Design, Synthesis, and Biological Evaluation of Chiral Urea and Thiourea Derivatives as Multitarget-Directed...

A series of novel chiral urea and thiourea compounds were designed as multitarget-directed ligands for Alzheimer's disease and evaluated against hAChE, hBChE, and hMAO-B, with exploratory assessment of HSD10. These chiral compounds were synthesized and fully characterized, and their enantiopurity was confirmed via HPLC. Biological screening revealed that several of the 38 single-enantiomer derivatives exhibited potent triple inhibition of hAChE, hBChE, and hMAO-B. Biological screening revealed that a significant number of the 38 synthesized single-enantiomer urea and thiourea derivatives potent inhibition across the primary target panel. Among them, compounds 14S (hAChE: 0.08 μM, hBChE: 0.93 μM, hMAO-B: 0.02 μM), 23R (hAChE: 0.06 μM, hBChE: 0.74 μM, hMAO-B: 0.08 μM), 6S (hAChE: 0.08 μM, hBChE: 0.65 μM, hMAO-B: 0.08 μM), and 30S (hAChE: 0.04 μM, hBChE: 0.74 μM, hMAO-B: 0.19 μM) showed the most promising multitarget profiles. Compounds 6S and 23R also displayed anti-Aβ aggregation activity, whereas 12S combined dual hMAO-B/hAChE inhibition with antioxidant activity. Compounds 3R and 11R were found to be highly active against hMAO-B (0.03 μM and 0.06 μM, respectively) and hBChE (0.68 and 0.72 μM). HSD10 inhibition was limited overall, with only compound 26S showing moderate activity (52.2%) at higher micromolar concentration. Molecular docking and molecular dynamics simulations supported the proposed ligand-target interactions for the most active compounds. In addition, selected derivatives showed no acute cytotoxicity under the tested conditions in HepG2 cells and high permeability potential in the PAMPA-BBB assay. Based on their in vitro profiles, seven compounds (3R, 6S, 11R, 12S, 14S, 23R, and 30S) were advanced to in vivo evaluation, and compounds 6S, 12S, and 23R, 30S produced significant improvements in the Morris Water Maze model. Collectively, these findings identify chiral urea and thiourea derivatives as promising lead structures for further anti-Alzheimer drug discovery.