The post-translational modification known as O-GlcNAcylation is a highly dysregulated process in tumors, and a key contributor to malignant transformation. In contrast, after three decades since its discovery, very little has been revealed about this process in the immune system. With the prospect of targeting O-GlcNAcylation as tumor therapy, greater understanding of how it regulates immune responses in the context of the tumor microenvironment will be needed. Here, we discuss recent discoveries from which a picture is emerging that O-GlcNAcylation, in either tumors or in immune cells, could negatively impact overall antitumor immune responses. We propose that interference with O-GlcNAcylation thus holds promise for cancer treatment from both perspectives.