The anti-Alzheimer potential of new spirooxindole derivatives containing 1,4-dihydropyridines as a targeted cholinesterase inhibitor

In this study, a series of new spirooxindole derivatives containing 1,4-dihydropyridines compounds were designed, synthesized and their cholinesterase inhibitory activities were evaluated. A novel, environmentally friendly, and high-yield method for the synthesis of spiro[indoline-3,4-pyridine]-3‑carbonitrile derivatives was developed. This method is based on multicomponent, cascade reactions between different isatins, malononitrile, and 1,1-enediamines (EDAMs), leading to the production of the target compounds in favorable to excellent yields. In vitro acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory activities were investigated, and the most potent inhibitor of BChE was compound 4i , which contained a 5-Cl group at R 1 and a 4-CH 3 group at R 2 , with an IC 50 value of 5.32 ± 0.48 μM. Kinetic experiments revealed 4i to be a mixed-type inhibitor with a K i of 4.2 μM. Molecular docking and molecular dynamics simulations also confirmed the effectiveness of 4i by interacting with key residues of the catalytic triad in the catalytic active site (CAS) and the peripheral anionic site (PAS) pockets of the enzyme, yielding low RMSD and RMSF values. In silico ADME-Tox predictions also confirmed the drug-likeness and safety profile of 4i . The cell-based assay in SH-SY5Y cells did not show toxicity at concentrations up to 100 μM, with moderate neuroprotective activity at 25 μM. In general, the combined biological screening, computational modeling, and cell-based studies identify compound 4i as an effective lead compound for the development of selective BChE inhibitors.