Fenugreek seed extract–doxorubicin synergy against hepatocellular carcinoma in HepG2 cells: in vitro and in silico mechanistic studies

Abstract Background Hepatocellular carcinoma (HCC) is one of the most prevalent malignancies worldwide. The therapeutic efficacy of conventional chemotherapeutic agents such as doxorubicin (DOX) is limited by dose-dependent toxicity and the development of drug resistance. Combination strategies incorporating bioactive natural products may enhance anticancer efficacy while enabling dose reduction. The present study aimed to evaluate the potential synergistic cytotoxicity of combining Fenugreek aqueous extract (FAE) with (DOX) against the HepG2 cell line. Methods Phytochemical characterization was performed using UHPLC-QTOF-MS/MS Profiling and HPLC. Cell viability and selectivity were assessed using the SRB assay. Apoptosis, necrosis, autophagy, and cell cycle distribution were analysed by flow cytometry and Western blotting. Drug–drug interaction was evaluated using the Chou–Talalay method. Molecular docking was performed to explore potential interactions between selected FAE constituents and apoptosis- and autophagy-related protein targets. Results FAE enhanced DOX’s cytotoxicity on HepG2 cells, with the interaction ranging from synergistic to additive depending on the concentration ratio. The DOX/FAE combination enhanced cell death through sub-G1 arrest and augmented apoptotic, necrotic, and autophagic responses compared with monotherapies. Western blot analysis demonstrated modulation of the Bax/Bcl-2 ratio and increased LC3-II expression. Docking simulations suggested favourable binding of selected steroidal saponins to Bcl-2 and LC3 proteins. Conclusion These findings indicate that FAE potentiates DOX-induced cytotoxicity in vitro through modulation of multiple regulated cell death pathways. While the results support the possibility of this combination as a dose-modulating strategy, further validation in additional HCC models and in vivo systems is required. Graphical abstract