Non-invasive detection of 2-hydroxyglutarate and other metabolites in IDH1 mutant glioma patients using magnetic resonance spectroscopy

Abstract

Mutations of the isocitrate dehydrogenase 1 and 2 genes (IDH1 and IDH2) are commonly found in primary brain cancers. We previously reported that a novel enzymatic activity of these mutations results in the production of the putative oncometabolite, R(-)-2-hydroxyglutarate (2-HG). Here we investigated the ability of magnetic resonance spectroscopy (MRS) to detect 2-HG production in order to non-invasively identify patients with IDH1 mutant brain tumors. Patients with intrinsic glial brain tumors (n = 27) underwent structural and spectroscopic magnetic resonance imaging prior to surgery. 2-HG levels from MRS data were quantified using LC-Model software, based upon a simulated spectrum obtained from a GAMMA library added to the existing prior knowledge database. The resected tumors were then analyzed for IDH1 mutational status by genomic DNA sequencing, Ki-67 proliferation index by immunohistochemistry, and concentrations of 2-HG and other metabolites by liquid chromatography-mass spectrometry (LC-MS). MRS detected elevated 2-HG levels in gliomas with IDH1 mutations compared to those with wild-type IDH1 (P = 0.003). The 2-HG levels measured in vivo with MRS were significantly correlated with those measured ex vivo from the corresponding tumor samples using LC-MS (r (2) = 0.56; P = 0.0001). Compared with wild-type tumors, those with IDH1 mutations had elevated choline (P = 0.01) and decreased glutathione (P = 0.03) on MRS. Among the IDH1 mutated gliomas, quantitative 2-HG values were correlated with the Ki-67 proliferation index of the tumors (r ( 2 ) = 0.59; P = 0.026). In conclusion, water-suppressed proton ((1)H) MRS provides a non-invasive measure of 2-HG in gliomas, and may serve as a potential biomarker for patients with IDH1 mutant brain tumors. In addition to 2-HG, alterations in several other metabolites measured by MRS correlate with IDH1 mutation status.

Fig. 1

Magnetic Resonance Imaging and MRS. Top panel (A and B): Axial MRI scans of two anaplastic astrocytomas (WHO grade III) show focal regions of fluid-attenuated inversion recovery (FLAIR) hyperintensity corresponding to areas of tumor. Other than the surrounding edema related to the size of the tumors, pre-operative structural MRI imaging characteristics were generally indistinguishable between IDH1 mutant (A) and wild-type (B) gliomas of the same grade and histopathology. Voxels of interest for MR spectroscopic analysis were localized by two neuroradiologists. Spectral voxels were placed in the center of the area of solid tumor, excluding regions of probable necrosis or vasogenic edema. Comparison of representative MR spectra from IDH1 mutant gliomas (C) versus wild-type spectra (D). Note the extra peaks in the region of Glu/Gln/2-HG (centered at 2.25 ppm) that are increased in the IDH1mutant tumors, compared to the wild-type MR spectra

Fig. 2

2-hydroxyglutarate, Glu and Gln levels in IDH1 wild-type versus IDH1 mutant gliomas. Quantitative 2-HG levels were calculated ex vivo using LC–MS (A) and in vivo using MRS (B). Good correlation was found between 2-HG levels measured by in vivo MRS and ex vivo LC–MS of tumor samples from the same patients (P < 0.0001) (C). IDH1 mutant cases are highlighted in red. ***P < 0.0001; **P = 0.003

Fig. 3

Tumor grade does not correlate with 2-HG concentrations in IDH1 mutant gliomas. Surgically resected glioma tissues were tested by LC–MS for the concentration of 2-HG in confirmed cases of IDH1 mutated gliomas. No significant differences in quantitative 2-HG levels were found due to differences in histology (p < 0.05 by ANOVA test)

Fig. 4

Metabolite levels in IDH1 wild-type vs. IDH1 mutant gliomas. Levels of A N-acetyl-aspartate (NAA), B choline (Cho), C glutathione (GSH), and D Lactate (Lac) were determined by MRS and quantified using LC-Model software. Metabolite ratios are represented with respect to the total concentrations of Cr. *P = 0.01 for Cho and P = 0.03 for GSH. Red dots represent wild-type samples with artifactually detectable 2-HG on MRS and high lactate levels, showing the metabolic profile of the “false positive” cases

Fig. 5

Relationship between Ki-67 proliferation index and 2-HG concentration. For the mutant IDH1 tumors (n = 9), the Ki-67 index for each tumor sample was determined by immunohistochemistry and compared to 2-HG concentrations calculated by MRS analysis. Correlation coefficients were calculated and are shown with a linear regression line drawn (P = 0.026)