title: Pharmacological characterization and radioligand binding properties of a high-affinity, nonpeptide, bradykinin B1 receptor antagonist.
abstract: Compound A (N-[2-[4-(4,5-dihydro-1H-imidazol-2-yl)phenyl]ethyl]-2-[(2R)-1-(2-napthylsulfonyl)-3-oxo-1,2,3,4-tetrahydroquinoxalin-2-yl]acetamide) is a member of a new class of aryl sulfonamide dihydroquinoxalinone bradykinin B1 receptor antagonists that should be useful pharmacological tools. Here we report on some of the pharmacological properties of compound A as well as the characterization of [35S]compound A as the first nonpeptide bradykinin B1 receptor radioligand. Compound A inhibited tritiated peptide ligand binding to the cloned human, rabbit, dog, and rat bradykinin B1 receptors expressed in CHO cells with Ki values of 0.016, 0.050, 0.56, and 29 nM, respectively. It was inactive at 10 microM in binding assays with the cloned human bradykinin B2 receptor. In functional antagonist assays with the cloned bradykinin B1 receptors, compound A inhibited agonist-induced signaling with activities consistent with the competition binding results, but had no antagonist activity at the bradykinin B2 receptor. Compound A was also found to be a potent antagonist in a rabbit aorta tissue bath preparation and to effectively block des-Arg9 bradykinin depressor responses in lipopolysaccharide-treated rabbit following intravenous administration. The binding of [35S]compound A was evaluated with the cloned bradykinin B1 receptors. In assays with human, rabbit, and dog receptors, [35S]compound A labeled a single site with Kd values of 0.012, 0.064, and 0.37 nM, respectively, and with binding site densities equivalent to those obtained using the conventional tritiated peptide ligands. Binding assays with the cloned rat bradykinin B1 receptor were not successful, presumably due to the low affinity of the ligand for this species receptor. There was no specific binding of the ligand detected in CHO cells expressing the human bradykinin B2 receptor. In assays with the cloned human bradykinin B1 receptor, the pharmacologies of the binding of [35S]compound A and [3H][Leu9]des-Arg10-kallidin were the same. The high signal-to-noise ratio obtained with [35S]compound A will allow this ligand to be a very useful tool for future investigations of the bradykinin B1 receptor.
authors.affiliation: Department of Molecular Neurology, Merck Research Laboratories, WP46-300, West Point, PA 19486, USA. rick_ransom@merck.com
authors.forename: Richard W
authors.initials: RW
authors.lastname: Ransom
authors.forename: Charles M
authors.initials: CM
authors.lastname: Harrell
authors.forename: Duane R
authors.initials: DR
authors.lastname: Reiss
authors.forename: Kathryn L
authors.initials: KL
authors.lastname: Murphy
authors.forename: Raymond S L
authors.initials: RS
authors.lastname: Chang
authors.forename: J Fred
authors.initials: JF
authors.lastname: Hess
authors.forename: Patricia J
authors.initials: PJ
authors.lastname: Miller
authors.forename: Stacey S
authors.initials: SS
authors.lastname: O'Malley
authors.forename: Pat J
authors.initials: PJ
authors.lastname: Hey
authors.forename: Priya
authors.initials: P
authors.lastname: Kunapuli
authors.forename: Dai-Shi
authors.initials: DS
authors.lastname: Su
authors.forename: M Kristine
authors.initials: MK
authors.lastname: Markowitz
authors.forename: Michael A
authors.initials: MA
authors.lastname: Wallace
authors.forename: Conrad E
authors.initials: CE
authors.lastname: Raab
authors.forename: Allen N
authors.initials: AN
authors.lastname: Jones
authors.forename: Dennis C
authors.initials: DC
authors.lastname: Dean
authors.forename: Douglas J
authors.initials: DJ
authors.lastname: Pettibone
authors.forename: Roger M
authors.initials: RM
authors.lastname: Freidinger
authors.forename: Mark G
authors.initials: MG
authors.lastname: Bock
mesh_terms: D000818:Animals; D001013:Aorta, Thoracic / Q000187:drug effects / Q000502:physiology; D001667:Binding, Competitive / Q000187:drug effects; D001794:Blood Pressure / Q000187:drug effects; D065169:Bradykinin B1 Receptor Antagonists; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D004285:Dogs; D004305:Dose-Response Relationship, Drug; D006801:Humans; D007093:Imidazoles / Q000378:metabolism / Q000494:pharmacology; D066298:In Vitro Techniques; D007609:Kallidin / Q000031:analogs & derivatives* / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D008297:Male; D011810:Quinoxalines / Q000378:metabolism / Q000494:pharmacology; D011817:Rabbits; D011869:Radioligand Assay; D051381:Rats; D043783:Receptor, Bradykinin B1 / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D014316:Tritium; D014661:Vasoconstriction / Q000187:drug effects
chemical_list: D065169:Bradykinin B1 Receptor Antagonists; D007093:Imidazoles; D008070:Lipopolysaccharides; C496286:N-(2-(4-(4,5-dihydro-1H-imidazol-2-yl)phenyl)ethyl)-2-((2R)-1-(2-napthylsulfonyl)-3-oxo-1,2,3,4-tetrahydroquinoxalin-2-yl)acetamide; D011810:Quinoxalines; D043783:Receptor, Bradykinin B1; C082229:bradykinin, Lys-Leu(8)-desArg(9)-; D014316:Tritium; D007609:Kallidin; C052787:kallidin, des-Arg(10)-