title: Identification and characterization of novel small-molecule protease-activated receptor 2 agonists.
abstract: We report the first small-molecule protease-activated receptor (PAR) 2 agonists, AC-55541 [N-[[1-(3-bromo-phenyl)-eth-(E)-ylidene-hydrazinocarbonyl]-(4-oxo-3,4-dihydro-phthalazin-1-yl)-methyl]-benzamide] and AC-264613 [2-oxo-4-phenylpyrrolidine-3-carboxylic acid [1-(3-bromo-phenyl)-(E/Z)-ethylidene]-hydrazide], each representing a distinct chemical series. AC-55541 and AC-264613 each activated PAR2 signaling in cellular proliferation assays, phosphatidylinositol hydrolysis assays, and Ca(2+) mobilization assays, with potencies ranging from 200 to 1000 nM for AC-55541 and 30 to 100 nM for AC-264613. In comparison, the PAR2-activating peptide 2-furoyl-LIGRLO-NH(2) had similar potency, whereas SLIGRL-NH(2) was 30 to 300 times less potent. Neither AC-55541 nor AC-264613 had activity at any of the other PAR receptor subtypes, nor did they have any significant affinity for over 30 other molecular targets involved in nociception. Visualization of EYFP-tagged PAR2 receptors showed that each compound stimulated internalization of PAR2 receptors. AC-55541 and AC-264613 were well absorbed when administered intraperitoneally to rats, each reaching micromolar peak plasma concentrations. AC-55541 and AC-264613 were each stable to metabolism by liver microsomes and maintained sustained exposure in rats, with elimination half-lives of 6.1 and 2.5 h, respectively. Intrapaw administration of AC-55541 or AC-264613 elicited robust and persistent thermal hyperalgesia and edema. Coadministration of either a tachykinin 1 (neurokinin 1) receptor antagonist or a transient receptor potential vanilloid (TRPV) 1 antagonist completely blocked these effects. Systemic administration of either AC-55541 or AC-264613 produced a similar degree of hyperalgesia as was observed when the compounds were administered locally. These compounds represent novel small-molecule PAR2 agonists that will be useful in probing the physiological functions of PAR2 receptors.
authors.affiliation: ACADIA Pharmaceuticals, Inc., San Diego, CA 92121, USA.
authors.forename: Luis R
authors.initials: LR
authors.lastname: Gardell
authors.forename: Jian-Nong
authors.initials: JN
authors.lastname: Ma
authors.forename: Jimmi Gerner
authors.initials: JG
authors.lastname: Seitzberg
authors.forename: Anne E
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authors.lastname: Knapp
authors.forename: Hans H
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authors.lastname: Schiffer
authors.forename: Ali
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authors.lastname: Tabatabaei
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authors.lastname: Davis
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authors.forename: Bryan
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authors.forename: Kenneth K
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authors.lastname: Wong
authors.forename: Birgitte
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authors.forename: Yan
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authors.forename: Jelveh
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authors.lastname: Lameh
authors.forename: Kara
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authors.lastname: Schmelzer
authors.forename: Roger
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authors.initials: ES
authors.lastname: Burstein