Pharmacology and autoradiography of human DP prostanoid receptors using [H-3]-BWA868C, a DP receptor-selective antagonist radioligand

1 A potent and highly selective DP prostanoid receptor antagonist radioliga nd, [H-3]-cyclohexyl-N-BWA868C (3-benzyl-5-(6-carboxyhexyl)-1-(2-cyclohexyl -2-hydroxyethyl-amino) hydantoin, ([H-3]-BWA868C)), has been generated for receptor binding and autoradiographic studies. 2 Specific [H-3]-BWA868C binding to human platelet membranes achieved equil ibrium within 60 min at 23 degreesC and constituted up to 95% of the total binding. The association (K (+ 1)) and dissociation (K (- 1)) rate constant s of binding were 0.758/-0.064 min(-1), mmol and 0.0042+/-0.0002 min(-1), r espectively, yielding dissociation constants (K(D)s) of 5.66+/-0.44 nM (n = 4). 3 Specific [H-3]-BWA868C bound to DP receptors with a high affinity (K-D = 1.45+/-0.01 nM, n = 3) and to a finite, saturable number of binding sites ( B-max = 21.1 +/- 0.6 nmol g(-1) wet weight). 4 DP receptor class prostanoids (e.g. ZK118182, BW245C, BWA868C, PGD,) exhi bited high (nanomolar) affinities for [H-3]-BWA866C binding, while prostano ids selective for EP, FP, IP and TP receptors showed a low (micromolar) aff inity. 5 Specific DP receptor binding sites were autoradiographically localized on the ciliary epithelium/ process, longitudinal and circular ciliary muscles , retinal choroid and iris in human eye sections using [(3H)]-BWA868C. Whil e [H-3]-PGD(2) yielded similar quantitative distribution of DP receptors as [H-3]-BWA868C, the level of non-specific binding observed with [H-3]-PGD(2 ) was significantly greater than that observed with [H-3]-BWA868C. 6 Tt is concluded that [H-3]-BWA868C is a high-affinity and very specific D P receptor radioligand capable of selectively labelling the DP receptor. [H -3]-BWA868C may prove useful for future homogenate-based and autoradiograph ic studies on the DP receptor.