Objective To analyze the significance and biological role of MAGEA6 expression in gastric cancer. Methods The tissue microarray samples of 90 cases of gastric cancer resected surgically in the hospital sample bank from December 2009 to June 2010 were collected. The expression of MAGEA6 in gastric cancer tissue microarray was detected by immunohistochemical staining, and the relationship between MAGEA6 expression and clinicopathological features of gastric cancer was analyzed.Kaplan-Meier analyzed the relationship between the expression of MAGEA6 and the prognosis of gastric cancer. BGC-823 gastric cancer cell line was cultured in vitro. Set si-MAGEA6 group (transfected with si-MAGEA6) and si-Ctrl group (transfected with vector), the cell proliferation ability and apoptosis rate were detected by CCK-8 and flow cytometry. Western blotting detected the expression of MAGEA6, apoptosis-related proteins [b lymphoma-2 gene (Bcl-2), bcl-2 related X protein (Bax)], autophagy-related proteins [microtubule associated protein light chain 3-Ⅱ (LC3-Ⅱ), p62 protein (p62), autophagy-related gene 5 (Atg5) protein, yeast Atg6 homolog (Beclin 1)], Akt/mTOR signaling pathway-related proteins [protein kinase B (Akt), phosphorylated protein kinase B (p-Akt), mammalian target of rapamycin (mTOR), phosphorylated mammalian target of rapamycin (p-mTOR)]. The autophagy flow and autophagosome formation were observed by laser confocal microscope and electron microscope. Results The immunohistochemical score of MAGEA6 in gastric cancer tissues was higher than that in adjacent tissues [(3.77±1.50) points vs.(2.58±1.11) points, P<0.05]. Patients with high expression of MAGEA6 were significantly related to age and TNM stage (P<0.05).The cell viability of gastric cancer cells in si-MAGEA6 group was lower than that in si-Ctrl group (P<0.05). The apoptosis rate of gastric cancer cells in si-MAGEA6 group was higher than that in si-Ctrl group (14.97%±0.86% vs. 4.63%±0.55%, P<0.05). The protein expression levels of MAGEA6, Bcl-2, p62, p-Akt, and p-mTOR in si-MAGEA6 group were lower than those in si-Ctrl group, the protein expression levels of Bax, Atg5, Beclin 1, and LC3-Ⅰ/LC3-Ⅱ were higher than those in si-Ctrl group (P<0.05). In the si-MAGEA6 group, autophagy bodies increased significantly, and autophagy bodies and lysosomes formed autophagy lysosomes. Conclusions Gastric cancer tissues showed a significantly increased level of MAGEA6. Silencing MAGEA6 expression inhibits the proliferation of gastric cancer cells, suggesting that MAGEA6 may be an effective biomarker and potential therapeutic target for gastric cancer.