Discovery of the First M₅-Selective and CNS Penetrant Negative Allosteric Modulator (NAM) of a Muscarinic Acetylcholine Receptor: (S)-9b-(4-Chlorophenyl)-1-(3,4-difluorobenzoyl)-2,3-dihydro-1H-imidazo[2,1-a]isoindol-5(9bH)-one (ML375)

Abstract

A functional high throughput screen and subsequent multi-dimensional, iterative parallel synthesis effort identified the first muscarinic acetylcholine receptor (mAChR) negative allosteric modulator (NAM) selective for the M5 subtype. ML375 is a highly selective M5 NAM with sub-micromolar potency (human M5 IC50 = 300 nM, rat M5 IC50 = 790 nM, M1–4 IC50 >30 μM), excellent multi-species PK, high CNS penetration, and enantiospecific inhibition.

Molecular Mechanisms of Action of M5 Muscarinic Acetylcholine Receptor Allosteric Modulators

“…Recent screening and medicinal chemistry efforts have resulted in the first subtype-selective small-molecule positive and negative allosteric modulators of muscarinic M 5 mAChRs (Bridges et al, 2010;Gentry et al, 2013Gentry et al, , 2014). ML380 has been described as a PAM of ACh-stimulated calcium mobilization at both human and rat muscarinic M 5 mAChRs, with moderate selectivity versus the M 1 and M 3 mAChR subtypes.…”

“…Radioligand-binding studies also suggested that ML380 could increase the affinity of ACh for the M 5 mAChR . ML375 is a highly subtype-selective NAM of ACh-stimulated calcium mobilization at both human and rat muscarinic M 5 mAChRs, which was shown to be allosteric by its ability to slow [ 3 H]-NMS dissociation from the receptor (Gentry et al, 2013). Despite these data, relatively little is known about the mechanisms of modulation by these allosteric ligands.…”

“…Most recently, screening and medicinal chemistry studies identified the first M 5 mAChR-selective allosteric ligands, including the exemplar PAM, ML380 [1-((1H-indazol-5-yl) sulfonyl)-N-ethyl-N-(2-(trifluoromethyl)benzyl)piperidine-4-carboxamide], and the negative allosteric modulator (NAM), (Gentry et al, 2013 (Fig. 1).…”

Crystal structure of 2-(8a-methyl-5-oxo-hexahydroimidazo [1,2-a]pyridin-1(5H)-yl)-2-oxoethyl acetate, C12H18N2O4

“…They were also used as anticancer agents, antibiotics, antibacterial agents, and so on [4][5][6][7]. Moreover, they also acted as a useful intermediate in the synthesis of compounds with special properties [8,9]. Dichloroacetyl diazabicyclo derivatives were reported as novel herbicide safener protecting crops from injury by chlorine acetamide herbicides, sulfonylurea herbicides, and imidazolinone herbicides since BASF employed 5-dichloroacetyl-3,3,6-trimethyl-9-oxo-1,5-diazabicyclo[4.3.0] nonane as safeners which increased the toleration of crops to acetanilides herbicides [10][11][12].…”

Further optimization of the M5 NAM MLPCN probe ML375: Tactics and challenges

“…20 A limited chemical optimization effort afforded ML375 ( 2 ), the first M 5 -selective NAM with favorable CNS exposure (brain:plasma K p = 1.8), moderate PK, high plasma protein binding (rat f u = 0.029, human f u = 0.013, rat brain f u = 0.003) and enantiospecific activity (only the ( S )-enantiomer of the 9b p -Cl phenyl wasactive). 20 Despite a major advance in the field, due to weak potency at rat M 5 , coupled with high plasma protein and brain homogenate binding, ML375 lacked the requisite free drug exposure to serve as an in vivo tool compound. 20 In this Letter, we report on the continued optimization of our first-in-class M 5 NAM, and detail key tactics and noteworthy challenges en route to an M 5 NAM in vivo probe.…”