SYNTHESIS, MOLECULAR MODELING STUDIES, AND MUSCARINIC RECEPTOR ACTIVITY OF AZAPROPHEN ANALOGS

Synthesis, radioligand binding, and pharmacologic activities of a series of muscarinic receptor ligands including and related to azaprophen (6-methyl-6-azabicyclo[3.2.1]octan-3-alpha-ol 2,2-diphenylpropionate, 1) have been measured to determine activity and selectivity for muscarinic receptor subtypes. Pharmacologic affinities of antagonists were determined as pA2 values for antagonism of methacholine-induced tension responses in guinea pig ileum. Binding affinities were measured by competition against [H-3]QNB binding in guinea pig ileum, rat heart and brain, and m1- or m3-transfected Chinese hamster ovary (CHO) cells. The efficacies of muscarinic agonists in brain were determined by the ratio of binding affinities against [H-3]QNB or [H-3]NMS and [H-3]oxotremorine-M ([H-3]Oxo-M). Nine muscarinic antagonists, including azaprophen, did not discriminate significantly between the subtypes of muscarinic receptors. K(I) values for receptor binding for azaprophen (1) were between 8.81 x 10(-11) and 4.72 x 10(-10) M in ileum, heart, brain, and m1- or m3-transfected CHO cells. The alpha- and beta-benzilate esters 5 and 6 are as potent as azaprophen, and diphenylacetate esters 3 and 4 and N-(6)-benzyl alpha-isomer 7 are less potent than azaprophen. Significant stereoselectivity was exhibited with (+)-azaprophen being approximately 200 times more potent than the (-)-enantiomers and the 3-beta-ol isomer 2 being ca. 50 times less potent than azaprophen in all systems. A molecular modeling-molecular mechanics study was conducted to account for the difference. Putative muscarinic agonists (analogues and isomers of 6-methyl-6-azabicyclo[3.2.1]octan-3-ol acetate) did not discriminate muscarinic receptor subtypes with K(I) values between 2.77 x 10(-6) and 4.33 X 10(-5)M without significant stereoselectivity in the systems examined. The most active analogue was (1R,3R,5S)-6-[1(R)-phenylethyl]-6-azabicyclo[3.2.1]octan-3-alpha-ol acetate. However, efficacies of these putative agonists were in general very low.