Characterization of [125I]-SB-258585 binding to human recombinant and native 5-HT6 receptors in rat, pig and human brain tissue

1 SB-258585 (4-Iodo-N-[4-methoxy-3-(4-methyl-piperazin-1-yl)-phenyl]-benzenesulphonamide) is a high affinity ligand at 5-HT6 receptors. It displays over 100 fold selectivity for the 5-HT6 receptor over all other 5-HT receptors tested so far. SB-258585 has been radiolabelled, to high specific activity, for its characterization as a 5-HT6 receptor selective radioligand. 2 [I-125]-SB-258585 bound, with high affinity, to a single population of receptors in a cell line expressing human recombinant 5-HT6 receptors. Kinetic and saturation binding experiments gave pK(D) values of 9.01+/-0.09 and 9.09+/-0.02, respectively. 3 In membranes derived from rat or pig striatum and human caudate putamen, [I-125]-SB-258585 labelled a single site with high levels (>60%) of specific binding. Saturation analysis revealed pK(D) values of 8.56+/-0.07 for rat, 8.60+/-0.10 for pig and 8.90+/-0.02 for human. B-max values for the tissues ranged from 173+/-23 and 181+/-25 fmol mg(-1) protein in rat and pig striatum, respectively, to 215+/-41 fmol mg(-1) protein in human caudate putamen. 4 The pK(i) rank order of potency for a number of compounds, determined in competition binding assays with [I-125]-SB-258585, at human caudate putamen membranes was: SB-271046 > SB-258585 > SB-214111 > methiothepin > clozapine > 5-Me-OT > 5-HT > Po 04-6790 > mianserin > ritanserin = amitriptyline > 5-CT > mesulergine. Similar profiles were obtained from pig and rat striatal membranes and recombinant 5-HT6 receptors; data from the latter correlated well with [H-3]-LSD binding. 5 Thus, [I-125]-SB-258585 is a high affinity, selective radioligand which can be used to label both recombinant and native 5-HT6 receptors and will facilitate further characterization of this receptor subtype in animal and human tissues.