Optimized Administration of the M₄ PAM VU0467154 Demonstrates Broad Efficacy, but Limited Effective Concentrations in Mecp2^+/– Mice

Abstract

Hypofunction of cholinergic circuits and diminished cholinergic tone have been associated with the neurodevelopmental disorder Rett syndrome (RTT). Specifically, deletion of Mecp2 in cholinergic neurons evokes the same social and cognitive phenotypes in mice seen with global Mecp2 knockout, and decreased choline acetyltransferase activity and vesamicol binding have been reported in RTT autopsy samples. Further, we recently identified significant decreases in muscarinic acetylcholine receptor subtype 4 (M 4 ) expression in both the motor cortex and cerebellum of RTT patient autopsies and established proof of concept that an acute dose of the positive allosteric modulator (PAM) VU0467154 (VU154) rescued phenotypes in Mecp2 +/− mice. Here, we expand the assessment of M 4 PAMs in RTT to address clinically relevant questions of tolerance, scope of benefit, dose response, chronic treatment, and mechanism. We show that VU154 has efficacy on anxiety, social preference, cognitive, and respiratory phenotypes in Mecp2 +/− mice; however, the therapeutic range is narrow, with benefits seen at 3 mg/kg concentrations, but not 1 or 10 mg/kg. Further, sociability was diminished in VU154treated Mecp2 +/− mice, suggestive of a potential adverse effect. Compound efficacy on social, cognitive, and respiratory phenotypes was conserved with a 44-day treatment paradigm, with the caveat that breath rate was moderately decreased with chronic treatment in Mecp2 +/+ and Mecp2 +/− mice. VU154 effects on respiratory function correlated with an increase in Gsk3β inhibition in the brainstem. These results identify the core symptom domains where efficacy and adverse effects may present with M 4 administration in RTT model mice and advocate for the continued evaluation as potential RTT therapeutics.

Potentiation of the muscarinic acetylcholine receptor 1 modulates neurophysiological features in a mouse model of Rett syndrome

“…This suggests that chronic dosing of VU846 may induce differential efficacy compared to acute administration; as VU846 has been optimized for use in chronic dosing paradigms, these could form the basis of future studies. We would note, however, that a similar inverted U-shaped dose response in RTT mice was reported in our study of the of M 4 PAM VU0467154 (VU154), which rescued anxiety, social preference, associative memory, and respiratory phenotypes in RTT mice after several weeks of dosing, but benefit was seen only with 3 ​mg/kg but not with 1 or 10 ​mg/kg doses [ 44 , 68 ]. Based on our findings, future studies evaluating compound behavior on both behavior and AEPs with RTT mice using dose-response and chronic dosing studies would be informative in positioning AEPs as a biomarker for muscarinic mechanisms.…”

Activation of M4 muscarinic receptors in the striatum reduces tic‐like behaviours in two distinct murine models of Tourette syndrome

“…The highly selective, competitive muscarinic M 1 receptor antagonist VU0255035 (Abcam, Boston, MA), and the potent selective M 4 receptor antagonist VU6028418 (Bio‐Techne) were suspended in 0.1% Tween80, 0.5% MC in water, and administered PO at 10 ml·kg −1 as in Spock et al (2021) or infused into the DLS. All dose ranges were informed by the available literature on rodent models (Cikowski et al, 2022; Montani et al, 2021; Ono et al, 2012; Sheffler et al, 2009) and established based on pilot dose‐curve investigations on CIN‐d and control mice.…”