Development of a Novel Benzanilide Derivative Targeting VEGFR-2: Computational Modeling and <i>In Vitro</i> Anticancer Validation

Abstract

The pursuit of selective vascular endothelial growth factor receptor-2 (VEGFR-2) inhibitors continues to be a key approach in anticancer drug discovery. In this context, a new benzanilide-based derivative, Z-CMPBC, was designed following essential VEGFR-2 pharmacophoric requirements and investigated through combined computational and biological approaches. Density functional theory (DFT) calculations at the B3LYP/6-31[Formula: see text]G(d, p) level indicated favorable electronic characteristics, including balanced chemical hardness and pronounced electrophilicity, suggesting a strong propensity for interaction within kinase active sites. Molecular docking, supported by 200 ns molecular dynamics simulations, demonstrated stable accommodation of the compound within the VEGFR-2 binding pocket. Binding free energy calculations using the MM-GBSA method revealed a significant interaction profile ([Formula: see text] kcal/mol), mainly governed by van der Waals forces and persistent hydrophobic interactions with critical residues. Biological assessment confirmed potent VEGFR-2 inhibition, with an IC[Formula: see text] value of [Formula: see text]M, closely comparable to sorafenib ([Formula: see text]M). Western blot analysis further verified a reduction in phosphorylated VEGFR-2 (pY1054/Y1059) levels in MCF-7 cells, supporting effective intracellular kinase inhibition. Additionally, Z-CMPBC exhibited pronounced cytotoxicity, particularly against MCF-7 breast cancer cells (IC[Formula: see text]M), along with a high selectivity index ([Formula: see text]) relative to normal Vero cells. Flow cytometry analysis demonstrated induction of G0/G1 phase arrest, accompanied by enhanced apoptotic activity (34.05%) compared to untreated cells. In silico ADMET and toxicity predictions suggested acceptable pharmacokinetic behavior and safety characteristics. Overall, these findings highlight Z-CMPBC as a promising VEGFR-2 inhibitor with potential for further development as an anticancer agent.