title: In vitro pharmacological characterization of novel isoxazolopyridone derivatives as allosteric metabotropic glutamate receptor 7 antagonists.
abstract: Novel isoxazolopyridone derivatives that are metabotropic glutamate receptor (mGluR) 7 antagonists were discovered and pharmacologically characterized. 5-Methyl-3,6-diphenylisoxazolo[4,5-c]pyridin-4(5H)-one (MDIP) was identified by random screening, and 6-(4-methoxyphenyl)-5-methyl-3-pyridin-4-ylisoxazolo[4,5-c]pyridin-4(5H)-one (MMPIP) was produced by chemical modification of MDIP. MDIP and MMPIP inhibited L-(+)-2-amino-4-phosphonobutyric acid (L-AP4)-induced intracellular Ca2+ mobilization in Chinese hamster ovary (CHO) cells coexpressing rat mGluR7 with Galpha(15) (IC50 = 20 and 26 nM). The maximal response in agonist concentration-response curves was reduced in the presence of MMPIP, and its antagonism is reversible. MMPIP did not displace [3H](2S)-2-amino-2-[(1S,2S)-2-carboxycycloprop-1-yl]-3-(xanth-9-yl) propanoic acid (LY341495) bound to mGluR7. These results suggested that these isoxazolopyridone derivatives are allosteric antagonists. In CHO cells expressing rat mGluR7, MDIP and MMPIP inhibited l-AP4-induced inhibition of forskolin-stimulated cAMP accumulation (IC50 = 99 and 220 nM). In CHO cells coexpressing human mGluR7 with Galpha(15), MDIP and MMPIP also inhibited the l-AP4-induced cAMP response. The maximal degree of inhibition by MMPIP was higher than that by MDIP in a cAMP assay. MMPIP was able to antagonize an allosteric agonist, the N,N'-dibenzhydryl-ethane-1,2-diamine dihydrochloride (AMN082)-induced inhibition of cAMP accumulation. In the absence of these agonists, MMPIP caused a further increase in forskolin-stimulated cAMP levels in CHO cells expressing mGluR7, whereas a competitive antagonist, LY341495, did not. This result indicates that MMPIP has an inverse agonistic activity. The intrinsic activity of MMPIP was pertussis toxin-sensitive and mGluR7-dependent. MMPIP at concentrations of at least 1 microM had no significant effect on mGluR1, mGluR2, mGluR3, mGluR4, mGluR5, and mGluR8. MMPIP is the first allosteric mGluR7-selective antagonist that could potentially be useful as a pharmacological tool for elucidating the roles of mGluR7 on central nervous system functions.
authors.affiliation: Tsukuba Research Institute, Banyu Pharmaceutical Co., Ltd., 3 Okubo, Tsukuba, Ibaraki 300-2611, Japan. gentaroh_suzuki@merck.com
authors.forename: Gentaroh
authors.initials: G
authors.lastname: Suzuki
authors.forename: Naohiro
authors.initials: N
authors.lastname: Tsukamoto
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Fushiki
authors.forename: Aki
authors.initials: A
authors.lastname: Kawagishi
authors.forename: Masayuki
authors.initials: M
authors.lastname: Nakamura
authors.forename: Hideki
authors.initials: H
authors.lastname: Kurihara
authors.forename: Morihiro
authors.initials: M
authors.lastname: Mitsuya
authors.forename: Mitsuru
authors.initials: M
authors.lastname: Ohkubo
authors.forename: Hisashi
authors.initials: H
authors.lastname: Ohta
mesh_terms: D000495:Allosteric Site; D000818:Animals; D001667:Binding, Competitive; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism; D003001:Cloning, Molecular; D006224:Cricetinae; D003412:Cricetulus; D000242:Cyclic AMP / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D006801:Humans; D015394:Molecular Structure; D010080:Oxazoles / Q000737:chemistry / Q000494:pharmacology*; D011728:Pyridones / Q000737:chemistry / Q000494:pharmacology*; D034741:RNA, Small Interfering / Q000494:pharmacology; D051381:Rats; D018094:Receptors, Metabotropic Glutamate / Q000037:antagonists & inhibitors* / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors*
chemical_list: D010080:Oxazoles; D011728:Pyridones; D034741:RNA, Small Interfering; D018094:Receptors, Metabotropic Glutamate; D011994:Recombinant Proteins; C102719:metabotropic glutamate receptor 7; C508120:oxazolopyridinone; D000242:Cyclic AMP; D002118:Calcium