Hesperidin Enhances Doxorubicin Cytotoxicity in MCF-7 Breast Cancer Cells: Evidence from Cellular, Gene Expression, Global DNA Methylation, and Molecular Docking Analyses
Reham M. Abdel Moneim, Hatim A. El-Baz, Mohamed Farouk ElshalIntroduction/Objective:
Doxorubicin efficacy in breast cancer is constrained by chemoresistance and toxicity. Hesperidin, a citrus flavonoid, exhibits anticancer and epigenetic regulatory properties. This study evaluated whether hesperidin enhances doxorubicin cytotoxicity in MCF-7 cells and characterized the associated apoptotic and Heat-Shock (HSP60, HSP90) transcripts, as well as global DNA methylation.
Methods:
Cytotoxicity and synergy were assessed using the MTT assay and the Chou-Talalay method (CI). Apoptosis and cell cycle were analyzed by flow cytometry. Expression of BAX, BCL2, HSP60, and HSP90 was quantified using RT-qPCR. Global DNA methylation (5-mC%) was assessed colorimetrically, alongside molecular docking simulations for HSP60 and HSP90.
Results:
Combining hesperidin with doxorubicin produced synergistic cytotoxicity (CI < 1) and a dosereduction index of 1.95 for doxorubicin. The combination induced apoptosis, accompanied by upregulation of BAX, downregulation of BCL2, and modulation of HSP60/HSP90 transcript levels. Additionally, the combination attenuated doxorubicin-induced global hypermethylation. Molecular docking demonstrated favorable binding affinities of hesperidin toward both HSP60 and HSP90, particularly HSP60; however, these findings remain correlative without proving direct target engagement.
Discussion:
The synergistic response, paired with modulated apoptotic markers and global DNA methylation, suggests that hesperidin sensitizes MCF-7 cells to doxorubicin through multi-targeted cellular and epigenetic pathways. While docking and transcriptomic shifts indicate potential chaperone involvement, these preliminary observations require further functional validation.
Conclusion:
Hesperidin enhances doxorubicin cytotoxicity in MCF-7 cells by modulating apoptosis, stressresponse transcripts, and DNA methylation. These in-vitro findings support further mechanistic and in-vivo evaluation of this combination strategy.