Design, synthesis, and biological evaluation of novel benzylidene-N-(3,4 dichlorophenyl) hydrazine-1-carbothioamide derivatives as new acetylcholinesterase inhibitors

Alzheimer’s disease (AD) is a continuous worsening condition of the nervous system in which brain cells degenerate over time, leading to impairments in memory and thinking, and it is linked to a decrease in acetylcholine concentrations in the brain, a neurotransmitter crucial for memory and learning. Targeting acetylcholinesterase (AChE) inhibition remains a fundamental treatment strategy. The goal of this study was to design and synthesize a series of new benzylidene-N-(3,4-dichlorophenyl)hydrazine-1-carbothioamide derivatives 3a-3g as potential acetylcholinesterase (AChE) inhibitors for Alzheimer’s disease therapy. By integrating synthesis, biological evaluation, docking analysis, and pharmacokinetic predictions, the study presents a complete strategy that not only discovers effective inhibitors but also confirms their drug-like features. This multi-faceted strategy strengthens the reliability of the findings and highlights compound 3g as a robust lead candidate. The synthesis of derivatives was validated by employing 1HNMR, 13CNMR and HRMS spectroscopy. Biological assays demonstrated AChE inhibitory properties of the synthesized molecules, with compound 3g showing the most potent inhibitor activity, exhibiting IC₅₀ values of 0.055±0.002 µM for AChE and no activity toward BChE enzyme. Docking studies supported these results by demonstrating favorable binding conformation and stability, while pharmacokinetic and drug-likeness analyses further established compound 3g as a promising lead candidate for AD therapy.