There is currently no curative treatment for patients diagnosed with triple-negative breast cancer brain metastases (TNBC-BM). CAR T cells hold potential for curative treatment given they retain the cytolytic activity of a T cell combined with the specificity of an antibody. In this proposal we evaluated the potential of EGFR re-directed CAR T cells as a therapeutic treatment against TNBC cells in vitro and in vivo. MethodsWe leveraged a TNBC-BM tissue microarray and a large panel of TNBC cell lines and identified elevated epidermal growth factor receptor (EGFR) expression. Next, we designed a second-generation anti-EGFR CAR T construct incorporating a clinically relevant mAb806 tumor specific single-chain variable fragment (scFv) and intracellular 4-1BB costimulatory domain and CD3 using a lentivirus system and evaluated in vitro and in vivo anti-tumor activity. ResultsWe demonstrate EGFR is enriched in TNBC-BM patient tissue after neurosurgical resection, with six of 13 brain metastases demonstrating both membranous and cytoplasmic EGFR. Eleven of 13 TNBC cell lines have EGFR surface expression ≥85% by flow cytometry. EGFR CAR T treated mice effectively eradicated TNBC-BM and enhanced mouse survival (log rank p<0.004). Conclusion Our results demonstrates anti-tumor activity of EGFR CAR T cells against TNBC cells in vitro and in vivo.Given EGFR CAR T cells are currently undergoing clinical trials in primary brain tumor patients without obvious toxicity, our results are immediately actionable against the TNBC-BM patient population.