Breaking the Hypoxic Code: Metformin as a Master Switch in Cancer Cell Metabolic Adaptation

Tumor hypoxia promotes metabolic reprogramming, epithelial-mesenchymal transition (EMT), and therapy resistance. Metformin, a widely used biguanide, has emerged as a candidate to counteract these adaptations, yet its phenotype-specific impact under stringent hypoxia remains unclear. Six human cancer cell lines reflecting oxidative (Pasteur-type; A549, MCF-7, SH-SY5Y) and glycolytic (Warburg-type; MDA-MB-231, MEL-JUSO, UPCI-SCC-131) metabolisms were exposed to chemical hypoxia induced by sodium sulfite. Dissolved-oxygen kinetics, resazurin viability, phase-contrast morphology, and immunofluorescence for HIF-1α/-2α, VEGF-A, GLUT1, HK2, LDH-A, E-cadherin, and N-cadherin were assessed in normoxia and hypoxia ± metformin (1 mM, 4 mM). A single 4 mM metformin dose prevented dissolved-oxygen levels from falling below the hypoxic threshold (< 10% DO) in all lines, whereas 1 mM was ineffective. Under normoxia, metformin significantly reduced viability in both the Pasteur-type cells and Warburg-type cells, to a similar extent (P < .0001), abolishing the Warburg survival advantage. Metformin reduced hypoxia-induced HIF-1α expression (2.2-fold) and significantly decreased VEGF-A, GLUT1, HK2, and LDH-A levels across cell lines, while HIF-2α showed a less consistent response. Concurrently, E-cadherin rose (1.6-fold) and N-cadherin declined (0.46-fold), consistent with attenuation of EMT-associated molecular features. Morphological stress features observed in hypoxia were attenuated, especially in hypoxia-sensitive lines. Metformin simultaneously limits oxygen consumption, suppresses HIF-1α-mediated glycolytic remodeling, and attenuates EMT-associated molecular changes, driving a phenotype-independent loss of cancer-cell viability under hypoxia. These findings position metformin as a metabolic gatekeeper capable of dismantling hypoxia-induced tumor resilience and warrant its integration into combination strategies targeting hypoxic niches.