Recurrent triple-negative breast cancer from cysteine deprivation loses tumorigenicity via downregulation of the CST4 signaling

Triple-negative breast cancer (TNBC) is an aggressive type of breast cancer with a high risk of recurrence following treatment. Inhibition of cysteine uptake by erastin effectively induces mesenchymal TNBC undergoing prompt ferroptosis. However, a small residual population of cancer cells keeps resistant and survived during the erastin treatments, which might mimic tumor relapse in patients after therapy. We enriched and established the erastin-resistant/recurrent TNBC cell models by multi-cycle challenges of erastin. Identified epigenetic compounds could abolish such resistance in recurrent cells. Intriguingly, recurrent TNBC failed to grow in anchorage-independent conditions, implying a loss of tumorigenicity. By analyzing transcriptomic profiling, recurrent TNBC displayed highly transcriptomic alterations and attenuated signaling processes, including K-Ras signaling. Knockout of CST4, a member of the cystatin gene family strongly suppressed in recurrent cells, significantly reduced the tumorigenic potential and K-Ras signaling. Our findings suggested that targeted cysteine therapy could be a valid treatment for TNBC with a low possibility of tumorigenicity formation. The MDA-MB-231 erastin-resistant/recurrent cells were established from their parental erastin-sensitive cells by multiple cycles of erastin treatment in vitro. RNA was extracted by RNAeasy kits (Qiagen) from the erastin-sensitive and erastin-resistant cells in triplicate. Transcriptomic profiling was analyzed by Affymetrix Clariom S Assay.