Tumorigenesis and Neoplastic Progression Nuclear Exclusion of TET 1 Is Associated with Loss of 5-Hydroxymethylcytosine in IDH 1 Wild-Type Gliomas

The recent identification of isocitrate dehydrogenase 1 (IDH1) gene mutations in gliomas stimulated various studies to explore the molecular consequences and the clinical implications of such alterations. The Cancer Genome Atlas Research Network showed evidence for a CpG island methylator phenotype in glioblastomas that was associated with IDH1 mutations. These alterations were associated with the production of the oncometabolite, 2-hydroxyglutarate, that inhibits oxygenases [ie, ten-eleven translocation (TET) enzymes involved in the oxidation of 5-methylcytosine to 5-hydroxymethylcytosine (5hmC)]. We investigated 60 gliomas for 5hmC presence, 5-methylcytosine content, TET1 expression, and IDH1 mutation to gain insight into their relationships on a histological level. Of gliomas, 61% revealed no immunoreactivity for 5hmC, and no correlation was observed between IDH1 mutations and loss of 5hmC. Interestingly, expression of TET1 showed remarkable differences regarding overall protein levels and subcellular localization. We found a highly significant (P 0.0007) correlation between IDH1 mutations and nuclear accumulation of TET1, but not with loss of 5hmC. Of 5hmC-negative gliomas, 70% showed either exclusive or dominant cytoplasmic expression, or no detectable TET1 protein (P 0.0122). Our data suggest that the loss of 5hmC is a frequent event in gliomas, independent of IDH1 mutation, and may be influenced by the nuclear exclusion of TET1 from the nuclei of glioma cells. (Am J Pathol 2012, 181: 675–683; http://dx.doi.org/10.1016/j.ajpath.2012.04.017) Glioblastoma (World Health Organization grade IV) is the most frequent and most malignant type of glioma. Because of poor response to radiation therapy and most forms of chemotherapy, glioblastomas have a dismal prognosis, with a median survival time of 12 months. Primary or de novo glioblastomas are characterized by multiple genetic alterations, commonly including loss of heterozygosity on chromosome 10, EGFR amplification and overexpression, CDKN2A deletion, and PTEN mutation. Only 5% of glioblastomas, the so-called secondary glioblastomas, develop through progression from preexisting astrocytomas of WHO grade 2 or 3. They typically lack EGFR amplification but frequently carry TP53 mutations. Recently, somatic mutations of the isocitrate dehydrogenase (IDH1/IDH2) genes have been reported in most secondary glioblastomas and diffuse astrocytic and oligodendroglial tumors as well. These mutations, however, are rare in primary glioblastomas. Interestingly, an IDH1 mutation appears to be associated with increased DNA methylation at 5=-CpG islands of multiple genes, the so-called CpG island methylator phenotype. An IDH1 mutation in cancer cells leads to increased levels of the oncometabolite, 2-hydroxyglutarate (2-HG), and to a decrease of -ketoglutarate ( -KG), inhibiting the activity of -KG–dependent oxygenases, which are involved in demethylation of genomic DNA and histone tails. The 2-oxoglutarate and FeII-dependent dioxygenases TET1 and TET2 (ten-eleven translocation) are responsible for the oxidation of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) and require -KG and oxygen. TET3 is involved in the oxidation of 5mC to 5hmC in zygotic paternal DNA after fertilization. The amount of