Novel trans-stilbene-phytophenol conjugates as potent tubulin-targeting antiproliferative agents

A series of novel trans-stilbene-phytophenol/phenolic acid conjugates linked through a triazole moiety were synthesized via Knoevenagel condensation and copper-catalyzed azide-alkyne cycloaddition. Among the 24 synthesized compounds 23-46, compound 24 emerged as the most potent candidate, exhibiting strong cytotoxicity against MCF-7 and MDA-MB-231 breast cancer cells, while showing substantially lower toxicity toward normal NKE cells. Compound 24 markedly suppressed clonogenic survival and induced apoptosis, as demonstrated by Annexin V/PI staining, enhanced caspase-3 activity, and nuclear condensation and fragmentation. Cell-cycle analysis revealed pronounced G2/M arrest with increased sub-G0 accumulation. Mechanistically, compound 24 significantly increased intracellular and mitochondrial reactive oxygen species (ROS), accompanied by substantial mitochondrial membrane depolarization, suggesting ROS-mediated mitochondrial dysfunction as an important contributor to apoptosis. Molecular docking demonstrated favorable binding of compound 24 within the colchicine-binding pocket of β-tubulin supporting stable protein-ligand interactions. Furthermore, in silico ADMET profiling indicated favorable drug-like and predicted safety characteristics. Collectively, compound 24 represents a promising tubulin-targeting anticancer lead that suppresses breast cancer cell survival through coordinated G2/M arrest, oxidative stress, mitochondrial dysfunctions, and caspase-dependent apoptosis, warranting further preclinical investigation.