Detection of phenotype-specific therapeutic vulnerabilities in breast cells using a CRISPR loss-of-function screen.

Cellular phenotype plasticity between epithelial and mesenchymal states has been linked to metastasis and heterogeneous responses to cancer therapy, and remains a challenge for the treatment of triple-negative breast cancer (TNBC). Here we used isogenic human breast epithelial cell lines, D492 and D492M, representing the epithelial and the mesenchymal phenotype, respectively. We employed a CRISPR-Cas9 loss-of-function screen targeting a 2240-genes "druggable genome" to identify phenotype-specific vulnerabilities. Cells with the epithelial phenotype were more vulnerable to the loss of genes related to EGFR-RAS-MAPK signaling, while the mesenchymal-like cells had increased sensitivity to knockout of G2 -M cell cycle regulators. Furthermore, we discovered knockouts that sensitize to the mTOR inhibitor everolimus and the chemotherapeutic drug fluorouracil in a phenotype-specific manner. Specifically, loss of EGFR and Fatty Acid Synthase (FASN) increased the effectiveness of the drugs in the epithelial and the mesenchymal phenotype, respectively. These phenotype-associated genetic vulnerabilities were confirmed using targeted inhibitors of EGFR (gefitinib), G2 -M transition (STLC) and FASN (Fasnall). In conclusion, a CRISPR-Cas9 loss-of-function screen enables the identification of phenotype-specific genetic vulnerabilities that can pinpoint actionable targets and promising therapeutic combinations.