Hypoxia-associated proteins are commonly expressed as a consequence of disturbances in microcirculation. However, the clinical relevance of the proteins has never been studied in primary central nervous system lymphoma (PCNSL). The expression of hypoxia-inducible factor 1α (HIF-1α) and its downstream proteins, vascular endothelial growth factor (VEGF) and glucose transporter 1 (GLUT-1), were evaluated in a central nervous system (CNS) lymphoma xenograft model and in human PCNSL tissue. In the CNS lymphoma xenograft model, the expression of HIF-1α, VEGF, and GLUT-1 co-localized in subsets of lymphoma cells adjacent to necrosis. In tumor specimens from 51 patients with PCNSL, positive HIF-1α staining was found in 26 patients (51.0%), positive VEGF in 30 (58.8%), and positive GLUT-1 in 17 (33.3%), and HIF-1α showed a significant correlation with VEGF (p < 0.05). However, no significant association was seen between hypoxia-associated protein positivity and unfavorable clinical characteristics. Thus, the results failed to show an association with shorter overall survival or time to progression, except that the percentage of lymphoma cells positive for GLUT-1 (>20%) was significantly associated with worse survival. In conclusion, hypoxia-associated proteins were expressed in PCNSL, suggesting a hypoxic microenvironment. However, the prognostic relevance of these proteins for PCNSL was not demonstrated in this study.