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Metformin Kills Breast Cancer Cells by Inducing NADH Accumulation, Pyruvate Depletion and Reductive Stress

AMP‐activated kinase (AMPK) is a central metabolic regulator activated by deficits in nutrient supply. The role of AMPK in cancer has been contentious with both oncogenic and tumor suppressive roles being proposed. Studies by our group indicated that in breast cancer, AMPK activation promotes tumor evolution towards more aggressive phenotypes. These studies, however, were heavily criticized based on the argument that metformin, a biguanidine drug used to treat diabetes, activates AMPK and kills breast cancer cells. Hence, we set up to test the alternative hypothesis that AMPK activation by metformin relieves metformin toxicity by alleviating energy deficits, which are severe enough to cause cell death despite of AMPK activation. To examine this possibility we used AMPK‐competent (MCF7ev) and AMPK‐deficient (MCF7shAMPK) MCF7 cells, which are a estrogen receptor positive breast cancer cell line. Our results showed that MCF7shAMPK are slightly more sensitive to metformin than parental cells particularly at higher doses (20 mM). In addition, we found that pyruvate suppresses metformin toxicity towards parental cells but not MCF7shAMPK. Combined with metabolomics studies that revealed significant AMPK‐dependent distortions in amino acid metabolism caused by metformin our results indicate that complex I inhibition with the accumulation of NADH are the likely causes of metformin toxicity. This is alleviated by pyruvate (a NADH oxidant) as long as AMPK is functional and activates NADH oxidation by lactate dehydrogenase. We also conclude that metformin may be best utilized to treat tumors deficient in AMPK activity.