10.1021/acschembio.5b00628.s001 Anna Ohtera Anna Ohtera Yusaku Miyamae Yusaku Miyamae Kotaro Yoshida Kotaro Yoshida Kazuhiro Maejima Kazuhiro Maejima Toru Akita Toru Akita Akira Kakizuka Akira Kakizuka Kazuhiro Irie Kazuhiro Irie Seiji Masuda Seiji Masuda Taiho Kambe Taiho Kambe Masaya Nagao Masaya Nagao Identification of a New Type of Covalent PPARγ Agonist using a Ligand-Linking Strategy American Chemical Society 2015 activates PPAR γ PPAR γ PPAR γ agonist Covalent PPAR γ Agonist Cys 285 residue covalent PPAR γ agonists luciferase reporter assay GW 9662 covalently PPAR γ ligands PPAR γ LBP 2015-12-18 00:00:00 Journal contribution https://acs.figshare.com/articles/journal_contribution/Identification_of_a_New_Type_of_Covalent_PPAR_Agonist_using_a_Ligand_Linking_Strategy/2097058 Peroxisome proliferator-activated receptor γ (PPARγ) is a ligand-activated transcription factor that plays an important role in adipogenesis and glucose metabolism. The ligand-binding pocket (LBP) of PPARγ has a large Y-shaped cavity with multiple subpockets where multiple ligands can simultaneously bind and cooperatively activate PPARγ. Focusing on this unique property of the PPARγ LBP, we describe a novel two-step cell-based strategy to develop PPARγ ligands. First, a combination of ligands that cooperatively activates PPARγ was identified using a luciferase reporter assay. Second, hybrid ligands were designed and synthesized. For proof of concept, we focused on covalent agonists, which activate PPARγ through a unique activation mechanism regulated by a covalent linkage with the Cys285 residue in the PPARγ LBP. Despite their biological significance and pharmacological potential, few covalent PPARγ agonists are known except for endogenous fatty acid metabolites. With our strategy, we determined that plant-derived cinnamic acid derivatives cooperatively activated PPARγ by combining with GW9662, an irreversible antagonist. GW9662 covalently reacts with the Cys285 residue. A docking study predicted that a cinnamic acid derivative can bind to the open cavity in GW9662-bound PPARγ LBP. On the basis of the putative binding mode, structures of both ligands were linked successfully to create a potent PPARγ agonist, which enhanced the transactivation potential of PPARγ at submicromolar levels through covalent modification of Cys285. Our approach could lead to the discovery of novel high-potency PPARγ agonists.