gms | German Medical Science

Objective: Giant cell glioblastoma (gcGBM), characterized by the predominance of bizarre multinucleated giant cells, is a rare variant of GBM, accounting for less than 5%. They develop in younger patients de novo after a short clinical history. They tend to be well delineated and carry a better prognosis than GBM. GcGBM carry frequent TP53 mutations (>80%) and PTEN mutations (>30%), but EGFR amplification and IDH1/2 mutation (<10%) are rare. GcGBM show characteristics of primary and secondary GBM and therefore take a hybrid position. The aim of the present study was to further assess genetic alterations characterizing this rare variant.

Method: Briefly, genomic DNA (gDNA) was extracted from frozen tissue of five gcGBM and blood of one healthy control. Library preparation was performed, followed by exome enrichment and exome sequencing using Illumina HiSeq2000, according to the manufacturer’s protocols. Sequencing reads were aligned to reference human genome (hg19). For variant detection only reads with a Qscore >20 and a reading depth >10 were included. For validation, gDNA of the corresponding FFPE tissues was extracted and PCR was performed using primers designed for the specific sites of interest, followed by direct sequencing.

Results: Exome sequencing detected TP53 mutations (100%) and PTEN mutation (40%) in gcGBM, whereas no gcGBM carried an IDH1 mutation. We then searched for alterations in the same gene in at least two of five gcGBM. Candidate genes were only considered when databases like ToppGene showed evidence for a role in cancer. The potential functional impact of amino acid changes on the protein was assessed by bioinformatic mutation prediction tools, like SIFT and Polyphen2. After applying the above mentioned criteria, we found three candidate genes: TTN, PLXNA2 and PKHD1. TTN mutations and PKHD1 mutations were validated by direct sequencing in 2 gcGBM (40%). PLXNA2 mutations were confirmed in 3 gcGBM (60%).

Conclusions: In this study, common genetic alterations in gcGBM, like TP53 and PTEN mutations, were confirmed by exome sequencing. Additionally, three candidate genes with a frequency of at least 40% in gcGBM were identified and validated by direct sequencing. All mutations were predicted to be damaging mutations and to have a significant impact on the corresponding protein. Future studies are needed to screen for mutations in these genes in a larger number of gcGBM and to assess the function of these genes in gliomas and especially in gcGBM.