Annulated heterocyclic bioisosteres of norarecoline. Synthesis and molecular pharmacology at five recombinant human muscarinic acetylcholine receptors

Article abstract of DOI:10.1021/jm00012a019

A series of O-alkylated analogs of 5,6,7,8-tetrahydro-4H-isoxazolo[4,5- c]azepin-3-ol (THAO) were synthesized and characterized as ligands for muscarinic acetylcholine receptors (mAChRs). O-Methyl-THAO (4a), O-ethyl- THAO (4b), O-isopropyl-THAO (4c), and O-propargyl-THAO (4d) were shown to be potent inhibitors of the binding of tritiated quinuclidinyl benzilate (QNB), pirenzepine (PZ), and oxotremorine-M (Oxo-M) to tissue membrane preparations. In the [³H]-Oxo-M binding assay, receptor affinities in the low nanomolar range were measured for 4a (IC₅₀ = 0.010 μM), 4b (IC₅₀ = 0.003 μM), 4c (IC₅₀ = 0.011 μM), and 4d (IC₅₀ = 0.0008 μM). Pharmacological effects (EC₅₀ or K(i) values) and intrinsic activities (per cent of maximal carbachol responses) were determined using five recombinant human mAChRs (m1- m5) and the functional assay, receptor selection and amplification technology (R-SAT). Compound 4c antagonized carbachol-induced responses at m1, m3, and m5. With the exception of 4b, which was an antagonist at m5, 4a,b,d showed partial agonism at m1-m5 with very similar subtype selectivity (m2 > m4 > ml ≥ m3 > m5). Agonist index values for 4a-d, which were calculated from [³H]QNB (brain) and [³H]Oxo-M (brain) binding data, were shown to be predictive of pharmacologically determined intrinsic activities at m1-m5, the same rank order of intrinsic activity being observed at all five mAChRs (4a > 4d > 4b > 4c). It is concluded that within this class of high-affinity mAChR (m1-m5) ligands, containing secondary amino groups, minor changes of the bioisosteric ester alkyl groups have marked effects on potency and, in particular, intrinsic activity.