To examine how E2 promotes brain metastasis by upregulating M2 microglia polarization, we treated breast cancer brain metastatic cells, SKBrM (ER−/Her2+) and 231BrM (ER−/PR−/Her2−), with conditioned medium (CM) that was generated from human HMC3 microglia treated with or without E2. Here, ESR1 is linked to breast cancer.