β-elemenine Induces Ferroptosis And Apoptosis In Triple-negative Breast Cancer Cell

Background:Triple-negative breast cancer（TNBC）is the most malignant breast cancer.Currently,there is a lack of effective targeted therapy,and it is urgent to explore drugs with unique targeting effects to overcome the treatment challenges of triple-negative breast cancer.Objective:In this study,in vivo and in vitro experiments were used to evaluate the therapeutic efficacy and mechanism ofβ-elemonic acid on triple-negative breast cancer cells.Methods:Firstly,human triple-negative breast cancer cell MDA-MB-231 and mouse triple-negative breast cancer cell 4T1 were cultured in vitro.MTT assay was used to detect the effects of different concentrations ofβ-elemonic acid on MDA-MB-231 and 4T1 at 24h,48h and 72h,respectively.The effects ofβ-elemonic acid on the proliferation of MDA-MB-231and 4T1 were detected by plate cloning assay.Scratch test,Transwell migration and invasion test were used to detect the effects ofβ-elemonic acid on the lateral migration,vertical migration and invasion abilities of MDA-MB-231 and 4T1.The apoptosis and cycle of MDA-MB-231 and 4T1 were detected by flow cytometry.The expression level of Fe²⁺in MDA-MB-231 and 4T1 after treatment withβ-elemonic acid was detected by Fe²⁺kit.ROS,Lipid ROS,GSH,MDA and 4-HNE were measured by lipid peroxidation assay kits,the above experiments were intervened with Ferrostatin-1.The expression of ferroptosis-related proteins GPX4,PTGS2,SLC7A11,P53 and apoptosis-related proteins P53,Bax,Bcl-2,Caspase3were detected by Western blot.Secondly,the in vivo effect was studied by constructing a mouse model of homologous transplantation tumor.HE staining and immunohistochemical experiments were used to verify whether there was toxic reaction ofβ-elemonic acid on internal organs and the potential mechanism of its effect.Results:MTT assay and plate cloning assay results showed thatβ-elemonic acid inhibited the proliferation of MDA-MB-231 and 4T1 in a concentration-dependent and time-dependent manner.Scratch test,Transwell migration and invasion test showed thatβ-elemonic acid inhibited the lateral and vertical migration and invasion abilities of MDA-MB-231 and 4T1cells in a concentration-dependent manner.Flow cytometry and cycle experiments showed thatβ-elemonic acid induced apoptosis of MDA-MB-231 and 4T1 in a concentration-dependent manner and stopped the cells from growing in the G1 phase.Western blot experiment showed thatβ-elemonic acid could up-regulate the expression levels of P53,Bax,Caspase3,and PTGS2 in MDA-MB-231 and 4T1,and down-regulate the expression levels of Bcl-2,GPX4,and SLC7A11.In vitro studies have shown thatβ-elemonic acid can activate the ferroptosis pathway and apoptosis pathway in breast cancer,significantly inhibit the growth of MDA-MB-231 and 4T1,and induce cell death.In vivo studies showed thatβ-elemonic acid effectively inhibited tumor growth in mice with syngeneic transplanted tumor without obvious toxicity.Conclusion:Through cell experiments and animal experiments showed thatβ-elemonic acid can induce the activation of ferroptosis and apoptosis pathways in triple-negative breast cancer,and can exert significant anti-tumor activity in triple-negative breast cancer.