Re-engineering breast cancer metabolism using metformin and dichloroacetate
Open Access
- Author:
- Banskota, Samagya
- Area of Honors:
- Bioengineering
- Degree:
- Bachelor of Science
- Document Type:
- Thesis
- Thesis Supervisors:
- Dr. Anneke Blackburn, Thesis Supervisor
Dr. William O Hancock, Thesis Supervisor
Peter J Butler, Thesis Honors Advisor - Keywords:
- Cancer
Breast Cancer Metabolism
Metformin
Dichloroacetate - Abstract:
- Cancer cells have a unique metabolic phenotype, the Warburg effect which, causes a high rate of glycolysis for ATP generation. This results in increased lactate production and reduced mi- tochondrial oxidation of pyruvate. This unique metabolic profile is thought to be associated with the resistance of cancer cells to apoptosis. Dichloroacetate(DCA) and metformin are two generic drugs that have recently demonstrated nontoxic anti-neoplastic effect by directly tar- geting cancer metabolism. DCA is an inhibitor of pyruvate dehydrogenase kinase (PDK), that can reverse the Warburg effect by increasing the flux of pyruvate into the mitochondria while metformin is an anti-diabetic drug that can inhibit proliferation in different types of tumor by targeting complex I of electron transport chain (ETC) and indirectly activating AMP-activated protein kinase (AMPK). The purpose of this study was to characterize the effects of DCA and metformin individually and in combination in an in vitro model of V14 breast cancer cells. Here, we report that after 72 hours of 5mM DCA and 6mM metformin treatment alone, there were 50% and 60% less viable cells present. When 5mM DCA was combined with metformin, it enhanced metformin’s anti-tumor activity at a lower dose, 0.5mM, there was at least 4 fold decrease in viable cell number compared with metformin treatment alone. The robustness of this result was validated by linear regression and measurement error model. DCA and metformin, however did not induce apoptosis, as seen from caspases 3 and 7 activity. The ability of DCA to inhibit PDKs and that of metformin to block complex I of ETC was assessed by measur- ing extracellular lactate concentration. DCA reduced the extracellular lactate concentration by targeting more pyruvate into the mitochondria, while metformin treated cells compensated for impaired mitochondrial respiration by increasing glycolysis as seen with increased extracellular lactate concentration. ATP levels also decreased significantly after the combination treatment indicating that there was a severe disruption of energy homeostasis in the cells. The action of metformin on its targeted molecule, AMPK, was assessed by western blot. Our results demonstrated that metformin activated AMPK in a dose and time dependent manner. Treatment with DCA also activated AMPK. We report a new molecular target of DCA, AMPK. However, the mechanism of AMPK activation by DCA is not fully understood yet but there are strong evidences in the literature that it could be via Ca2+ signaling. Together, these data suggest that targeting two aspects of metabolism using dichloroacetate and metformin may be useful in the treatment of cancer.