M4 PAM 23i(Monash University)

The M₄ muscarinic acetylcholine receptor (mAChR) is a biological target for neurocognitive disorders. Compound 1 is an ago-PAM for the M₄ mAChR. Herein, we report the design, synthesis, and evaluation of novel putative M₄ mAChR PAMs based on 1. These analogs were screened and then fully characterized in two functional assays (G_oB protein activation and CAMYEL activation) to quantify their allosteric and ago-PAM properties against ACh. A selection of 7 M₄ PAMs were assessed for their ability to modulate ACh-mediated β-arrestin recruitment and revealed 4 distinct clusters of M₄ PAM activity: (1) analogs similar to 1 (24d), (2) analogs demonstrating only allosteric agonism (23d), (3) analogs with increased allosteric properties in CAMYEL activation (23b/23f and 24a/24b), and (4) analogs with a biased modulatory effect toward β-arrestin recruitment (23i). These novel M₄ chemical tools disclose discrete molecular determinants, allowing further interrogation of the therapeutic roles of cAMP and β-arrestin pathways in neurocognitive disorders.