Background: Mammary carcinoma is the most communal carcinoma of higher women mortality. Rehmanniae Radix (RR) a traditional Chinese medicine. Objectives: To examine the anti-proliferative and anti-apoptotic effect of RR plant extract on mammary cancer cells MCF-7 and its action on PI3K/AKT/mTOR and GSK3 β mechanisms. Materials and Methods: The anti-proliferative of plant extract of RR was calculated, and morphological changes were measured by 3-(4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide assay. ROS (dichloro-dihydro fluorescein diacetate), MMP (Rh-123), apoptosis by Propidium Iodide, 4' 6-diamidino-2-phenylindole, and Acridine Orange/Ethidium Bromide staining assays assessed cellular cell death in MCF-7 cells. Western blotting analyses exhibited both phosphorylated forms compared with total PI3K/AKT/mTOR and GSK3 β were recognized. Results: The inhibitory action of breast cancer cells that RR extracts designated in a concentration manner. The IC50 value of RR extract on MCF-7 cells at 150 μg/mL concentration. Furthermore, staining assays seemed that RR extract-treated at 150 and 200 μg/mL, ROS production and apoptosis were augmented, and MMP and cell development were meaningfully reduced compared to control cells. Western blot analysis severely reduced the expression of p-PI3K, p-AKT, p-mTOR, and p-GSK3 β, but all total was extremely expressed than control cells. Altogether, the initiation of programmed cell death and suppression of cell propagation by the action of RR extract through the PI3K/AKT/mTOR mechanism recommended the anti-proliferative action of RR. Conclusion: Our results gave an innovative place of anti-tumor effect of RR extract is a gifted therapeutic and preventive agent in mammary cancer cell treatment.