Ginsenoside Rg3 attenuates the stemness of breast cancer stem cells by activating the hippo signaling pathway

Ginsenoside Rg3 (Rg3), a bioactive compound from ginseng, is gaining attention for its potential in targeting cancer stem cells in cancer therapy. The therapeutic effect of Rg3 on breast cancer stem cells (BCSCs) has not been systematically explored using a suitable approach. Our study leverages a multi-faceted strategy, including network pharmacology, molecular docking, and in vitro experiments validation, to explore the effect of Rg3 against BCSCs. We identified 38 common targets of Rg3 and BCSCs through public databases mining. The analysis of protein-protein interaction network revealed Myc, Stat3, Bcl2, Cdh1, Egf, Il6, Egfr, Nfkb1, Sox2 and Sirt1 as the top 10 potential targets. Molecular docking further validated Rg3 has robust binding potential with these targets. Utilizing the BCSC-enriched MCF-7 and MDA-MB-231 mammosphere model, in vitro experiments substantiated Rg3's ability to induce apoptosis, suppress proliferation, and inhibit mammospheres formation of BCSCs. Rg3 also decreased the ALDH^high and CD44⁺/CD24^−/low subpopulations and downregulated the expression of cancer stem cell markers such as c-MYC, ALDH1A1, NANOG in BCSCs. After Rg3 treatment, most of the top 10 genes in BCSC-enriched MCF-7 mammospheres showed a significant reduction in expression, with Cdh1 (E-cadherin) being the most markedly downregulated. The E-cadherin/catenin complex acts as an upstream regulator of the Hippo signaling pathway, which is crucial for BCSC function and is among the top 20 enriched pathways identified by KEGG analysis. Mechanistically, Rg3 attenuates the stemness of BCSCs by activating the Hippo signaling pathway. This study provides a comprehensive evaluation of Rg3 as a promising therapeutic agent against BCSCs.