawley , Homeostasis/drug effects , Cell Line , Lung/pathology , Lung/drug effects , Disease Models, Animal , Lipopolysaccharides , Humans , FlavanonesABSTRACT
Colorectal cancer (CRC) is a common malignant tumor with high morbidity and mortality rates worldwide. Although surgical resection and adjuvant radiotherapy/chemotherapy are the mainstays of CRC treatment, the efficacy is unsatisfactory due to several limitations, including high drug resistance. Accordingly, there is a dire need for new drugs or a novel combination approach to treat this patient population. Herein, we found that cinnamaldehyde (CA) could exert an antitumor effect in HCT-116 cell lines. Target fishing, molecular imaging, and live-cell tracing using an alkynyl-CA probe revealed that the heat shock 60 kDa protein 1 (HSPD1) protein was the target of CA. The covalent binding of CA with HSPD1 altered its stability. Furthermore, our results demonstrated that CA could induce cell apoptosis by inhibiting the PI3K/Akt signaling pathway and enhanced anti-CRC activity both in vitro and in vivo. Meanwhile, CA combined with different chemotherapeutic agents was beneficial to patients resistant to anti-CRC drug therapy.
ABSTRACT
Cyclin-dependent kinases 2 (CDK2) is a serine/threonine-protein kinase, which plays a key role in the regulation of cell cycle and is related to the occurrence and development of melanoma. In this study, we identified potent inhibitors for CDK2 by combining a multistage virtual screening strategy with bioassay validations. The biochemical activity of compounds was validated with ADP-Glo™ Kinase assay in vitro, and the results indicated that the biochemical activity of compound 1 (C1) was better than other selected compounds. Cell viability assay showed that the minimum inhibition concentration of C1 for CDK2 was lower than 4 µM. Further functional test results showed that C1 exerted significant antiproliferative, pro-apoptosis, and anti-migration activity in melanoma cell lines (A375 cells, WM35 cells, and A875 cells). Our findings suggested that the C1, virtually screened from compound libraries, as the novel inhibitor of CDK2, may be further developed as an effective therapeutic agent in the treatment of melanoma lines.
Subject(s)
CDC2-CDC28 Kinases , Melanoma , Humans , Cyclin-Dependent Kinases/metabolism , Protein Serine-Threonine Kinases/metabolism , Melanoma/drug therapy , Cell Line , Cyclin-Dependent Kinase 2ABSTRACT
Protein arginine methyltransferase 5 (PRMT5) is a major type II enzyme responsible for symmetric dimethylation of arginine (SDMA), and plays predominantly roles in human cancers, including in ovarian cancer. However, the exactly roles and underlying mechanisms of PRMT5 contributing to the progression of ovarian cancer mediated by reprogramming cell metabolism remain largely elusive. Here, we report that PRMT5 is highly expressed and correlates with poor survival in ovarian cancer. Knockdown or pharmaceutical inhibition of PRMT5 is sufficient to decrease glycolysis flux, attenuate tumor growth, and enhance the antitumor effect of Taxol. Mechanistically, we find that PRMT5 symmetrically dimethylates alpha-enolase (ENO1) at arginine 9 to promotes active ENO1 dimer formation, which increases glycolysis flux and accelerates tumor growth. Moreover, PRMT5 signals high glucose to increase the methylation modification of ENO1. Together, our data reveal a novel role of PRMT5 in promoting ovarian cancer growth by controlling glycolysis flux mediated by methylating ENO1, and highlights that PRMT5 may represent a promising therapeutic target for treating ovarian cancer.
