Affinity, potency and efficacy of tramadol and its metabolites at the cloned human µ-opioid receptor

Abstract.

The present study was conducted to characterise the centrally active analgesic drug tramadol hydrochloride [(1RS,2RS)-2-[(dimethyl-amino)-methyl]-1-(3-methoxyphenyl)-cyclohexanol hydrochloride] and its metabolites M1, M2, M3, M4 and M5 at the cloned human µ-opioid receptor. Membranes from stably transfected Chinese hamster ovary (CHO) cells were used to determine the four parameters of the ligand-receptor interaction: the affinity of (±)-tramadol and its metabolites was determined by competitive inhibition of [³H]naloxone binding under high and low salt conditions. The agonist-induced stimulation of [³⁵S]GTPγS binding permits the measurement of potency (EC₅₀), efficacy (E _max = maximal stimulation) and relative intrinsic efficacy (effect as a function of receptor occupation). The metabolite (+)-M1 showed the highest affinity (K _i=3.4 nM) to the human µ-opioid receptor, followed by (±)-M5 (K _i=100 nM), (-)-M1 (K _i=240 nM) and (±)-tramadol (K _i=2.4 µM). The [³⁵S]GTPγS binding assay revealed an agonistic activity for the metabolites (+)-M1, (-)-M1 and (±)-M5 with the following rank order of intrinsic efficacy: (+)-M1>(±)-M5>(-)-M1. The metabolites (±)-M2, (±)-M3 and (±)-M4 displayed only weak affinity (K _i>10 µM) and had no stimulatory effect on GTPγS binding. These data indicate that the metabolite (+)-M1 is responsible for the µ-opioid-derived analgesic effect.