Polypeptide N-acetylgalactosaminyltransferases (GalNAc-Ts), a group of isoenzymes that initiate mucin-type O-glycosylation, have been shown to mediate tumor growth and metastasis in various cancer types. However, data on the clinical significance and features of GalNAc-Ts remain scant. Here, we used Oncomine and The Cancer Genome Atlas (TCGA) databases to analyze the transcription and survival effect of GALNTs (N-acetylgalactosaminyltransferase genes) in pan-cancer. The data showed that the GALNTs were aberrantly expressed in various human cancers and significantly associated with patients' clinical outcomes. The expression of 13 GALNTs were correlated with prognosis in brain low grade glioma (LGG) patients. In addition, based on the expression profiles of GALNT family genes in TCGA-LGG dataset, we identified 2 molecular subtypes (cluster1/2) by consensus clustering and analyzed tumor heterogeneity. Our results demonstrated that cluster 2 group was associated with poor prognosis, CD8(+) T cells, macrophages and DCs infiltration, up-regulated expression of immune checkpoints, and higher tumor immune dysfunction and exclusion score, indicating that GalNAc-Ts might contribute to tumor immune escape. Furthermore, we employed LASSO regression and time-dependent ROC analysis to construct a GALNTs-related prognostic signature with the TCGA-LGG dataset, and then validated the signature using 2 external cohorts. Taken together, our study successfully developed a novel prognostic biomarker for LGG and provides a basis for personalized immunotherapy in brain cancer.