Investigation of ONO‐AE1‐259, A Selective Prostanoid EP 2 ‐Receptor Agonist, on the Guinea‐Pig Isolated Saphenous Vein

Abstract

There are no selective agonists for the prostanoid EP4-receptor, and the ones available for the EP2-receptor are either of low potency (e.g. AH 13205) or are C1-methyl esters subject to variable de-esterification within tissues (e.g. butaprost). EP4 vasorelaxant systems are often highly sensitive, with IC50 values for PGE2 as low as 0.1 nM, whereas EP2 systems are usually some 30–100 times less sensitive. Determining whether EP2-receptors co-exist with EP4-receptors is therefore quite difficult. Some information can be gained using the EP4 antagonist AH 23848, but this blocker has a low affinity (pA2= 5.0–5.4), making large dose ratios difficult to obtain. We have recently investigated the utility of a new EP2 agonist, ONO-AE1-259 in this situation. ONO-AE1-259 (free acid) is claimed to be highly selective on the basis of radioligand competition and second messenger measurements on cloned mouse receptors; its EC50 for cyclic AMP production = 1.8 nM for EP2 and >10 μM for EP4-receptors (S. Narumiya; 11th International Conference on Prostaglandins, Florence, 2000). Rings of guinea-pig lateral saphenous vein were set up in a 4-channel Mulvaney myograph system (bath volume = 5 ml) in Krebs-Henseleit solution containing 1 μM indomethacin and 0.2 μM GR 32191 (TP antagonist). Tone was generated with 3 μM phenylephrine and agonists were added cumulatively. PGE2 induced ∼95% relaxation at 5 nM and had an EC50 of about 0.1 nM. ONO-AE1-259 consistently showed small relaxant effects at 1 nM, but its log concentration-response curve was shallower than that of PGE2 and only about 85% relaxation was found at 5 μM (IC50= 20 nM relative to its own maximum). The vein preparation did not contact to sulprostone, and therefore is unlikely to contain opposing EP1 and/or EP3 systems. In the presence of 30 μM AH 23848, the log concentration-response curve for PGE2 was displaced to the right in a parallel manner (dose ratio = 7.6, pA2= 5.3). However, the ONO-AE1-259 curve was only slightly displaced to the right by AH 23848 (dose ratio = 1.6), while at the same time maximum relaxation was reduced by 15%. ONO-AE1-259 was about 25 times more potent than butaprost. It is possible therefore that ONO-AE1-259 is selectively activating a relaxant EP2-receptor in the guinea-pig saphenous vein. The saphenous vein preparation is also highly sensitive to the prostacyclin analogues AFP-07 (IC50= 0.65 nM) and cicaprost (IC50= 7 nM). We are investigating whether these analogues activate IP and/or EP4 receptors, in the light of our recent finding that they are potent and moderately potent EP4 agonists respectively on both piglet and rabbit saphenous veins (Jones & Chan, 2001, Br. J. Pharmacol., in press).