Abstract 3368: Down-regulation of LDH-A reduced lactate production and changed the expression of glycolysis-associated transporters

Abstract

Abstract In the clinic, tumors showing a pronounced Warburg effect (high lactate tumors) are associated with a higher risk of metastasis and a decreased probability of survival of patients. Therefore, manipulations of the glycolytic pathway may alter tumor cell metabolism and might be a way to influence tumors’ aggressiveness, e.g. by down-regulating intracellular lactate levels. To test for this hypothesis B16.SIY E12 murine melanoma cells were stably transfected with shRNA down-regulating LDH-A expression (Lac Low cells). As a control cell line B16.SIY E12 was transfected with a scrambled shRNA construct (Lac High cells). These cell lines were characterized regarding their metabolic properties in vitro, especially lactate production in medium supernatants, the extracellular acidification rates (ECAR) and oxygen consumption rates (OCR). Furthermore, the expression of glycolysis-associated transporters (GLUT1, GLUT3, MCT1, MCT4 and CD147) was determined at the protein level. In a first in vivo set-up ATP, lactate and glucose content of syngeneic experimental tumors were measured using induced metabolic bioluminescence imaging (imBI). Changing LDH-A expression induced differences in lactate production in the two tumor cell lines in vitro with Lac Low cells producing significantly less lactate in 24 h than Lac High cells. Accordingly, Lac Low cells revealed a significantly lower ECAR and a higher OCR than Lac High cells. The protein expression of glycolysis-associated transporters was also influenced by changes in LDH-A expression with Lac High cells showing a significantly higher protein expression of MCT1 and MCT4. The general expression of GLUT1 was unchanged (Western Blot) but membrane localization was higher in Lac High cells (immunofluorescence staining). Measuring metabolite content in experimental tumors of the same cell lines showed that the cells maintained their glycolytic properties with Lac Low tumors showing lower lactate content than Lac High tumors. In summary, down-regulation of LDH-A expression reduced lactate production and membrane localization of glycolysis-associated transporters. The fact, that these characteristic are maintained during the transition from in vitro to in vivo makes this model system useful for systematic studies on therapeutic manipulation of tumor glycolysis and associated pathways. Supported by the Deutsche Forschungsgemeinschaft KFO262, DE 835/2-1 (WMK) + KR1418/8-1 (MK) Citation Format: Henrike Schroer, Christian G. Fabian, Marina Kreutz, Kristina Goetze, Wolfgang Mueller-Klieser. Down-regulation of LDH-A reduced lactate production and changed the expression of glycolysis-associated transporters. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3368. doi:10.1158/1538-7445.AM2014-3368