This study aims to determine the cellular functions and clinical significance of microRNA-409 (miR-409) in breast cancer by targeting special AT-rich sequence-binding protein 1 (SATB1). Breast cancer tissues and adjacent normal tissues, breast cancer cell lines (MDA-MB-453, MDA-MB-231, BT-549, BR3, and MCF-7) were used. miR-409 mimics, miR-409 inhibitor, SATB1, and siSATB1 were transiently transduced into cancer cells independently or together. RT-qPCR, Western blot, Cell Counting Kit-8 (CCK8), and Transwell assays were carried out to analyze the expression, cellular proliferation, and invasion. The results showed that the expression of miR-409 in breast cancer tissues is lower than that in adjacent tissues. The application of a target prediction algorithm predicts that the candidate gene regulated by miR-409 may be SATB1. The expression level of miR-409 in MDA-MB-453 cells is lower, while in BT-549 cells it is higher, when compared with MDA-MB-231, BR3, and MCF-7. The proliferation rate and invasive ability of MDA-MB-453 cells transfected with the miR-409 mimic was significantly lower than that of the miRNA negative control (miR-NC) cells, while the proliferation rate and invasive ability of BT-549 cells transfected with the miR-409 inhibitor were significantly increased. Cell proliferation and invasion of miR-409 mimic and SATB1 co-transfected MDA-MB-453 cells increased compared with that of miR-409 mimic-transfected cells, while miR-409 inhibitor and siSATB1 co-transfected BT-549 cells showed the opposite result. All these results indicated that miR-409 regulates breast cancer proliferation and invasion by targeting SATB1 and might be a potential therapeutic target for the treatment of breast cancer.