ABSTRACT
Previous report showed that a variety of icotinib derivatives bearing different 1,2,3-triazole moieties, which could be readily prepared via copper (I)-catalyzed cycloaddition (CuAAC) reaction between icotinib and different azides, exhibited interesting activity against different lung cancer cell lines such as H460, H1975, H1299, A549 or PC-9. To further expand the application scope of the compounds and to validate the function of triazole groups in drug design, the anti-cancer activity of these compounds against esophageal squamous carcinoma (ESCC) cells was tested herein. Preliminary MTT experiments suggested that these compounds were active against different ESCC cell lines such as KYSE70, KYSE410, or KYSE450 as well as their drug-resistant ones. Especially, compound 3l showed interesting anticancer activity against these cell lines. The mode of action was studied via molecular docking, SPR experiments and other biochemical studies, and 3l exhibited higher binding potential to wild-type EGFR than icotinib did. In vivo anticancer study showed that 3l could inhibit tumor growth of cell-line-derived xenografts in ESCC. Study also suggested that 3l was a potent inhibitor for EGFR-TK pathway. Combining these results, 3l represents a promising lead compound for the design of anti-cancer drugs against ESCC.
ABSTRACT
SCOPE: Glutamate (Glu) and γ-aminobutyric acid (GABA) are the major excitatory and inhibitory neurotransmitters that control information flow in the brain. GABA dysfunction is a general vulnerability factor for mental illness. Cinnamaldehyde (CA) is found to have sedation in a mental illness model. However, the specific targets and molecular mechanisms related to the sedative effects of CA have not been elucidated. METHODS AND RESULTS: Metabolomics analysis and target fishing showed CA could increase the expression of GABA in vivo, and α-enolase (ENO1) is the primary target protein of CA associated with sedation. CA mainly binds with ENO1 in the cerebellar granular layer of brain, which influences the first transformations of the input signals arriving in the cerebellar cortex. The α,ß-unsaturated aldehyde group of CA blocks the hydroxy group of Ser40, which induces a loss in ENO1 activation. CA also disturbs the glycolysis pathway and influences the tricarboxylic acid cycle and oxidative phosphorylation, which activate gluconeogenesis to provide energy to the brain. This mechanism is verified in zebrafish with ENO1 or glutamic acid decarboxylase (GAD) deficiency. CONCLUSIONS: CA demonstrates sedation and alleviates GABA dysfunction via covalent binding ENO1, which shows the potential to improve the therapy of mental illness.
Subject(s)
Zebrafish , gamma-Aminobutyric Acid , Acrolein/analogs & derivatives , Animals , Glutamate Decarboxylase/metabolism , Phosphopyruvate Hydratase/genetics , Phosphopyruvate Hydratase/metabolism , Zebrafish/metabolism , gamma-Aminobutyric Acid/metabolism , gamma-Aminobutyric Acid/pharmacologyABSTRACT
Acute lung injury/acute respiratory distress syndrome (ALI/ARDS) is one of the most common complications in COVID-19. Elastase has been recognized as an important target to prevent ALI/ARDS in the patient of COVID-19. Cyclotheonellazole A (CTL-A) is a natural macrocyclic peptide reported to be a potent elastase inhibitor. Herein, we completed the first total synthesis of CTL-A in 24 linear steps. The key reactions include three-component MAC reactions and two late-stage oxidations. We also provided seven CTL-A analogues and elucidated preliminary structure-activity relationships. The in vivo ALI mouse model further suggested that CTL-A alleviated acute lung injury with reductions in lung edema and pathological deterioration, which is better than sivelestat, one approved elastase inhibitor. The activity of CTL-A against elastase, along with its cellular safety and well-established synthetic route, warrants further investigation of CTL-A as a candidate against COVID-19 pathogeneses.
