title: Simplified assays of lipolysis enzymes for drug discovery and specificity assessment of known inhibitors.
abstract: Lipids are used as cellular building blocks and condensed energy stores and also act as signaling molecules. The glycerolipid/ fatty acid cycle, encompassing lipolysis and lipogenesis, generates many lipid signals. Reliable procedures are not available for measuring activities of several lipolytic enzymes for the purposes of drug screening, and this resulted in questionable selectivity of various known lipase inhibitors. We now describe simple assays for lipolytic enzymes, including adipose triglyceride lipase (ATGL), hormone sensitive lipase (HSL), sn-1-diacylglycerol lipase (DAGL), monoacylglycerol lipase, α/β-hydrolase domain 6, and carboxylesterase 1 (CES1) using recombinant human and mouse enzymes either in cell extracts or using purified enzymes. We observed that many of the reported inhibitors lack specificity. Thus, Cay10499 (HSL inhibitor) and RHC20867 (DAGL inhibitor) also inhibit other lipases. Marked differences in the inhibitor sensitivities of human ATGL and HSL compared with the corresponding mouse enzymes was noticed. Thus, ATGListatin inhibited mouse ATGL but not human ATGL, and the HSL inhibitors WWL11 and Compound 13f were effective against mouse enzyme but much less potent against human enzyme. Many of these lipase inhibitors also inhibited human CES1. Results describe reliable assays for measuring lipase activities that are amenable for drug screening and also caution about the specificity of the many earlier described lipase inhibitors.
authors.affiliation: Molecular Nutrition Unit and Montreal Diabetes Research Center, CRCHUM, Montréal, Québec, Canada H2X 0A9 Departments of Nutrition, Biochemistry, and Molecular Medicine, Université de Montréal, Montréal, Québec, Canada H2X 0A9.
authors.forename: Jose
authors.initials: J
authors.lastname: Iglesias
authors.affiliation: Molecular Nutrition Unit and Montreal Diabetes Research Center, CRCHUM, Montréal, Québec, Canada H2X 0A9 Departments of Nutrition, Biochemistry, and Molecular Medicine, Université de Montréal, Montréal, Québec, Canada H2X 0A9.
authors.forename: Julien
authors.initials: J
authors.lastname: Lamontagne
authors.affiliation: Molecular Nutrition Unit and Montreal Diabetes Research Center, CRCHUM, Montréal, Québec, Canada H2X 0A9 Departments of Nutrition, Biochemistry, and Molecular Medicine, Université de Montréal, Montréal, Québec, Canada H2X 0A9.
authors.forename: Heidi
authors.initials: H
authors.lastname: Erb
authors.affiliation: Molecular Nutrition Unit and Montreal Diabetes Research Center, CRCHUM, Montréal, Québec, Canada H2X 0A9 Departments of Nutrition, Biochemistry, and Molecular Medicine, Université de Montréal, Montréal, Québec, Canada H2X 0A9.
authors.forename: Sari
authors.initials: S
authors.lastname: Gezzar
authors.affiliation: Molecular Nutrition Unit and Montreal Diabetes Research Center, CRCHUM, Montréal, Québec, Canada H2X 0A9 Departments of Nutrition, Biochemistry, and Molecular Medicine, Université de Montréal, Montréal, Québec, Canada H2X 0A9.
authors.forename: Shangang
authors.initials: S
authors.lastname: Zhao
authors.affiliation: Molecular Nutrition Unit and Montreal Diabetes Research Center, CRCHUM, Montréal, Québec, Canada H2X 0A9 Departments of Nutrition, Biochemistry, and Molecular Medicine, Université de Montréal, Montréal, Québec, Canada H2X 0A9.
authors.forename: Erik
authors.initials: E
authors.lastname: Joly
authors.affiliation: NuChem Therapeutics, Montréal, Québec, Canada, H4P 2R2.
authors.forename: Vouy Linh
authors.initials: VL
authors.lastname: Truong
authors.affiliation: NuChem Therapeutics, Montréal, Québec, Canada, H4P 2R2.
authors.forename: Kathryn
authors.initials: K
authors.lastname: Skorey
authors.affiliation: NuChem Therapeutics, Montréal, Québec, Canada, H4P 2R2.
authors.forename: Sheldon
authors.initials: S
authors.lastname: Crane
authors.affiliation: Molecular Nutrition Unit and Montreal Diabetes Research Center, CRCHUM, Montréal, Québec, Canada H2X 0A9 Departments of Nutrition, Biochemistry, and Molecular Medicine, Université de Montréal, Montréal, Québec, Canada H2X 0A9 marc.prentki@umontreal.ca murthy.madiraju@crchum.qc.ca.
authors.forename: S R Murthy
authors.initials: SR
authors.lastname: Madiraju
authors.affiliation: Molecular Nutrition Unit and Montreal Diabetes Research Center, CRCHUM, Montréal, Québec, Canada H2X 0A9 Departments of Nutrition, Biochemistry, and Molecular Medicine, Université de Montréal, Montréal, Québec, Canada H2X 0A9 marc.prentki@umontreal.ca murthy.madiraju@crchum.qc.ca.
authors.forename: Marc
authors.initials: M
authors.lastname: Prentki
mesh_terms: D000273:Adipose Tissue / Q000378:metabolism; D000818:Animals; D002265:Carboxylic Ester Hydrolases / Q000037:antagonists & inhibitors / Q000378:metabolism; D055808:Drug Discovery / Q000379:methods*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005227:Fatty Acids / Q000378:metabolism; D006801:Humans; D008049:Lipase / Q000037:antagonists & inhibitors / Q000378:metabolism; D050155:Lipogenesis / Q000502:physiology; D008066:Lipolysis / Q000187:drug effects* / Q000502:physiology; D008071:Lipoprotein Lipase / Q000037:antagonists & inhibitors / Q000378:metabolism; D051379:Mice; D008994:Monoacylglycerol Lipases / Q000378:metabolism; D002787:Sterol Esterase / Q000037:antagonists & inhibitors / Q000378:metabolism; D014280:Triglycerides / Q000378:metabolism; D000217:Acyltransferases
chemical_list: D002265:Carboxylic Ester Hydrolases; D004791:Enzyme Inhibitors; D005227:Fatty Acids; D008049:Lipase; D008071:Lipoprotein Lipase; D008994:Monoacylglycerol Lipases; D002787:Sterol Esterase; D014280:Triglycerides; C474609:CES1 protein, human; C494794:PNPLA2 protein, human; D000217:Acyltransferases; C494445:PNPLA2 protein, mouse
keywords: adipose triglyceride lipase; alpha/beta hydrolase domain 6; diacylglycerol lipase; fatty acid amide hydrolase; hormone sensitive lipase; monoacylglycerol lipase