Nonpeptidic urotensin-II receptor antagonists I: In vitro pharmacological characterization of SB-706375

SB-706375 potently inhibited [ ¹²⁵I]hU-II binding to both mammalian recombinant and 'native' UT receptors (K _i 4.7 ± 1.5 to 20.7 ± 3.6 nM at rodent, feline and primate recombinant UT receptors and K _i 5.4 ± 0.4 nM at the endogenous UT receptor in SJRH30 cells). Prior exposure to SB-706375 (1 μM, 30 min) did not alter [ ¹²⁵I]hU-II binding affinity or density in recombinant cells (K _D 3.1 ± 0.4 vs 5.8 ± 0.9 nM and B _max 3.1 ± 1.0 vs 2.8 ± 0.8 pmol mg ^-1) consistent with a reversible mode of action. The novel, nonpeptidic radioligand [ ³H]SB-657510, a close analogue of SB-706375, bound to the monkey UT receptor (K _D 2.6 ± 0.4 nM, B _max 0.86 ± 0.12 pmol mg ^-1) in a manner that was inhibited by both U-II isopeptides and SB-706375 (K _i 4.6 ± 1.4 to 17.6 ± 5.4 nM) consistent with the sulphonamides and native U-II ligands sharing a common UT receptor binding domain. SB-706375 was a potent, competitive hU-II antagonist across species with pK _b 7.29-8.00 in HEK293-UT receptor cells (inhibition of [Ca ²⁺] _i-mobilization) and pK _b 7.47 in rat isolated aorta (inhibition of contraction). SB-706375 also reversed tone established in the rat aorta by prior exposure to hU-II (K _app ∼ 20 nM). SB-706375 was a selective U-II antagonist with ≥ 100-fold selectivity for the human UT receptor compared to 86 distinct receptors, ion channels, enzymes, transporters and nuclear hormones (K _i/IC ₅₀ > 1 μM). Accordingly, the contractile responses induced in isolated aortae by KCl, phenylephrine, angiotensin II and endothelin-1 were unaltered by SB-706375 (1 μM). In summary, SB-706375 is a high-affinity, surmountable, reversible and selective nonpeptide UT receptor antagonist with cross-species activity that will assist in delineating the pathophysiological actions of U-II in mammals.