Design, synthesis, and biological evaluation of novel aryl isoxazoles as potential Hsp90 inhibitors

Heat shock protein 90 (Hsp90) is a molecular chaperone critical for the stabilization of numerous oncoproteins, making it a promising target for anticancer drug development. This study aimed to design, synthesize, and evaluate novel 3,5-diarylisoxazole derivatives as potential Hsp90 inhibitors.

A series of 3,5-diarylisoxazole compounds (5R_1-6) was designed using molecular docking to predict binding affinity. Compounds were synthesized via cyclization of 4-chloroacetophenone with hydroxylamine hydrochloride, followed by condensation with various aldehydes. Structures were confirmed by melting point, FT-IR, and 1H-NMR spectroscopy. The MTT assay assessed cytotoxicity, and Hsp90 inhibitory activity was evaluated using an Hsp ELISA kit. The stability of the 5R3 compound in the Hsp90 active site was investigated using molecular dynamics simulation.

All compounds showed favorable binding energies. Compound 5R3 showed the highest binding affinity (ΔG = -7.83 kcal/mol) and the highest cytotoxicity against MCF-7 cells (IC50 = 0.014 μM) and significantly reduced Hsp90 concentration from 5.54 ng/mL (untreated) to 1.56 ng/mL. Furthermore, molecular dynamics simulation studies confirmed the stability of the Hsp90-5R₃ complex during a 100 ns simulation.

Compound 5R₃ with an electronegative substituent (F) on the aromatic ring showed the highest cytotoxic effect, also decreased the concentration of Hsp90 protein more than others.