Characterisation of [3H]-darifenacin as a novel radioligand for the study of muscarinic M3 receptors.

Darifenacin, (S)-2-[1-[2,3-dihydrobenzofuran-5-yl]-3- pyrrolidinyl]-2,2-diphenylacetamide, is a novel muscarinic M3 antagonist. In this study we have compared the binding of [3H]-darifenacin to the five cloned human muscarinic receptors (m1-m5) expressed in CHO cells. [3H]-darifenacin binds with 6 fold higher affinity to m3 (KD = 0.33 nmol/l) over m1 (KD = 1.6 nmol/l) receptors. There was no specific binding of [3H]-darifenacin to m2 receptors and specific binding to m4 and m5 receptors was insufficient to determine a KD. Binding of [3H]-darifenacin to m1 and m3 was displaced by atropine (m1 pKi = 9.36, m3 pKi = 9.4), 4-DAMP (m1 pKi = 9.04, m3 pKi = 9.19), pirenzepine (m1 pKi = 8.63, m3 pKi = 6.85), methoctramine (m1 pKi = 7.28, m3 pKi = 6.63), and darifenacin (m1 pKi = 8.36, m3 pKi = 9.14), demonstrating that [3H]-darifenacin represents the first selective m3 radioligand.