Abstract 5429: The Warburg Effect: protons suck
Authors: Russell Shonagh , Xu Liping , Gillies Rober J. 
Published: July 2017

Aerobic glycolysis (the Warburg Effect) is a hallmark of cancer and is associated with local invasion and metastasis. This metabolic phenotype results in acidification of the microenvironment in solid tumors, which is responsible for many of the known sequelae. For example, systemic buffer therapy directly and specifically increases extracellular tumor pH and reduces spontaneous and experimental metastasis in vivo. Further, extracellular acidosis can be a potent inhibitor of anti-tumor immunity. Removal of glycolytically-derived acids requires the activity of proton transporting mechanisms, such as NHE, V-ATPase and CA-IX. While it is commonly believed that these transporters are responding to the demands imposed by increased glycolytic flux, an alternative hypothesis states that glycolytic flux is increased to satisfy the demand driven by these transporters. Hence, expression of transporters that drive extracellular acidosis would induce a more glycolytic phenotype. To test this, we have transfected lowly glycolytic and non-metastatic MCF-7 breast adenocarcinoma cells with two different proton transporters.

In the first model, we used the yeast plasma membrane proton ATPase 1 (PMA1); a Type 1 P-type ATPase. PMA1 was stably over-expressed in MCF7 cell line and over-expression of PMA1 transformed the cells into a more aggressive phenotype with increased glucose consumption, lactate production, and proton production. Phenotypically, the transfected clones had elevated rates of migration and invasion in vitro and increased metastasis in vivo. This experimental evidence supports the hypothesis that proton export alone could drive the Warburg phenotype. Hence, in this system, glucose is metabolized to replenish either the ATP expended or the exported hydrogen-ions.

In the second model we used CA-IX; an exofacial carbonic anhydrase that is highly expressed in invasive cancers and materially participates in acidifying the microenvironment. CA-IX is upregulated in multiple cancer types with minimal expression in normal tissue, and is therefore an attractive and biologically relevant therapeutic target. In vitro, we have observed

that many cells expressing a glycolytic phenotype have elevated CA-IX expression, leading us to postulate that these phenotypes are coupled. To test this, we stably overexpressed CA-IX in MCF7 cells and again this resulted in a stable, glycolytic phenotype with increased glucose consumption, lactate production, and proton production observed using assay kits and live cell metabolic phenotyping with the Seahorse Extracellular Flux Analyzer. Through this study we have shown that acidosis leads to a more aggressive behavior and that CA-IX expression alone can induce a glycolytic phenotype. Hence, we hypothesise this is due to the enzymatic activity of CA-IX “sucking in” glucose to replenish intracellular H+ that are exported by this mechanism.

Citation Format: Shonagh Russell, Liping Xu, Rober J. Gillies. The Warburg Effect: protons suck [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5429. doi:10.1158/1538-7445.AM2017-5429