GPX4 Inhibitor Resistance and Metastatic Features in Triple‐Negative Breast Cancer

ABSTRACT Leveraging ferroptosis as a cancer therapy has faced challenges due to the limited bioavailability and systemic toxicities of small‐molecule ferroptosis modulators. Small molecule inhibitors such as RSL3 and ML210 trigger ferroptosis by targeting glutathione peroxidase 4 (GPX4), a key enzyme that neutralizes lipid peroxides. While many studies have focused on targeting primary tumors, much less is known about the extent to which GPX4‐inhibitor resistance may contribute to metastasis. To address this, we cultured triple‐negative breast cancer cell lines with GPX4 inhibitors to generate cell lines (M231, 4T1) that were resistant to GPX4 inhibitors (GPX4i). Tumors derived from GPX4i‐resistant cells compared to parental cells had unique metabolic and lipidomic profiles, were associated with a shift toward an epithelial‐like state (decreased vimentin, increased EpCAM expression), formed decreased spontaneous metastases from primary tumors, but had no differences in overall metastatic burden upon intravenous injection. Collectively, these data demonstrate that long‐term maintenance with GPX4‐inhibitors in vitro leads to altered metastatic profiles in vivo.