Law_2026_J.Mol.Struct__145611

Alzheimers disease (AD) is a progressive neurodegenerative disorder marked by cognitive and non-cognitive decline. Cholinesterase inhibitors remain among the few available treatments, acting by preventing the hydrolysis of acetylcholine to alleviate cholinergic deficits. As butyrylcholinesterase (BChE) activity progressively increases in advanced AD while acetylcholinesterase activity declines, selective BChE inhibition has emerged as a promising therapeutic approach. Benzimidazole is a versatile pharmacophore with diverse biological activities, including cholinesterase inhibition. This study aimed to develop more potent benzimidazole-based cholinesterase inhibitors by modifying the 1- and 2-positions of the benzimidazole core. Among the synthesized compounds, 5IIa exhibited the most potent selective BChE inhibition at a low micromolar level (IC50=0.4 microM). Human BChE (hBChE) was co-crystallized with 5IIa to resolve the structure of the proteinligand complex by X-ray crystallography. Owing to its labile nature, 5IIa was subsequently modified to yield a series of derivatives (5IIa17), which demonstrated potent and selective inhibition of equine BChE (eqBChE), with IC50 values ranging from 1.07 to 7.86 microM. Lead compound 5IIa6 inhibited hBChE with an IC50 of 5.9 microM and was identified as a mixed-mode inhibitor based on kinetic studies. 5IIa6 was also predicted to have high blood-brain barrier permeability in an in vitro model, while cell viability assay against neuroblastoma and microglial cells indicated no significant toxicity. Taken together, these findings identify 5IIa6 as a promising and selective BChE inhibitor that warrants further investigation as a potential therapeutic agent for AD