title: Development of a RapidFire mass spectrometry assay and a fluorescence assay for the discovery of kynurenine aminotransferase II inhibitors to treat central nervous system disorders.
abstract: Kynurenine aminotransferases convert kynurenine to kynurenic acid and play an important role in the tryptophan degradation pathway. Kynurenic acid levels in brain have been hypothesized to be linked to a number of central nervous system (CNS) disorders. Kynurenine aminotransferase II (KATII) has proven to be a key modulator of kynurenic acid levels in brain and, thus, is an attractive target to treat CNS diseases. A sensitive, high-throughput, label-free RapidFire mass spectrometry assay has been developed for human KATII. Unlike other assays, this method is directly applicable to KATII enzymes from different animal species, which allows us to select proper animal model(s) to evaluate human KATII inhibitors. We also established a coupled fluorescence assay for human KATII. The short assay time and kinetic capability of the fluorescence assay provide a useful tool for orthogonal inhibitor validation and mechanistic studies.
authors.affiliation: Lead Discovery and Optimization, Bristol-Myers Squibb R&D, Pennington, NJ 08534, USA. Electronic address: hao.lu@bms.com.
authors.forename: Hao
authors.initials: H
authors.lastname: Lu
authors.affiliation: Lead Discovery and Optimization, Bristol-Myers Squibb R&D, Pennington, NJ 08534, USA.
authors.forename: Lisa
authors.initials: L
authors.lastname: Kopcho
authors.affiliation: Disease Sciences and Technology, Biocon Bristol-Myers Squibb R&D Centre, Bangalore, 560099, India.
authors.forename: Kaushik
authors.initials: K
authors.lastname: Ghosh
authors.affiliation: Protein Science, Bristol-Myers Squibb R&D, Princeton, NJ 08648, USA.
authors.forename: Mark
authors.initials: M
authors.lastname: Witmer
authors.affiliation: Discovery Chemistry, Bristol-Myers Squibb R&D, Wallingford, CT 06492, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Parker
authors.affiliation: Disease Sciences and Technology, Biocon Bristol-Myers Squibb R&D Centre, Bangalore, 560099, India.
authors.forename: Sumit
authors.initials: S
authors.lastname: Gupta
authors.affiliation: Disease Sciences and Technology, Biocon Bristol-Myers Squibb R&D Centre, Bangalore, 560099, India.
authors.forename: Marilyn
authors.initials: M
authors.lastname: Paul
authors.affiliation: Disease Sciences and Technology, Biocon Bristol-Myers Squibb R&D Centre, Bangalore, 560099, India.
authors.forename: Prasad
authors.initials: P
authors.lastname: Krishnamurthy
authors.affiliation: Disease Sciences and Technology, Biocon Bristol-Myers Squibb R&D Centre, Bangalore, 560099, India.
authors.forename: Basanth
authors.initials: B
authors.lastname: Laksmaiah
authors.affiliation: Protein Science, Bristol-Myers Squibb R&D, Princeton, NJ 08648, USA.
authors.forename: Dianlin
authors.initials: D
authors.lastname: Xie
authors.affiliation: Protein Science, Bristol-Myers Squibb R&D, Princeton, NJ 08648, USA.
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Tredup
authors.affiliation: Lead Discovery and Optimization, Bristol-Myers Squibb R&D, Pennington, NJ 08534, USA.
authors.forename: Litao
authors.initials: L
authors.lastname: Zhang
authors.affiliation: Lead Discovery and Optimization, Bristol-Myers Squibb R&D, Pennington, NJ 08534, USA. Electronic address: lynn.abell@bms.com.
authors.forename: Lynn M
authors.initials: LM
authors.lastname: Abell
mesh_terms: D001921:Brain / Q000187:drug effects / Q000378:metabolism; D002493:Central Nervous System Diseases / Q000188:drug therapy / Q000201:enzymology; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D057075:Enzyme Assays / Q000379:methods*; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D057166:High-Throughput Screening Assays / Q000379:methods; D006801:Humans; D007736:Kynurenic Acid / Q000378:metabolism; D013058:Mass Spectrometry / Q000379:methods; D013050:Spectrometry, Fluorescence / Q000379:methods; D000637:Transaminases / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D000637:Transaminases; C021481:kynurenine-oxoglutarate transaminase; D007736:Kynurenic Acid
keywords: Fluorescence; High throughput; KATII; Mass spectrometry; RapidFire