Crystal structure of the M₅ muscarinic acetylcholine receptor

Abstract

32The human M5 muscarinic acetylcholine receptor (mAChR) has recently emerged as an 33 exciting therapeutic target for treating a range of disorders, including drug addiction. However, 34 a lack of structural information for this receptor subtype has limited further drug development 35 and validation. Here we report a high-resolution crystal structure of the human M5 mAChR 36 bound to the clinically used inverse agonist, tiotropium. This structure allowed for a 37 comparison across all five mAChR family members that revealed important differences in both 38 orthosteric and allosteric sites that could inform the rational design of selective ligands. These 39 structural studies together with chimeric swaps between the extracellular regions of the M2 and 40 M5 mAChR further revealed the structural insight into "kinetic-selectivity", where ligands 41show differential residency times between related family members. Collectively, our study 42 provides important insights into the nature of orthosteric and allosteric ligand interaction across 43 the mAChR family that could be exploited for the design of selective ligands. 44 45 Significance Statement 46The five subtypes of the muscarinic acetylcholine receptors (mAChRs) are expressed 47 throughout the central and peripheral nervous system where they play a vital role in physiology 48 and pathologies. Recently, the M5 mAChR subtype has emerged as an exciting drug target for 49 the treatment of drug addiction. We have determined the atomic structure of the M5 mAChR 50 bound to the clinically used inverse agonist tiotropium. The M5 mAChR structure now allows 51 for a full comparison of all five mAChR subtypes and reveals subtle differences in the 52 extracellular loop (ECL) regions of the receptor that mediate orthosteric and allosteric ligand 53 selectivity. Together these findings open the door for future structure-based design of selective

Activation of M1muscarinic receptors reduce pathology and slow progression of neurodegenerative disease

“…The M1-receptor in particular with its high expression in memory centres and pro-cognitive properties has been proposed as a target for the treatment of memory loss in AD that would avoid the dose-limiting adverse responses associated with current clinically approved cholinesterase inhibitors 23,24 . The challenge has been to develop M1-receptor selective drugs since the orthosteric acetylcholine binding site is highly conserved across the five muscarinic receptor subtypes 20 . As such orthosteric ligands, such as xanomeline, have failed in the clinic as AD therapeutics primarily due to cholinergicadverse responses mediated by peripheral M2-and M3-receptors 21 .…”

“…The barrier to testing this hypothesis in the clinic is the development of agonists that selectively activate the M1-receptor since the orthosteric acetylcholine binding site is nearly identical between the 5 muscarinic receptor subtypes 20 . As such, generation of subtype-selective orthosteric agonists is very challenging 21 .…”

“…As such, generation of subtype-selective orthosteric agonists is very challenging 21 . An alternative approach we, and others, have adopted is to target an allosteric binding pocket on the extracellular surface of the receptor 20 . Agents that bind to this site can act by increasing the sensitivity of the receptor to acetylcholine 22 .…”

Steroids as the novel class of high-affinity allosteric modulators of muscarinic receptors

“…asthma, chronic obstructive pulmonary disease, over-reactive bladder) 2,3 . All five subtypes of muscarinic receptors share high structural homology in the orthosteric binding site that makes their pharmacological targeting via orthosteric binding site extremely difficult [4][5][6][7] . Therefore, in past decades a lot of effort was dedicated to the research of allosteric modulators that bind to the less conserved sites on muscarinic receptors.…”

Muscarinic Acetylcholine Receptor M5 Involved in Spermatogenesis through the Modification of Cell-Cell Junctions

“…M1-M4 have been studied for a long time, however, M5 is the least investigated mAChR subtype and was the last one to be cloned (Bender et al, 2019;Vuckovic et al, 2019). However, mAChR M5 appears to have many important roles which makes it an attractive therapeutic target (Vuckovic et al, 2019;Fujii et al, 2017;Berizzi et al, 2016). mAChRs are present in the male reproductive system; in particular, M1-M4 are found in efferent ductules, epididymides, vas deferens, seminal vesicles, and the prostate (Lucas et al, 2008).…”