Simple SummaryMultiple non-cancerous cells are known to be involved in brain metastasis, and the roles of neutrophils during this event are poorly understood. We aim to understand how tumor-infiltrated neutrophils promote breast cancer brain metastasis and how tumor cells affect the properties of neutrophils. Utilizing patient-based analyses together with our unique animal models, we discovered that several c-Met mediated inflammatory cytokines, including CXCL1/2 and G/GM-CSF, are critical to the neutrophil recruitment and activity in the metastatic lesions. In return, neutrophils activated by those factors secrete a high level of lipocalin 2 (LCN2), which in turn enhances the stemness of tumor cells. Our study revealed novel interactions between neutrophils and brain metastatic cells, which may offer new insight into treating brain metastasis.AbstractThe brain is one of the most common metastatic sites among breast cancer patients, especially in those who have Her2-positive or triple-negative tumors. The brain microenvironment has been considered immune privileged, and the exact mechanisms of how immune cells in the brain microenvironment contribute to brain metastasis remain elusive. In this study, we found that neutrophils are recruited and influenced by c-Met high brain metastatic cells in the metastatic sites, and depletion of neutrophils significantly suppressed brain metastasis in animal models. Overexpression of c-Met in tumor cells enhances the secretion of a group of cytokines, including CXCL1/2, G-CSF, and GM-CSF, which play critical roles in neutrophil attraction, granulopoiesis, and homeostasis. Meanwhile, our transcriptomic analysis demonstrated that conditioned media from c-Met high cells significantly induced the secretion of lipocalin 2 (LCN2) from neutrophils, which in turn promotes the self-renewal of cancer stem cells. Our study unveiled the molecular and pathogenic mechanisms of how crosstalk between innate immune cells and tumor cells facilitates tumor progression in the brain, which provides novel therapeutic targets for treating brain metastasis.