Madecassoside Induces Apoptosis and Inhibits Migration by Regulating ROS-Mediated Signaling Pathways in MDA-MB-231...

Madecassoside (MC) is an anti-inflammatory and antibacterial active substance extracted from the traditional Chinese medicine Centella asiatica. Based on network pharmacology and experimental validation, this study assessed MC's anti-breast cancer (BC) actions and related molecular pathways. CCK-8, Trypan Blue, and Hoechst33342/PI assays showed that MC significantly reduced BC cell activity, but it had little inhibitory effects on normal cells. Network pharmacology analysis predicted 40 intersection targets between MC and BC, 291 GO-related biological processes, and 105 KEGG signaling pathways. The anti-breast cancer activity of MC is closely related to reactive oxygen species (ROS), AKT and MAPK signaling pathways. Through Annexin V-FITC/PI, flow cytometry, transwell, wound healing and western blotting experiments, it was found that MC induced mitochondria-dependent apoptosis, G2/M phase arrest and inhibited cell migration of MDA-MB-231 cells through MAPK/STAT3/NF-κB signaling pathway, PI3K/AKT signaling pathway and AKT/GSK-3β/β-catenin signaling pathway, respectively. The induction and blocking effects were inhibited by the addition of ROS scavenger N-acetyl cysteine (NAC). Molecular docking showed that MC had significant binding ability with STAT3, CASP3, BCL2 and JUN targets in BC. In summary, MC induced apoptosis, cell cycle arrest, and migration inhibition via ROS-mediated MAPK/STAT3/NF-κB, PI3K/AKT, and AKT/GSK-3β/β-catenin signaling pathways in MDA-MB-231 cells.