Abstract
BACKGROUND/OBJECTIVES: Trachystemon orientalis (TO) is native to Bulgaria, the western Caucasus, and Turkey. It possesses antimicrobial, antifungal, and antidiabetic properties. Due to their efficacy and lower toxicity, the search for natural compounds with anticancer potential, especially for breast cancer, has increased. This study evaluated the cytotoxic effects on breast cancer cells. METHODS: MCF-7 breast cancer cells and CCD-1072Sk normal fibroblast cells were treated with TO extracts prepared using water, methanol, ethanol, and dimethyl sulfoxide (DMSO). Cytotoxicity was assessed in a dose- and time-dependent manner, and IC₅₀ values were calculated. The most potent extract, water, was analyzed for its effects on cancer-related pathways via RT-qPCR, gene enrichment, and gene-metabolite association analyses. Flow cytometry was used to assess cell cycle distribution and apoptosis, and fluorescence staining was used to evaluate nuclear, cytoskeletal, and mitochondrial morphology. RESULTS: TO exhibited dose-dependent cytotoxicity in MCF-7 cells, with IC₅₀ values ranging from 28.88 ± 1.91 ng/µL (water extract) to 185.30 ± 6.60 ng/µL (ethanol extract), demonstrating lower toxicity in normal cells. KEGG pathway enrichment analysis showed that the water extract was associated with the modulation of several cancer-related signaling pathways, including p53, HIF-1, TNF, PI3K–Akt, and MAPK pathways, which induced S-phase arrest and increased apoptosis by 11% as confirmed by morphological changes. CONCLUSIONS: Taken together, these findings indicate that TO has cytotoxic and apoptosis-inducing effects on breast cancer cells through the regulation of multiple signaling pathways and is a potential candidate for further validation in breast cancer research. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12906-026-05334-1.