Investigation of the Interaction of a Putative Allosteric Modulator, N-(2,3-Diphenyl-1,2,4-thiadiazole-5-(2H)-ylidene) Methanamine Hydrobromide (SCH-202676), with M1 Muscarinic Acetylcholine Receptors

Alfred Lanzafame, Arthur Christopoulos

Research output: Contribution to journalArticleResearchpeer-review

46 Citations (Scopus)

Abstract

The interaction between a novel G protein-coupled receptor modulator, N-(2,3-diphenyl-1,2,4-thiadiazole-5-(2H)-ylidene) methanamine hydrobromide (SCH-202676), and the M1 muscarinic acetylcholine receptor (mAChR) was investigated. In contrast to the prototypical mAChR allosteric modulator, heptane 1,7-bis-(dimethyl-3′-phthalimidopropyl)-ammonium bromide (C 7/3-phth), SCH-202676 had no effect on the dissociation kinetics of [3H]N-methylscopolamine ([3H]NMS) at M1 mAChRs stably expressed in Chinese hamster ovary (CHO) cell membranes. However, SCH-202676 completely inhibited the binding of [3HH]NMS in membrane preparations, with a Hill slope significantly greater than unity, indicative of positive cooperativity in the binding of the inhibitor. Moreover, SCH-202676 caused dextral shifts of the [3H]NMS saturation binding curve that were greater than expected for a competitive interaction. The addition of C 7/3-phth (100 μM) had no significant effect on the inhibitory potency of SCH-202676. In contrast to the findings in cell membranes, the interaction between SCH-202676 and [3H]NMS in intact M1 CHO cells yielded saturation and inhibition isotherms that were compatible with the predictions for a competitive interaction. Intact cell assays of acetylcholine-mediated phosphoinositide hydrolysis in the absence or presence of SCH-202676 revealed a mixed competitive/noncompetitive mode of interaction that was dependent on the concentration of SCH-202676. These data reveal that the nature of the interaction between SCH-202676 and the M1 mAChR is dependent on whether it is studied using intact versus broken cell preparations. It is proposed that SCH-202676 uses a dual mode of ligand-receptor interaction involving both extra- and intracellular attachment points on the M1 mAChR that are distinct from the allosteric binding site recognized by prototypical mAChR modulators such as C7/3-phth.

Original languageEnglish
Pages (from-to)830-837
Number of pages8
JournalJournal of Pharmacology and Experimental Therapeutics
Volume308
Issue number3
DOIs
Publication statusPublished - 1 Mar 2004
Externally publishedYes

Cite this