Pharmacological characterization and radioligand binding properties of a high-affinity, nonpeptide, bradykinin B 1 receptor antagonist

• Published in: European journal of pharmacology Vol. 499; no. 1; pp. 77 - 84
• Main Authors: Ransom, Richard W., Harrell, Charles M., Reiss, Duane R., Murphy, Kathryn L., Chang, Raymond S.L., Hess, J. Fred, Miller, Patricia J., O'Malley, Stacey S., Hey, Pat J., Kunapuli, Priya, Su, Dai-Shi, Markowitz, M. Kristine, Wallace, Michael A., Raab, Conrad E., Jones, Allen N., Dean, Dennis C., Pettibone, Douglas J., Freidinger, Roger M., Bock, Mark G.
• Format: Journal Article
• Language: English
• Published: Elsevier B.V 19.09.2004
• Subjects: Antagonist; Bradykinin B 1 receptor; Radioligand; Radioligand; Bradykinin B 1 receptor; Antagonist
• ISSN: 0014-2999; 1879-0712
• DOI: 10.1016/j.ejphar.2004.07.104

Compound A ( N-{2-[4-(4,5-dihydro-1 H-imidazol-2-yl)phenyl]ethyl}-2-[(2 R)-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 B 1 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 B 1 receptor radioligand. Compound A inhibited tritiated peptide ligand binding to the cloned human, rabbit, dog, and rat bradykinin B 1 receptors expressed in CHO cells with K i values of 0.016, 0.050, 0.56, and 29 nM, respectively. It was inactive at 10 μM in binding assays with the cloned human bradykinin B 2 receptor. In functional antagonist assays with the cloned bradykinin B 1 receptors, compound A inhibited agonist-induced signaling with activities consistent with the competition binding results, but had no antagonist activity at the bradykinin B 2 receptor. Compound A was also found to be a potent antagonist in a rabbit aorta tissue bath preparation and to effectively block des-Arg 9 bradykinin depressor responses in lipopolysaccharide-treated rabbit following intravenous administration. The binding of [ 35S]compound A was evaluated with the cloned bradykinin B 1 receptors. In assays with human, rabbit, and dog receptors, [ 35S]compound A labeled a single site with K d 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 B 1 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 B 2 receptor. In assays with the cloned human bradykinin B 1 receptor, the pharmacologies of the binding of [ 35S]compound A and [ 3H][Leu 9]des-Arg 10-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 B 1 receptor.