Tl. Davis et Na. Sharif, Pharmacological characterization of [H-3]-prostaglandin E-2 binding to thecloned human EP4 prostanoid receptor, BR J PHARM, 130(8), 2000, pp. 1919-1926
1 Prostaglandin (PG) E-2 (PGE(2)) is a potent prostanoid derived from arach
idonic which can interact with EP1, EP2, EP3 and EP4 prostanoid receptor su
btypes.
2 Recombinant human EP4 receptors expressed in human embryonic kidney (HEK-
293) cells were evaluated for their binding characteristics using [H-3]-PGE
(2) and a broad panel of natural and synthetic prostanoids in order to defi
ne their pharmacological properties.
3 [H-3]-PGE(2) binding was optimal in 2-[N-Morpholino]ethanesulphonic acid
(MES) buffer (pH 6.0) yielding 98 +/- 0.7% specific binding. The receptor d
isplayed high affinity (K-d = 0.72 +/- 0.12 nM; n = 3) for [H-3]-PGE(2) and
interacted with a saturable number of binding sites (B-max = 6.21 +/- 0.84
pmol mg(-1) protein).
4 In competition studies, PGE(2) (K-i = 0.75 +/- 0.03 nM; n = 12) and PGE(1
) (K-i = 1.45 +/- 0.24 nM; n = 3) displayed high affinities, as did two der
ivatives of PGE(1), namely 11-deoxy-PGE(1) (K-i = 1.36 +/- 0.34 nM) and 13,
14-dihydro-PGE(1) (K-i = 3.07 +/- 0.29 nM).
5 Interestingly, synthetic DP receptor-specific agonists such as BW245C (K-
i = 64.7 +/- 1.0 nM; n = 3) and ZK118182 (K-i = 425 +/- 42 nM; n = 4), and
the purported EP3 receptor-specific ligand enprostil (K-i = 43.1 +/- 4.4 nM
), also displayed high affinity for the EP4 receptor.
6 Two known EP4 receptor antagonists were weak inhibitors of [H-3]-PGE(2) b
inding akin to their known functional potencies, thus: AH23848 (K-i = 2690
+/- 232 nM); AH22921 (K-i = 31,800 +/- 4090 nM).
7 These studies have provided a detailed pharmacological characterization o
f the recombinant human EP4 receptor expressed in HEK-293 cells.