Melanoma-associated antigen (MAGE) family genes have been considered as potentially promising targets foranticancer immunotherapy. MAGED4 was originally identified as a glioma-specific antigen. Current knowledgeabout MAGED4 expression in glioma is only based on mRNA analysis and MAGED4 protein expression has notbeen elucidated. In the present study, we investigated this point and found that MAGED4 mRNA and protein wereabsent or very lowly expressed in various normal tissues and glioma cell line SHG44, but overexpressed in gliomacell lines A172,U251,U87-MG as well as glioma tissues, with significant heterogeneity. Furthermore, MAGED4protein expression was positively correlated with the glioma type and grade. We also found that the expressionof MAGED4 inversely correlated with the overall methylation status of the MAGED4 promoter CpG island.Furthermore, when SHG44 and A172 with higher methylation were treated with the DNA demethylating agent5-aza-2'-deoxycytidine (5-AZA-CdR) reactivation of MAGED4 mRNA was mediated by significant demethylationin SHG44 instead of A172. However, 5-AZA-CdR treatment had no effect on MAGED4 protein in both SHG44and A172 cells. In conclusion, MAGED4 is frequently and highly expressed in glioma and is partly regulated byDNA methylation. The results suggest that MAGED4 might be a promising target for glioma immunotherapycombined with 5-AZA-CdR to enhance its expression and eliminate intratumor heterogeneity.