6′'-O-acetylsaikosaponin D targets STAT3-mediated transcriptional remodeling to induce ferroptosis and apoptosis

Background Natural products from traditional medicinal plants represent an important source of anti-cancer agents and lead compounds for drug development. Saikosaponin D (SSD), a triterpenoid saponin isolated from Radix Bupleuri, exhibits anti-tumor activity but is limited by a narrow therapeutic window and cytotoxicity. Objective This study aimed to evaluate the anti-tumor activity of 6′'-O-acetylsaikosaponin D (6′'-O-acSSD), a naturally occurring acetylated derivative of Saikosaponin D that is identified in Radix Bupleuri , and to elucidate its underlying molecular mechanisms. Methods The anti-cancer effects of 6′'-O-acSSD were assessed using multiple cancer cell models, three-dimensional tumor spheroids, and patient-derived tumor organoids. Transcriptomic and bioinformatic analyses were performed to identify regulated cell death pathways. Molecular docking, molecular dynamics simulation, cellular thermal shift assays, western blotting, and CUT&Tag profiling were used to identify molecular targets and define transcriptional reprogramming. Results 6′'-O-acSSD markedly suppressed cancer cell proliferation, migration, and invasion and elicited a transcriptional program distinct from that induced by SSD. Transcriptomic and biochemical analyses revealed preferential activation of ferroptosis-associated signatures, including enhanced lipid peroxidation. Mechanistically, 6′'-O-acSSD engaged STAT3, inhibited its phosphorylation, and extensively remodeled STAT3 chromatin occupancy. Functional analyses showed that STAT3 inhibition was required for the full anti-tumor activity of 6′'-O-acSSD. Notably, blockade of ferroptosis only partially rescued cell viability, whereas STAT3 targeting additionally triggered apoptosis-associated transcriptional and biochemical changes. The anti-tumor effects of 6′'-O-acSSD were further validated in patient-derived tumor organoids. Conclusion 6′'-O-acSSD exerts potent anti-tumor effects by targeting STAT3-dependent transcriptional programs and inducing coordinated ferroptosis and apoptosis. These findings highlight the therapeutic potential of structurally optimized natural products for anti-cancer treatment.