Proteomics based approaches are emerging as useful tools to identify the targets of bioactive compounds and elucidate their molecular mechanisms of action. Here, we applied a chemical proteomic strategy to identify the peroxisome proliferator-activated receptor. (PPAR gamma) as a molecular target of the pro-apoptotic agent 15-ketoatractyligenin methyl ester (compound 1). We demonstrated that compound 1 interacts with PPAR gamma, forms a covalent bond with the thiol group of C285 and occupies the sub-pocket between helix H3 and the beta-sheet of the ligand-binding domain (LBD) of the receptor by Surface Plasmon Resonance (SPR), mass spectrometry-based studies and docking experiments. 1 displayed partial agonism of PPAR gamma in cell-based transactivation assays and was found to inhibit the AKT pathway, as well as its downstream targets. Consistently, a selective PPAR gamma antagonist (GW9662) greatly reduced the anti-proliferative and pro-apoptotic effects of 1, providing the molecular basis of its action. Collectively, we identified 1 as a novel PPAR gamma partial agonist and elucidated its mode of action, paving the way for therapeutic strategies aimed at tailoring novel PPAR gamma ligands with reduced undesired harmful side effects.

A compound-based proteomic approach discloses 15-ketoatractyligenin methyl ester as a new PPARγ partial agonist with anti-proliferative ability

Sabatino, Lina;Ziccardi, Pamela;Colantuoni, Vittorio;Lupo, Angelo;
2017-01-01

Abstract

Proteomics based approaches are emerging as useful tools to identify the targets of bioactive compounds and elucidate their molecular mechanisms of action. Here, we applied a chemical proteomic strategy to identify the peroxisome proliferator-activated receptor. (PPAR gamma) as a molecular target of the pro-apoptotic agent 15-ketoatractyligenin methyl ester (compound 1). We demonstrated that compound 1 interacts with PPAR gamma, forms a covalent bond with the thiol group of C285 and occupies the sub-pocket between helix H3 and the beta-sheet of the ligand-binding domain (LBD) of the receptor by Surface Plasmon Resonance (SPR), mass spectrometry-based studies and docking experiments. 1 displayed partial agonism of PPAR gamma in cell-based transactivation assays and was found to inhibit the AKT pathway, as well as its downstream targets. Consistently, a selective PPAR gamma antagonist (GW9662) greatly reduced the anti-proliferative and pro-apoptotic effects of 1, providing the molecular basis of its action. Collectively, we identified 1 as a novel PPAR gamma partial agonist and elucidated its mode of action, paving the way for therapeutic strategies aimed at tailoring novel PPAR gamma ligands with reduced undesired harmful side effects.
2017
Apoptosis
Binding Sites
Cell Proliferation
Diterpenes, Kaurane
Esters
HEK293 Cells
HT29 Cells
Humans
Jurkat Cells
Kinetics
Ligands
Molecular Docking Simulation
PPAR gamma
Protein Stability
Proteomics
Reproducibility of Results
Rosiglitazone
Surface Plasmon Resonance
Thermodynamics
Thiazolidinediones
Time Factors
Transcriptional Activation
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/20.500.12070/55217
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