Rosmarinic acid promotes mitochondrial fission and induces ferroptosis in triple-negative breast cancer cells

Breast cancer is the most common malignant tumor in women. Among its subtypes, triple-negative breast cancer (TNBC) is more aggressive and poses a serious threat to women's health. Rosmarinic acid (RA) is a natural polyphenolic compound known for its diverse pharmacological activities, with its antioxidant and anticancer properties being particularly notable. This study investigated the effects of RA on TNBC cell lines and explored its potential mechanisms. CCK-8 and colony formation assays were used to evaluate the potential inhibitory effects of RA on TNBC cells and to measure intracellular reactive oxygen species (ROS) levels. Flow cytometry was employed to analyze the cell cycle and apoptosis. RNA-seq analysis was performed to investigate the potential mechanisms of RA on MDA-MB-231 cells. RA inhibited the proliferation of TNBC cells in a concentration-dependent manner and reduced intracellular ROS levels. RA induced cell cycle arrest at the G1/G0 phase and promoted apoptosis by decreasing mitochondrial membrane potential. RNA-seq differential expression analysis, identified 1,929 differentially expressed genes, including 601 upregulated genes and 1,328 downregulated genes. Kyoto Encyclopedia of Genes and Genomes (KEGG), and gene set enrichment analysis (GSEA) showed that these differentially expressed genes were significantly enriched in pathways associated with ferroptosis, ABC transporters, and fatty acid metabolism. Additionally, RA significantly upregulated the expression of dynamin-related protein 1 (DRP1) in MDA-MB-231 cells, promoting mitochondrial fission, disrupting mitochondrial dynamics, and leading to dysfunction. Furthermore, RA increased the expression of intracellular ferroportin and heme oxygenase 1 (HMOX-1), resulting in elevated intracellular iron levels. The study suggests that RA inhibits the proliferation of TNBC cells through multiple mechanisms and may have potential therapeutic effects in the treatment of TNBC.