Abstract
Aim of studyMutations of isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2) gene were recently discovered in vast majority of World Health Organization (WHO) grade II/III gliomas. This study is to understand the effects of IDH1 R132H mutation in gliomagenesis and to develop new strategies to treat glioma with IDH1 R132H mutation.Materials and methodsOver expression of IDH1 R132H in U87MG cells was done by transfecting cells with IDH1 R132H plasmid. MTT assay, scratch repair assay and western blot were performed to study effects of IDH1 R132H mutation on cell proliferation, migration, regulating AKT-mTOR signaling pathway and cell death respectively. NADP+/NADPH and GSH quantification assays were performed to evaluate effects of IDH1 R132H mutation on the production of antioxidant NADPH and GSH.ResultsWe found that over expression of IDH1 R132H mutation decreased cell proliferation consistent with previous reports; however, it increased cell migration and enhanced AKT-mTOR signaling pathway activation. Mutations in isocitrate dehydrogenase (IDH) 1 also change the function of the enzymes and cause them to produce 2-hydroxyglutarate and not produce NADPH. We tested the level of NADPH and GSH and demonstrated that IDH1 R132H mutant stable cells had significantly low NADPH and GSH level compared to control or IDH1 wild type stable cells. The reduced antioxidants (NADPH and GSH) sensitized U87MG cells with IDH R132H mutant to 5-FU treatment.ConclusionOur study highlights the important role of IHD1 R132H mutant in up- regulating AKT-mTOR signaling pathway and enhancing cell migration. Furthermore, we demonstrate that IDH1 R132H mutation affects cellular redox status and sensitizes gliomas cells with IDH1 R132H mutation to 5FU treatment.
Highlights
Gliomas make up about 80% of all malignant brain tumors.[1]
We found that over expression of IDH1 R132H mutation decreased cell proliferation consistent with previous reports; it increased cell migration and enhanced AKT-mTOR signaling pathway activation
We tested the level of NADPH and GSH and demonstrated that IDH1 R132H mutant stable cells had significantly low NADPH and GSH level compared to control or IDH1 wild type stable cells
Summary
Gliomas make up about 80% of all malignant brain tumors.[1]. The exact causes of gliomas are not well known and it is believed that several oncogenes cooperate and contribute to the development of gliomas. [2] It was found that either isocitrate dehydrogenase (IDH) 1 or 2 genes mutations frequently occur in gliomas. [3] Isocitrate dehydrogenase (IDH) enzyme catalyzes the oxidative decarboxylation of isocitrate to produce α-ketoglutartate and at the same time use NADP+ as a cofactor to generate NADPH and maintain cellular redox status.[4]. [2] It was found that either isocitrate dehydrogenase (IDH) 1 or 2 genes mutations frequently occur in gliomas. [3] Isocitrate dehydrogenase (IDH) enzyme catalyzes the oxidative decarboxylation of isocitrate to produce α-ketoglutartate and at the same time use NADP+ as a cofactor to generate NADPH and maintain cellular redox status.[4] IDH1 mutations occurred in vast majority of World Health Organization (WHO) grade II/III gliomas and secondary glioblastomas. B. et al found that IDH1 R132H mutation in U87 cell line significantly decreased cell proliferation, accompanying changes in cell morphology and cell migration patterns. We are still at the infancy stage of understanding the role of IDH1 and IDH1 R132H mutation in gliomagenesis and further in-depth understanding of its molecular mechanisms in regulating cell proliferation and migration will be critical to develop future targeted therapy. We used multiple approaches to investigate the role of IDH1 and IDH1 R132H mutant in affecting cell proliferation, migration and major cell signaling pathway AKT-mTOR by stably overexpressing IDH1 either wild type or R132H mutant in U87MG cells or knocking down IDH1 by siRNA
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