Abstract 3538: Enhanced dependence on lipid metabolism is a cellular adaptation to acidic microenvironment
Authors: Pillai Smitha , Wojtkowiak Jonathan W. , Damaghi Mehdi , Gatenby Robert , Gillies Robert 
Published: July 2017

Abstract
Malignant tumors exhibit altered metabolism and consume higher levels of glucose compared to surrounding normal tissue, resulting in highly acidic microenvironment. Adaptation to acidic conditions is a pre-requisite for tumor cells to survive and thrive and to out-compete the stroma into which they invade. Acid adaptation is associated with chronic activation of autophagy as well as redistribution of the lysosomal proteins to the plasma membrane. These processes are major survival mechanisms adopted by tumor cells under acidic conditions. We have also observed that under acidic conditions, there is a rapid, reversible and robust increase in the accumulation of cytoplasmic lipid droplets (adiposomes) in a panel of breast cancer cells. Adiposomes are dynamic organelles that store neutral lipids surrounded by a shell of proteins and a phospholipid monolayer. Breast cancer cells when grown in acidic media accumulated adiposomes as revealed by nile red and perilipin-2 staining followed by confocal microscopy. The acid-induced lipogenic phenotype persists even when the cells are grown in de-lipidated serum, indicating that the source of lipids is de-novo and endogenous. When cells were treated with inhibitors of fatty acid synthesis such as TOFA, an inhibitor of Acetyl CoA Carboxylase or FAS inhibitor C75, adiposome formation at low pH was attenuated. Inhibition of either lipid anabolic or catabolic pathways was specifically cytotoxic in acid-adapted cells, but not in control cells where the FAS inhibitor, C75, is selectively toxic under acidic conditions. Additionally, when treated with etomoxir, an inhibitor of carnitine palmitoyltransferase 1, the rate limiting step in βeta-oxidation, acid adapted cells showed increased sensitivity. To investigate the role of de novo fatty acid synthesis further, we employed high resolution NMR spectroscopy to measure 13C enriched lactate isotopomers following metabolism of D-[1,2-13C] glucose. These analyses showed that glucose flux through the pentose phosphate pathway (PPP) was significantly (&gt;2.5 fold) higher in low pH exposed cells, compared to controls, representing a major shift in glucose metabolism from Embden Meyerhof to PPP, which results in increased production of NADPH, necessary for de novo lipid synthesis. Additional metabolic profiling using the Seahorse XF revealed that cells at low pH had higher rates of oxygen consumption (OCR) and that this was reversible. Further, we investigated the role of various acid sensing G-protein coupled receptors such as OGR1, TDAG8 and GPR4 in transducing the acid signal that results in the accumulation of lipid droplets. CRISPR/Cas9 mediated depletion of these receptors indicates that they might play a major role in inducing adiposome accumulation under acidic conditions. Taken together, increased dependence on lipid metabolism by cancer cells under acidic conditions reveals novel therapeutic vulnerabilities.
Citation Format: Smitha Pillai, Jonathan W. Wojtkowiak, Mehdi Damaghi, Robert Gatenby, Robert Gillies. Enhanced dependence on lipid metabolism is a cellular adaptation to acidic microenvironment [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 3538. doi:10.1158/1538-7445.AM2017-3538