Subject(s)
Acute Lung Injury/drug therapy , Leukocyte Elastase/antagonists & inhibitors , Peptides, Cyclic/pharmacology , Respiratory Distress Syndrome/drug therapy , Serine Proteinase Inhibitors/pharmacology , Acute Lung Injury/chemically induced , Acute Lung Injury/metabolism , Animals , Bleomycin , COVID-19/metabolism , COVID-19/pathology , Cell Line , Disease Models, Animal , Humans , Leukocyte Elastase/metabolism , Male , Mice , Mice, Inbred C57BL , Peptides, Cyclic/chemical synthesis , Peptides, Cyclic/chemistry , Respiratory Distress Syndrome/chemically induced , Respiratory Distress Syndrome/metabolism , Serine Proteinase Inhibitors/chemical synthesis , Serine Proteinase Inhibitors/chemistry , COVID-19 Drug TreatmentABSTRACT
The endocannabinoid system plays an essential role in the regulation of analgesia and human immunity, and Cannabinoid Receptor 2 (CB2) has been proved to be an ideal target for the treatment of liver diseases and some cancers. In this study, we identified CB2 antagonists using a three-step "deep learning-pharmacophore-molecular docking" virtual screening approach. From the ChemDiv database (1,178,506 compounds), 15 hits were selected and tested by radioligand binding assays and cAMP functional assays. A total of 7 out of the 15 hits were found to exhibit binding affinities in the radioligand binding assays against CB2 receptor, with a pKi of 5.15-6.66, among which five compounds showed antagonistic activities with pIC50 of 5.25-6.93 in the cAMP functional assays. Among these hits, Compound 8 with the 4H-pyrido[1,2-a]pyrimidin-4-one scaffold showed the best binding affinity and antagonistic activity with a pKi of 6.66 and pIC50 of 6.93, respectively. The new scaffold could serve as a lead for further development of CB2 drugs. Additionally, we hope that the model in this study could be further utilized to identify more novel CB2 receptor antagonists, and the developed approach could also be used to design potent ligands for other therapeutic targets.
Subject(s)
Drug Discovery , Ligands , Molecular Docking Simulation , Molecular Dynamics Simulation , Receptor, Cannabinoid, CB2/chemistry , Amino Acids , Binding Sites , Computational Biology/methods , Deep Learning , Drug Discovery/methods , Humans , Molecular Conformation , Molecular Structure , Protein Binding , Quantitative Structure-Activity Relationship , Receptor, Cannabinoid, CB2/antagonists & inhibitors , Small Molecule Libraries , WorkflowABSTRACT
Adenosine receptors (ARs) have been demonstrated to be potential therapeutic targets against Parkinson's disease (PD). In the present study, we describe a multistage virtual screening approach that identifies dual adenosine A1 and A2A receptor antagonists using deep learning, pharmacophore models, and molecular docking methods. Nineteen hits from the ChemDiv library containing 1,178,506 compounds were selected and further tested by in vitro assays (cAMP functional assay and radioligand binding assay); of these hits, two compounds (C8 and C9) with 1,2,4-triazole scaffolds possessing the most potent binding affinity and antagonistic activity for A1/A2A ARs at the nanomolar level (pKi of 7.16-7.49 and pIC50 of 6.31-6.78) were identified. Further molecular dynamics (MD) simulations suggested similarly strong binding interactions of the complexes between the A1/A2A ARs and two compounds (C8 and C9). Notably, the 1,2,4-triazole derivatives (compounds C8 and C9) were identified as the most potent dual A1/A2A AR antagonists in our study and could serve as a basis for further development. The effective multistage screening approach developed in this study can be utilized to identify potent ligands for other drug targets.
