PO-253 Characteristics of cellular respiration, glycolytic activity and related metabolic features in wild type and IDH1 mutant glioma cells

Abstract

IntroductionIDH mutations are expressed in 80% of low grade gliomas and secondary glioblastomas with a favourable prognosis comparing to non IDH mutant gliomas [According to WHO (2016)] classification, IDH1 mutation is an important marker to classify gliomas. Our main interest is to study the role of certain metabolic alterations in glioma models, especially mitochondrial functions, glycolytic activity, IDH1 mutations and its regulation.Material and methodsU87 MG, U373 MG and U251 MG and U251 MG IDH1 R132H (kind gift of Dr. W. Leenders) overexpressing cells were used in vitro. Cellular oxygen consumption, glycolytic activity, energy substrates oxidation were measured by Seahorse technique. Furthermore, the amount and the activity of several proteins related mTOR (mammalian target of rapamycin) and other metabolic pathways were studied. The proliferation and apoptosis rates of in vitro cell cultures were also studied. We analysed extra- and intracellular metabolite levels (citrate, succinate, fumarate, alfa-ketoglutarate, malate, 2-hydroxyglutarate – D-2-HG, glutamate) using LC-MS measurements. Addressing the question what are the sources of D-2-HG in IDH1 mutant U251 MG cells, we fed the cells with 13C-labelled energy substrates.Results and discussionsThe glycolytic activity and oxygen consumption rates of the four cell lines show differences. The wild type U251 MG cell line has higher glycolytic activity and lower basal respiration than its IDH1 mutant variant glioma cell line pair. Addition of D-2-HG (72 hour) to wild type U251 MG cells increased the basal respiration and decreased the glycolytic activity of U251 MG cells. Using various bioenergetic substrates, it has been shown that U251 MG cells can oxidise glutamine, glutamate and malate at significantly higher level than IDH1 mutant U251 MG cells. This may be due to the alternative use of glutamine, we found that glutamine is the main source of D-2-HG production in these IDH1 mutant glioma cells. Furthermore, differences were found in the expression pattern of proteins which regulate energy metabolism in the studied glioma cells.ConclusionBased on the results of the examined glioma cells, IDH1 mutant cells showed a lower glycolytic capacity, a higher respiration and altered glutamine metabolism, which could be therapeutic targets in the future.Supported by ÚNKP-17–2, ÚNKP-17–3, Richter Centenárium Foundation, Bolyai-590/2015 and STIA-KF-17.