We found that (1) LNT-229 glioma cells expressed TKTL1, (2) TKTL1 expression was upregulated in hypoxia and depended on the presence of HIF-1α, (3) TKTL1 suppression was accompanied by a downregulation of PPP intermediate sedoheptulose 7-phosphate, by an elevated glucose turnover and higher lactate levels indicating accelerated glycolysis and by an increase in intracellular ROS under hypoxic conditions, and (4) TKTL1 knockdown facilitated hypoxia-induced cell death and lowered clonogenic survival following irradiation. Here, TKTL1 is linked to central nervous system cancer.