Synthesis, Antitumor Activity Evaluation and Mechanistic Study of Novel Bis‐Heterocyclic Chalcones Against Liver Cancer

ABSTRACT Background Chalcones and heterocyclic compounds exhibit remarkably high activity in medicinal chemistry. In recent years, bis‐chalcones have been reported to possess excellent anticancer activity. We synthesized a series of bis‐heterocyclic chalcones via asymmetric chain synthesis, with the aim of making new discoveries in anticancer activity. Methods Bis‐heterocyclic chalcones were synthesized via Claisen‐Schmidt condensation and alkylation reactions. The inhibitory activities of the synthesized compounds against Huh‐1, Huh‐7, and HepG2 cell lines were evaluated using the CCK‐8 assay. Furthermore, the mechanism of action of these compounds was explored through live/dead cell staining, flow cytometric analysis, and Western blotting experiments. Results Twelve bis‐heterocyclic chalcone compounds were synthesized. All synthesized compounds were fully characterized by spectroscopic methods and evaluated for their cytotoxic activities against Huh‐1, Huh‐7, and HepG2 cell lines using the CCK‐8 assay at concentrations ranging from 0 to 100 μM. Among them, derivative 3f exhibited the most potent cytotoxicity against Huh‐7 (IC 50 = 8.40 μM) and Huh‐1 (IC 50 = 6.75 μM), whereas HepG2 cells were most sensitive to compound 3d (IC 50 = 27.99 μM). The mechanisms underlying the antitumor effects of 3d and 3f were further investigated through live/dead cell staining, flow cytometry, and western blot analysis. The results demonstrate that both compounds effectively induce apoptosis in liver cancer cells. Conclusions Bis‐heterocyclic chalcone compounds exhibit favorable inhibitory activity against hepatocellular carcinoma cells by inducing cell apoptosis, and thus can serve as a class of pharmaceutically active structural units for further in‐depth research and screening.