Tong Liu, Yuan Li
Objective To investigate the effect of polyphyllin I on MCF-7 spheroids enriched with cancer stem cells and to preliminarily observe its effect on the hybrid epithelial-mesenchymal transition （EMT） state and its regulatory role on cell stemness. Methods MCF-7 spheroids were cultured under serum-free，non-adherent conditions.Cell viability，invasion，and key protein expression were assessed through the cell counting kit-8 assay，Transwell assay，and Western blot，respectively.The CD44+/CD24-/low cell population was analyzed by flow cytometry. Results The MCF-7 spheroids exhibited high proliferative and invasive capacities，co-expressing the epithelial marker E-cadherin and the mesenchymal marker vimentin，which is indicative of a hybrid EMT state and high stemness.Polyphyllin I treatment significantly suppressed the viability and invasion of MCF-7 spheroids.From a mechanism perspective，polyphyllin I downregulated the E-cadherin/vimentin ratio，significantly reduced the levels of stemness-associated proteins CD44 antigen，aldehyde dehydrogenase 1 family member A1，as well as the ratio of phosphorylated signal transducer and activator of transcription 3 to total signal transducer and activator of transcription 3.Simultaneously，polyphyllin I decreased the proportion of CD44+/CD24-/low cells.No significant changes were observed in key proteins of the intrinsic apoptosis pathway. Conclusions Polyphyllin I reshapes the hybrid EMT state in MCF-7 spheroids，which is accompanied by the disruption of their high stemness equilibrium.This compound may reduce cellular stemness through the EMT remodeling process，thereby exerting antitumor effects，and this mechanism is independent of the intrinsic apoptosis pathway.