Design, synthesis and evaluation of novel 2-phenyl-3-(1H-pyrazol-4-yl)pyridine positive allosteric modulators for the M4 mAChR

Translation of muscarinic acetylcholine receptor (mAChR) agonists into clinically used therapeutic agents has been difficult due to their poor subtype selectivity. M4 mAChR subtype-selective positive allosteric modulators (PAMs) may provide better therapeutic outcomes, hence investigating their detailed pharmacological properties is crucial to advancing them into the clinic. Herein, we report the synthesis and comprehensive pharmacological evaluation of M4 mAChR PAMs structurally related to 1e, Me–C-c, [11C]MK-6884 and [18F]12. Our results show that small structural changes to the PAMs can result in pronounced differences to baseline, potency (pEC50) and maximum effect (Emax) measures in cAMP assays when compared to the endogenous ligand acetylcholine (ACh) without the addition of the PAMs. Eight selected PAMs were further assessed to determine their binding affinity and potential signalling bias profile between cAMP and β-arrestin 2 recruitment. These rigorous analyses resulted in the discovery of the novel PAMs, 6k and 6l, which exhibit improved allosteric properties compared to the lead compound, and probative in vivo exposure studies in mice confirmed that they maintain the ability to cross the blood-brain barrier, making them more suitable for future preclinical assessment. [Display omitted] •The M mAChR a promising target for the therapy of neurological diseases such as schizophrenia and Alzheimer's disease.•M4 mAChR PAMs with different allosteric profiles in terms of affinity, co-operativity and allosteric agonism were identified.•Compounds 6k and 6l were found to possess biased signalling profiles between cAMP and β-arrestin 2 recruitment.•Compounds 1e, 6k and 6l were able to cross the blood brain barrier and distribute into the brain parenchyma of mice.