Subject(s)
Adenosine A1 Receptor Antagonists , Adenosine A2 Receptor Antagonists , Deep Learning , Drug Discovery/methods , Humans , Molecular Docking Simulation , Parkinson Disease , Protein Binding , Receptor, Adenosine A2A/chemistry , Receptor, Adenosine A2A/genetics , Receptor, Adenosine A2A/metabolismABSTRACT
Alcoholic liver disease (ALD) is one of the pathogenic factors of chronic liver disease with the highest clinical morbidity worldwide. Ursolic acid (UA), a pentacyclic terpenoid carboxylic acid, has shown many health benefits including antioxidative, anti-inflammatory, anticancer, and hepatoprotective activities. We previously found that UA was metabolized in vivo into epoxy-modified UA containing an epoxy electrophilic group and had the potential to react with nucleophilic groups. In this study we prepared an alkynyl-modified UA (AM-UA) probe for tracing and capturing the target protein of UA from liver in mice, then investigated the mode by which UA bound to its target in vivo. By conducting proteome identification and bioinformatics analysis, we identified caspase-3 (CASP3) as the primary target protein of UA associated with liver protection. Molecule docking analysis showed that the epoxy group of the UA metabolite reacted with Cys-163 of CASP3, forming a covalent bond with CASP3. The binding mode of the UA metabolites (UA, CM-UA, and EM-UA) was verified by biochemical evaluation, demonstrating that the epoxy group produced by metabolism played an important role in the inhibition of CASP3. In alcohol-treated HepG2 cells, pretreatment with the UA metabolite (10 µM) irreversibly inhibited CASP3 activities, and subsequently decreased the cleavage of PARP and cell apoptosis. Finally, pre-administration of UA (20-80 mg· kg-1 per day, ig, for 1 week) dose-dependently alleviated alcohol-induced liver injury in mice mainly via the inhibition of CASP3. In conclusion, this study demonstrates that UA is a valuable lead compound for the treatment of ALD.
Subject(s)
Apoptosis/drug effects , Caspase 3/metabolism , Caspase Inhibitors/therapeutic use , Liver Diseases, Alcoholic/drug therapy , Liver/drug effects , Triterpenes/therapeutic use , Amino Acid Sequence , Animals , Caspase 3/chemistry , Caspase Inhibitors/metabolism , Cysteine/chemistry , Epoxy Compounds/chemistry , Epoxy Compounds/therapeutic use , Hep G2 Cells , Hepatocytes/drug effects , Humans , Liver/enzymology , Liver/pathology , Liver Diseases, Alcoholic/enzymology , Liver Diseases, Alcoholic/pathology , Male , Mice , Molecular Docking Simulation , Poly(ADP-ribose) Polymerases/metabolism , Protein Binding , Sequence Alignment , Triterpenes/metabolism , Ursolic AcidABSTRACT
Idiopathic pulmonary fibrosis is a progressive-fibrosing lung disease with high mortality and limited therapy, which characterized by myofibroblasts proliferation and extracellular matrix deposition. Myricetin, a natural flavonoid, has been shown to possess a variety of biological characteristics including anti-inflammatory and anti-tumor. In this study we explored the potential effect and mechanisms of myricetin on pulmonary fibrosis in vivo and vitro. The in vivo studies showed that myricetin effectively alleviated bleomycin (BLM)-induced pulmonary fibrosis. KEGG analysis of RNA-seq data indicated that myricetin could regulate the transforming growth factor (TGF)-ß signaling pathway. In vitro studies indicated that myricetin could dose-dependently suppress TGF-ß1/Smad signaling and attenuate TGF-ß1-induced fibroblast activation and epithelial-mesenchymal transition (EMT). Molecular docking indicated that heat shock protein (HSP) 90ß may be a potential target of myricetin, and MST assay demonstrated that the dissociation constant (Kd) of myricetin and HSP90ß was 331.59 nM. We demonstrated that myricetin interfered with the binding of HSP90ß and TGF-ß receptor II and impeded fibroblast activation and EMT. In conclusion, myricetin impedes TGF-ß1-induced lung fibroblast activation and EMT via targeting HSP90ß and attenuates BLM-induced pulmonary fibrosis in mice.
