Abstract 40: Developmental profile and regulation of the glycolytic enzyme hexokinase 2 and its association with aerobic glycolysis

Abstract

Abstract Proliferating embryonic and tumor tissues rely on aerobic glycolysis, or the metabolism of glucose to lactate under oxygenated conditions, to assist in the synthesis of biosynthetic precursors necessary for proliferation. The reliance on aerobic glycolysis may be mediated by the expression of specific metabolic enzymes. Mammalian Hexokinases HK1 and HK2 are 100kDa proteins that phosphorylate glucose to glucose-6-phosphate as the first step of the glycolytic pathway. In normal adult tissues, HK1 is ubiquitously expressed but is particularly prominent in the brain and kidney. HK2 is generally expressed at low levels within adipose tissue and skeletal tissue and negligently in normal brain. In this study, we wished to determine the ontogeny of these isoforms within the developing brain and determine how their expression relates to the extent of aerobic glycolysis in Glioblastoma Multiforme (GBM), a highly aggressive malignant brain tumor. Mining of existing published microarray data found strong expression of HK1 in 1, 2, 4 cell stage but a switch to stronger expression of HK2 in the blastocyst stage, previously reported to rely heavily on aerobic glycolysis. We performed HK2 immunohistochemistry in mice from gestational age E12, E16, post natal 1 month and 2 months. Our results demonstrated an age and cell specific HK1 and HK2 immunoreactivity in embryonic brain tissue with decreased expression of HK2 post-natally. Subsequently, we investigated the expression of HK1 and HK2 isoforms in a panel of GBM cell lines with varying levels of dependence on aerobic glycolysis, as measured by lactate levels and O2 consumption. HK2 but not HK1 expression was higher in GBM cells that had low O2 consumption and high extracellular lactate levels, supporting an association of HK2 with aerobic glycolysis. As HK2 expression is nearly silent in adult brain but expressed in fetal tissue and GBM cells, we hypothesized that DNA methylation/demethylation events may be playing important in its regulation. Adult normal human brain and GBM cell lines that had no HK2 expression were found to be methylated at CpG islands within intron 1 by bisulfite sequencing in contrast to fetal tissue and HK2-expressing GBM cells. The degree of methylation correlated with transcript expression in GBM cell lines. Treatment of U343 and A172 cell lines with 5-azacytidine restored HK2 transcript expression, supporting that HK2 maybe epigenetically regulated. Overall, our results demonstrate that the expression of the HK2 isoform, in contrast to HK1, may be particularly important in tissues relying on aerobic glycolysis for proliferation including embryonic tissue and GBMs. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 40.