Mass spectrometry-based assay for the molecular diagnosis of glioma: concomitant detection of chromosome 1p/19q codeletion, and IDH1, IDH2, and TERT mutation status

Abstract

The World Health Organization recently revised the diagnosis of glioma, to integrate molecular parameters, including IDH mutations and codeletion (loss of heterozygosity; LOH) of chromosome arms 1p/19q, into the definitions of adult glioma histological subtypes. Mutations in the TERT promoter may also be useful for glioma diagnosis and prognosis. The integration of molecular markers into routine diagnosis requires their rapid and reliable assessment. We propose a MassARRAY (MS)-based test that can identify 1p/19q codeletion using quantitative SNP genotyping and, simultaneously, characterize hotspot mutations in the IDH1, IDH2, and TERT genes in tumor DNA. We determined the reliability of the MS approach testing 50 gliomas and comparing the MS results with those obtained by standard methods, such as short tandem repeat genotyping, array comparative genomic hybridization (array-CGH) and Fluorescence In Situ Hybridization (FISH) for 1p/19q codeletion and Sanger sequencing for hotspots mutations. The results indicate that MS is suitable for the accurate, rapid, and cost-effective evaluation of chromosome deletions combined with hotspot mutation detection. This MS approach could be similarly exploited in evaluation of LOH in other situations of clinical and/or research importance.

Combined Mutational and Clonality Analyses Support the Existence of Intra-Tumor Heterogeneity in Papillary Thyroid Cancer

“…At variance, case #36, harboring both BRAF and TERT mutations with NAF 50% and 80%, respectively, was polyclonal, and we hypothesized the presence of two different clones, one with clonal BRAF V600E mutation and one with TERT c.-124C>T mutation at high NAF. The finding of NAF > 55% for the TERT mutation likely indicates the loss of heterozygosity (LOH) of the wild type allele, as previously shown [36].…”

TERT Promoter Alterations in Glioblastoma: A Systematic Review

“…Tumor heterogeneity or the scarcity of tumor DNA due to difficulties in tumor collection may lead to a lack of sensitivity of this historical technique [ 24 , 25 , 26 , 27 ]. Alternative sequencing methods were recently developed to increase the mutation detection rate in cases of low mutant allele frequency (MAF); these methods include Droplet Digital PCR (ddPCR), mass-spectrometry-based tests [ 28 ], and next-generation sequencing (NGS). ddPCR techniques have a higher sensitivity than Sanger sequencing in the detection of IDH1 and TERTp mutations in glioma [ 29 , 30 , 31 ].…”

The Genetic Landscape of Human Glioblastoma and Matched Primary Cancer Stem Cells Reveals Intratumour Similarity and Intertumour Heterogeneity

“…The assessment of hotspot mutations in IDH1 , IDH2 , and TERT promoter was performed as previously described [ 18 ]. In brief, DNA was amplified by multiplexed primer mix targeting codon 132 and 172 of IDH1 and IDH2 , respectively, and position c.-124 and c.-146 of TERT promoter.…”