Saikosaponin D potentiates the antineoplastic effects of doxorubicin in drug-resistant breast cancer through perturbing NQO1-mediated intracellular redox balance

Background: Drug resistance to doxorubicin (DOX) significantly limits its therapeutic efficacy in breast cancer (BC) patients. Saikosaponin D (SSD), a triterpene saponin derived from the traditional herb Radix Bupleuri, , has shown promise as a chemotherapeutic sensitizer in preclinical studies due to its notable antitumor activity. However, the role and mechanism of SSD in DOX-resistant BC cells remain largely unexplored. Purpose: This study aimed to investigate the chemosensitizing effect of SSD on DOX-resistant BC and the underlying molecular mechanisms both in vitro and in vivo. . Methods: In vitro assays, including cell viability, clone formation, three-dimensional tumor spheroid growth, and apoptosis analysis, were conducted to evaluate the synergistic effect of SSD and DOX on resistant BC cells. Reactive oxygen species (ROS), GSH/GSSG, NADPH/NADP+, + , and NADH/NAD+ + detections were employed to assess the impact of SSD on cellular redox homeostasis. Western blotting, cell cycle distribution assay, and DOX uptake assay were performed to further elucidate the possible antineoplastic mechanism of SSD. Finally, a subcutaneous MCF7/DOX cell xenografted model in nude mice was established to identify the in vivo anticarcinogenic effect of SSD combined with DOX. . Results: SSD significantly inhibited cell viability, proliferation, and clone formation, enhancing DOX's anticancer efficacy in vitro and in vivo. . Mechanistically, SSD reduced STAT1, NQO1, and PGC-1 alpha protein levels, leading to cellular redox imbalance, excessive ROS generation, and depletion of GSH, NADPH, and NADH. SSD induced DNA damage by disrupting redox homeostasis, resulting in G0/G1 phase cell cycle arrest. Additionally, SSD increased DOX accumulation in BC cells via inhibiting P-gp protein expression and efflux activity. Conclusion: We demonstrated for the first time that SSD enhances the sensitivity of chemoresistant BC cells to DOX by disrupting cellular redox homeostasis through inactivation of the STAT1/NQO1/PGC-1 alpha signaling pathway. This study provides evidence for SSD as an adjuvant agent in drug-resistant BC treatment.