Disli_2026_Bioorg.Chem_181_110350

Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterized by neuronal loss and cognitive decline. A key pathological feature of AD is the reduction of acetylcholine (ACh) levels resulting from the enzymatic activity of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). Therefore, inhibition of these enzymes remains a well-established therapeutic strategy for AD management. In this study, a novel series of phenothiazine-tetrazole hybrids was designed and synthesized as potential cholinesterase inhibitors. Their inhibitory activities against AChE and BChE were evaluated in vitro, and IC50 values were determined in the micromolar range. Among the tested derivatives, Compound E3a (IC50 = 10.664 microM) and Compound E5b (IC50 = 10.315 microM) exhibited the strongest AChE inhibition, whereas Compound D5b showed the highest activity toward BChE (IC50 = 18.734 microM). Enzyme kinetic analysis revealed that E5b and E3a act as competitive AChE inhibitors, with Ki values of 1.313 +/- 0.032 microM and 1.520 +/- 0.033 microM, respectively. To elucidate the molecular basis of inhibition, molecular docking studies were performed, followed by 500 ns molecular dynamics simulations, MM-PBSA binding free energy calculations, and free energy landscape analyses. The computational results revealed that Compound E3a forms a more dynamically and thermodynamically stabilized complex with AChE, primarily driven by favorable van der Waals interactions. ADMETlab 3.0 predictions further indicated acceptable preliminary drug-like features, although lipophilicity, solubility, CYP liability, and toxicity-related endpoints require further optimization and experimental validation. Collectively, these findings highlight phenothiazine-tetrazole hybrids as promising scaffolds for the rational development of next-generation cholinesterase inhibitors for Alzheimer's therapy.