Multi-pronged molecular insights into flavonoid-mediated inhibition of squalene epoxidase: a pathway to novel therapeutics

Abstract

Squalene epoxidase (SQLE) is a crucial enzyme in the sterol biosynthesis pathway and a promising target for therapeutic intervention in hypercholesterolemia and fungal infections. This study evaluates the inhibitory potential of six flavonoids namely silibinin, baicalin, naringenin, chrysin, apigenin-7-O-glucoside, and isorhamnetin against SQLE using an integrative approach combining <i>in silico</i> and experimental methods. Molecular docking revealed that apigenin-7-O-glucoside, silibinin, and baicalin displayed the highest binding affinities (-10.7, -10.2, and -10.0 kcal mol^-1, respectively) and robust interactions with the SQLE binding site. These findings were corroborated by 200 ns molecular dynamics (MD) simulations, which demonstrated stable binding trajectories, minimal structural fluctuations, a thermodynamically favored potential energy landscape (PEL) and favorable MM/PBSA binding free energies for three flavonoids. Experimental validation <i>via in vitro</i> inhibition assays confirmed the computational predictions, with apigenin-7-O-glucoside emerging as the most potent inhibitor (IC₅₀ = 1.74 ± 0.05 μM), followed by silibinin (IC₅₀ = 1.88 ± 0.28 μM) and baicalin (IC₅₀ = 2.50 ± 0.46 μM). Enzyme kinetics studies revealed distinct mechanisms of action: apigenin-7-O-glucoside exhibited competitive inhibition, while silibinin and baicalin showed mixed inhibition. Furthermore, <i>in silico</i> ADMET analysis indicated favorable pharmacokinetic and pharmacodynamic profiles for these flavonoids, with silibinin demonstrating particularly high bioavailability and lipophilicity. This study highlights apigenin-7-O-glucoside, silibinin, and baicalin as potent SQLE inhibitors with promising therapeutic potential. The congruence between <i>in silico</i> predictions and experimental results underscores the reliability of computational approaches in drug discovery, paving the way for future preclinical development of these compounds as novel SQLE-targeted therapeutics.