Despite recent advances, the efficacy of androgen/androgen receptor (AR)-targeted therapy remains limited for many patients with metastatic prostate cancer. This is in part because prostate cancers adaptively switch to the androgen/AR-independent pathway for survival and growth, thereby conferring therapy resistance. Tumor hypoxia is considered as a major cause of treatment resistance. However, the exact mechanism is largely unclear. Here we report that chronic-androgen deprivation therapy (ADT) in the condition of hypoxia induces adaptive androgen/AR-independence, and therefore confers resistance to androgen/AR-targeted therapy, e.g., enzalutamide. Mechanistically, this is mediated by glucose-6-phosphate isomerase (GPI), which is transcriptionally repressed by AR in hypoxia, but restored and increased by AR inhibition. In turn, GPI maintains glucose metabolism and energy homeostasis in hypoxia by redirecting the glucose flux from androgen/AR-dependent pentose phosphate pathway (PPP) to hypoxia-induced glycolysis pathway, thereby reducing the growth inhibitory effect of enzalutamide. Inhibiting GPI overcomes the therapy resistance in hypoxia in vitro and increases enzalutamide efficacy in vivo.