As expected, we found that after reducing GPR30 expression, calycosin caused a decrease the phosphorylation levels of SRC, EGFR, ERK1/2, and Akt in breast cancer cell lines with and without ER expression (MCF-7, T47D, MDA-MB-468, and SKBR3), but calycosin did not affect the expression of p-SRC, p-EGFR, p-ERK1/2, or p-Akt in GPR30-deficient MDA-MB-231 cells. This evidence concerns the gene GPER1 and breast cancer.