AN APPROACH TO DISCOVER NOVEL MUSCARINIC M1 RECEPTOR-POSITIVE ALLOSTERIC MODULATORS WITH POTENT COGNITIVE IMPROVEMENT AND MINIMIZED GASTROINTESTINAL DYSFUNCTION

Abstract

Activation of the muscarinic M1 receptor (M1R) is a promising approach to improve cognitive impairment in Alzheimer's disease (AD). However, we found that an M1 selective positive allosteric modulator (PAM) induced severe diarrhea through M1R activation, by using wild-type and M1R knock-out (KO) mice. In this study, we established a drug screening strategy to discover novel M1 PAMs with potent cognitive improvement and minimized gastrointestinal dysfunction. PAM parameters of M1 PAMs were assessed by using in vitro binding and functional analysis in human M1R expressing Chinese hamster ovary (CHO-K1) cells. Effects of M1 PAMs on contraction of mouse ileum were assessed by using in vitro Magnus assay. Y-maze task was used to evaluate effects of M1 PAMs on cognitive function in scopolamine-treated mice. Seven M1 PAMs with ≥ 100-fold selectivity over other muscarinic receptor subtypes were used. Their PAM parameters such as PAM activity (the inflection point-value), cooperativity value (α-value), and efficacy value (β-value) were assessed by the in vitro assay. Evaluation of these M1 PAMs in the Magnus assay revealed that there was a strong and positive correlation between % ileum contraction at 1 μM and their cooperativity value (log α), a PAM parameter associated with the effect of allosteric modulation on the affinity of the orthosteric site. Next, we characterized in vivo profiles of two M1 PAMs: compound A (log α = 1.18) and compound B (log α = 3.30). Compound A at 30 mg/kg significantly improved scopolamine-induced cognitive deficits in Y-maze without prominent signs of diarrhea in mice. In contrast, compound B at 10 mg/kg showed both significant amelioration of scopolamine-induced cognitive deficits in Y-maze and severe diarrhea. Fine adjustment of the α-value could be a key to discover M1 PAMs with potent cognitive improvement and lower risk of diarrhea. The current study provides the original screening strategy to identify safer M1 PAMs for the treatment of cognitive deficits related to AD. Based on this strategy, we have identified TAK-071. TAK-071 is currently in clinical development (ClinicalTrials.gov Identifier: NCT02769065).