Abstract
The Warburg effect occurs in 90% of tumours and causes a high rate of glycolysis even in the presence of oxygen, resulting in increased lactate production and reduced mitochondrial oxidation of pyruvate. Glucose metabolites are diverted to anabolic processes as a consequence, reducing pyruvate oxidation, hyperpolarizing the mitochondrial membrane potential, causing apoptotic resistance. Dichloroacetate (DCA) is a drug that can reverse the Warburg effect by inhibiting the pyruvate dehydrogenase kinases (PDKs), promoting oxidative metabolism of pyruvate. We are investigating in breast cancer cells (a) the effects of DCA on cell growth, (b) factors governing DCA sensitivity and (c) if DCA can enhance apoptosis induced by 4-(N-(S-penicillaminylacetyl)amino) phenylarsonous acid (PENAO), a novel anti-mitochondrial agent. At 5 mM DCA (48 hr treatment) there were 3-40% less viable cells present in MDA-MB-231, MCF7, MDA-MB-468, MCF10AT1, and T-47D breast cancer cell cultures. Growth of MCF10A non-cancerous cells was not affected, showing DCA selectively targets cancer cells. The PDKs have different sensitivities towards DCA inhibition (PDK2>PDK4>PDK1>PDK3). To determine if PDKs governed DCA sensitivity, PDK expression was examined by western blotting. In T-47D cells the expression of PDK2 (Ki 0.2 mM) and low levels of PDK1 (Ki 1 mM) and 3 (Ki 8 mM) correlated with their high sensitivity to DCA treatment. In MCF7 and MDA-MB-468 cells, high expression of PDK3 (Ki 8 mM) may explain their relative insensitivity to DCA. Extracellular lactate was also reduced by 50% at 1 mM and 5 mM DCA in T-47D and MCF7 cells respectively after 24 hr, indicating reversal of the Warburg effect, correlating with the PDK profiles. Induction of PDK1 in MCF7 cells under hypoxia increased sensitivity to DCA, showing the PDK profile still correlated with DCA sensitivity. The ability of DCA to enhance apoptosis induced by PENAO was also examined. The IC50 for PENAO (48 hr) was 3-13 µM for MDA-MB-468, MDA-MB-231, T-47D, MCF7 and MCF10AT1 cells, whereas 12 µM reduced cell viability by only 7% in the non-cancerous MCF10A cells. When combined with 5 mM DCA, the IC50 of PENAO for all cancer cell lines decreased by 15-70%, while toxicity to MCF10A cells was not increased. To measure apoptosis, cells were stained with annexin V and sorted by FACS. Treatment for 48 hr with 5 mM DCA and 5 µM PENAO doubled the proportion of apoptotic cells compared to PENAO alone on T-47D and MDA-MB-231 cells. DCA alone did not inhibit growth or induce apoptosis of MDA-MB-231 cells, thus showing potentiation of apoptosis. We have shown that DCA reverses the Warburg effect, inhibiting growth and enhancing apoptosis. PDKs may be a useful biomarker in determining whether DCA alone will be effective against different tumour types.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3228. doi:1538-7445.AM2012-3228