Extracts from Artemisia annua Linné (AAE) have various functions (anti-malaria, anti-virus, and anti-oxidant). However, the mechanism of the effects of AAE is not well known. Thus, we determined the apoptotic effects of AAE in AGS human gastric carcinoma cells. In this study, we suggested that AAE may exert cancer cell apoptosis through the Akt/mammalian target of rapamycin (mTOR)/glycogen synthase kinase (GSK)-3β signal pathway and mitochondria-mediated apoptotic proteins. Activation by Akt phosphorylation resulted in cell proliferation through phosphorylation of tuberous sclerosis complex 2 (TSC2), mTOR, and GSK-3β. Thus, de-phosphorylation of Akt inhibited cell proliferation and induced apoptosis through inhibition of Akt, mTOR, phosphorylation of GSK-3β at serine9, and control of Bcl-2 family members. Inhibition of GSK-3β attenuated loss of mitochondrial membrane potential and release of cytochrome C. Bax and pro-apoptotic proteins were activated by their translocation into mitochondria from the cytosol. Translocation of Bax induced outer membrane transmission and generated apoptosis through cytochrome C release and caspase activity. We also measured 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay, lactate dehydrogenase assay, Hoechst 33342 staining, AnnexinⅤ-PI staining, 5, 5'', 6, 6''-tetrachloro-1, 1'', 3, 3''-tetraethyl-imidacarbocyanine iodide (JC-1) staining, and Western blotting. Accordingly, our study showed that AAE treatment to AGS cells resulted in inhibition of Akt, TSC2, GSK-3β-phosphorylated, Bim, Bcl-2, and pro-caspase 3 as well as activation of Bax and Bak expression. These results indicate that AAE induced apoptosis via a mitochondrial event through regulation of the Akt/mTOR/GSK-3β signaling pathways.