Subject(s)
Flavonoids/pharmacology , HSP90 Heat-Shock Proteins/chemistry , Protective Agents/pharmacology , Pulmonary Fibrosis/drug therapy , Transforming Growth Factor beta/genetics , A549 Cells , Animals , Binding Sites , Bleomycin/administration & dosage , Cadherins/genetics , Cadherins/metabolism , Cell Proliferation/drug effects , Epithelial-Mesenchymal Transition , Extracellular Matrix/drug effects , Extracellular Matrix/metabolism , Fibronectins/genetics , Fibronectins/metabolism , Gene Expression Regulation , HSP90 Heat-Shock Proteins/antagonists & inhibitors , HSP90 Heat-Shock Proteins/genetics , HSP90 Heat-Shock Proteins/metabolism , Humans , Lung/drug effects , Lung/metabolism , Lung/pathology , Male , Mice , Mice, Inbred C57BL , Molecular Docking Simulation , Protein Binding , Pulmonary Fibrosis/chemically induced , Pulmonary Fibrosis/metabolism , Pulmonary Fibrosis/pathology , Signal Transduction , Smad Proteins/genetics , Smad Proteins/metabolism , Transforming Growth Factor beta/metabolism , Vimentin/genetics , Vimentin/metabolismABSTRACT
The use of antagonists for each adenosine receptor (AR) subtype as potent clinical candidates is of growing interest due to their involvement in the treatment of various diseases. The recent resolution of several A1 and A2A ARs X-ray structures provides opportunities for structure-based drug design. In this study, we describe the discovery of novel A1AR antagonists by applying a multistage virtual screening approach, which is based on random forest (RF), e-pharmacophore modeling and docking methods. A multistage virtual screening approach was applied to screen the ChemDiv library (1,492,362 compounds). Among the final hits, 22 compounds were selected for further radioligand binding assay analysis against human A1AR, and 18 compounds (81.82% success) exhibited nanomolar or low micromolar binding potency (Ki). Then, we selected six compounds (pKi > 6) to further evaluate their antagonist profile in a cAMP functional assay, and we found that they had low micromolar antagonistic activity (pIC50 = 5.51-6.38) for the A1AR. Particularly, four of six compounds (pKi > 6) showed very good affinity (pKi = 6.11-7.13) and selectively (>100-fold) for A1AR over A2AAR. Moreover, the novelty analysis suggested that four of six compounds (pKi > 6) were dissimilar to existing A1AR antagonists and hence represented novel A1AR antagonists. Further molecular docking and molecular dynamics (MD) studies showed that the three selective compounds 15, 20 and 22 were stabilized (RMSlig value ≤ 2 Å) inside the binding pocket of A1AR with similar orientations to the docking pose in 100-ns MD simulations, whereas they escaped from the binding area of A2AAR with larger values of RMSlig (RMSlig ≥ 2 Å). We hope that these findings provide new insights into the discovery of drugs targeting A1AR and facilitate research on new drugs and treatments for A1AR-related human pathologies.
Subject(s)
Purinergic P1 Receptor Antagonists/pharmacology , Receptor, Adenosine A1/metabolism , Dose-Response Relationship, Drug , Drug Evaluation, Preclinical , Humans , Ligands , Models, Molecular , Molecular Structure , Purinergic P1 Receptor Antagonists/chemistry , Structure-Activity RelationshipABSTRACT
The adenosine A1 receptor (A1R) is one of four adenosine receptors in humans, which are involved in the function of the cardiovascular, respiratory and central nervous systems. Experimental results indicate that A1R can form a homodimer and that the protomer-protomer interaction in the A1R dimer is related to certain pharmacological characteristics of A1R activation. In this work, we performed docking, metadynamics simulation, conventional molecular dynamics simulations, Gaussian-accelerated molecular dynamics simulations, potential of mean force calculations, dynamic cross-correlation motions analysis and community network analysis to study the binding mode of 5'-N-ethylcarboxamidoadenosine (NECA) to A1R and the effect of dimerization on the activation of A1R. Our results show that NECA binds to A1R in a similar mode to adenosine in the A1R crystal structure and NECA in the A2AR crystal structure. The A1R homodimer can be activated by one or two agonists with NECA occupying its orthosteric pockets in one (which we call the NECA-A1R system) or both protomers (which we call the dNECA-A1R system). In the NECA-A1R system, activation is predicated in the protomer without NECA bound. In the dNECA-A1R system, only one protomer achieves the active state. These findings suggest an asymmetrical activation mechanism of the homodimer and a negative cooperativity between the two protomers. We envision that our results may further facilitate the drug development of A1R.