Pharmacological profile of GR117289 in vitro: a novel, potent and specific non‐peptide angiotensin AT1 receptor antagonist
Authors: Robertson M.J. , Barnes J.C. , Drew G.M. , Clark K.L. , Marshall F.H. , Michel A. , Middlemiss D. , Ross B.C. , Scopes D. , Dowle M.D. 
Published: July 2012

This paper describes the effects of GR117289 (1‐[[3‐bromo‐2‐[2‐(1H‐tetrazol‐5‐yl)phenyl]‐5‐benzofuranyl]methyl]‐2‐butyl‐4‐chloro‐1H‐imidazole‐5‐carboxylic acid) at angiotensin receptors and binding sites in rabbit aorta, rat liver and bovine cerebellum preparations in vitro.
In rabbit isolated aortic strips, GR117289 (0.3, 1 and 3 nm) caused a concentration‐related, insurmountable suppression of the concentration‐response curve to angiotensin II (AII). When the contact time was increased, a greater degree of antagonism of AII was observed, suggesting that GR117289 is slow to reach equilibrium. A pKB of 9.8 ± 0.1 was calculated for GR117289 after 3 h incubation. GR117289 (1 μm) did not affect contractile responses to phenylephrine or 5‐hydroxytryptamine (5‐HT) in the rabbit aorta.
GR117289 (1 nm) alone caused a marked suppression and a slight rightward displacement of the AII concentration‐response curve. Co‐incubation with the competitive, surmountable AT1 receptor antagonist, losartan (10 nm, 100 nm and 1 μm), resulted in a concentration‐related upward and rightward displacement of the concentration‐response curve to subsequently administered AII. In separate experiments in which preparations were pre‐incubated with GR117289 (1 nm), subsequent addition of losartan (1 μm) for 2, 15 or 45 min caused a further, but similar, rightward displacement of the concentration‐response curve to subsequently administered AII with a time‐dependent increase in the maximum response.
Suppression of AII‐induced contractile responses, caused by superfusion with GR117289 (0.3, 1 or 3 nm) was not reversed by continuously washing the tissues for 3 h; in fact, the potency of GR117289 was slightly enhanced after this period.
In rat liver membranes, GR117289 was a potent competitor with [3H]‐AII for AT1 binding sites (pKi = 8.7 ± 0.1) but in bovine cerebellum membranes, it was a very weak competitor for AT2 binding sites (pKi < 6). Pre‐incubation of rat liver membranes with GR117289 had little effect on its affinity (pKi = 9.1 ± 0.21), but increasing the concentration of bovine serum albumen in the assay buffer from 0.001% to 0.1% w/v decreased affinity (pKi = 7.5 ± 0.1).
In saturation binding experiments in rat liver membranes, GR117289 (12 nm) increased the Kd of [3H]‐AII from 0.28 ± 0.06 nm to 0.37 ± 0.02 nm, and decreased Bmax from 10.0 ± 0.1 to 5.6 ± 0.3 fmol mg−1 tissue. In other experiments, GR117289 (1 μm) did not alter the rate of dissociation of [3H]‐AII from AT1 binding sites, following addition of excess unlabelled AII.
In rabbit aorta vascular smooth muscle membranes, GR117289 competed with [125I]‐Sar1Ile8 AII for binding to AT1 binding sites. In the presence of 0.1% w/v bovine serum albumen, a pIC50 of 7.6 ± 0.1 was calculated. Under the same conditions, but with rat liver membranes, a pIC50 of 7.8 ± 0.1 was determined.
Taken together, these results show that GR117289 is a potent, specific, selective and insurmountable antagonist at angiotensin AT1 receptors. Its profile in the rabbit aorta is consistent with the proposal that GR117289 is a slowly reversible (pseudo‐irreversible) antagonist at these receptors.