This study investigates the role of AMPK in regulating NADPH homeostasis to promote tumor cell survival under energy stress. AMPK activation during energy stress, such as glucose deprivation, is shown to extend cell survival by maintaining NADPH levels through alternative pathways when the pentose phosphate pathway (PPP) is impaired. Specifically, AMPK inhibits acetyl-CoA carboxylases (ACC1 and ACC2) to decrease NADPH consumption in fatty acid synthesis and increase NADPH production through fatty acid oxidation (FAO). Knockdown of either ACC1 or ACC2 compensates for AMPK activation, enhancing anchorage-independent growth and solid tumor formation in vivo. The findings suggest that AMPK's role in NADPH maintenance is critical for cancer cell survival under conditions of metabolic stress, such as glucose limitations and matrix detachment. This mechanism may explain why AMPK-deficient cells are resistant to oncogenic transformation and why patients with Peutz–Jeghers syndrome, who have inherited LKB1 deficiency, develop only benign tumors.This study investigates the role of AMPK in regulating NADPH homeostasis to promote tumor cell survival under energy stress. AMPK activation during energy stress, such as glucose deprivation, is shown to extend cell survival by maintaining NADPH levels through alternative pathways when the pentose phosphate pathway (PPP) is impaired. Specifically, AMPK inhibits acetyl-CoA carboxylases (ACC1 and ACC2) to decrease NADPH consumption in fatty acid synthesis and increase NADPH production through fatty acid oxidation (FAO). Knockdown of either ACC1 or ACC2 compensates for AMPK activation, enhancing anchorage-independent growth and solid tumor formation in vivo. The findings suggest that AMPK's role in NADPH maintenance is critical for cancer cell survival under conditions of metabolic stress, such as glucose limitations and matrix detachment. This mechanism may explain why AMPK-deficient cells are resistant to oncogenic transformation and why patients with Peutz–Jeghers syndrome, who have inherited LKB1 deficiency, develop only benign tumors.