ANO1 Reprograms Cholesterol Metabolism and the Tumor Microenvironment to Promote Cancer Metastasis

Tumor metastasis is a complex process. A better understanding of the mechanistic underpinnings of the metastatic cascade could uncover approaches to better prevent and target metastases. Here, we integrated transcriptomic profiling in primary esophageal squa-mous cell carcinoma (ESCC) and matched lymph node metastatic tissues and genome-wide CRISPR/Cas9 functional screening to identify the key drivers of cancer metastasis. Anoctamin 1 (ANO1) expression was found to be elevated in primary tumors and further increased in metastatic tissues. High expression of ANO1 was associated with poor patient survival, and ANO1 overexpression promoted cancer invasion and metastasis. Mechanistically, ANO1 inactivated the LXR pathway and increased intracellular cholesterol accumulation by interacting with JUN to repress the transcription of cholesterol hydroxylase, CYP27A1. In a paracrine manner, IL1I3 secreted from ANO1-expressing ESCC cancer cells activated NF -1(B signaling in fibroblasts to stimulate production of the chemokine CCL1, which in turn enhanced invasion of ESCC cancer cells. Furthermore, molecular docking and in silico screening of 1,500,000 small molecules identified K786-4469 as a lead com-pound that targeted ANO1 and suppressed cancer metastasis in vitro and in vivo. Overall, these findings uncover a noncanonical function of ANO1 in regulating cholesterol metabolism and fibro-blast function, revealing ANO1 as a potential prognostic biomarker and therapeutic target for metastatic ESCC. Significance: Metastatic cancer cells upregulate ANO1 to acti-vate cell-intrinsic and-extrinsic mechanisms that alter cholesterol metabolism and stimulate fibroblasts, which can be targeted with ANO1 inhibitors to inhibit metastatic growth.