Solution-phase multifunctional modulation of cholinesterases and carbonic anhydrases by thiourea–pyrimidine benzenesulfonamides: Kinetics and explicit-solvent molecular dynamics

In aqueous media, enzyme–ligand recognition is shaped not only by direct contacts but also by solvent-mediated interaction networks that persist across time scales. Here, cholinesterases (AChE and BChE) and cytosolic human carbonic anhydrases ( h CA I and h CA II) were investigated as two disease-relevant enzyme systems in solution, and twelve thiourea–pyrimidine benzenesulfonamide derivatives were synthesized and structurally characterized. Enzyme inhibition was quantified by steady-state kinetics; inhibition constants ( K I) were obtained by nonlinear global fitting of untransformed rate data, with Lineweaver–Burk plots used only for visualization. To connect solution-phase inhibition with molecular recognition, docking was validated by redocking (heavy-atom RMSD <2.0 Å across targets), and representative complexes were examined by 500 ns explicit-solvent MD simulations (TIP3P, 0.15 M NaCl, NPT), revealing stable binding modes and persistent, frequently water-mediated interaction patterns consistent with experimental trends. Cellular relevance was assessed in MKN-28 gastric cancer cells with L929 fibroblasts as a non-malignant control. Compounds with improved cytotoxic selectivity toward MKN-28 were identified, and the lead compound ( 11 ) was further evaluated by Annexin V/PI flow cytometry, showing a shift from viable cells toward apoptotic populations under the tested conditions. In silico ADMET profiling suggested moderate lipophilicity across the series, while highlighting elevated polar surface area and compound-dependent toxicity flags as parameters to monitor during optimization. Overall, thiourea–pyrimidine benzenesulfonamides emerge as multifunctional modulators of ChEs and cytosolic h CAs within the applied biochemical and cellular assays. © 2026 Elsevier B.V.