Study of <b><i>hTERT</i></b> and Histone 3 Mutations in Medulloblastoma

Abstract

Hotspot activating mutations of the telomerase reverse transcriptase (hTERT) promoter region were recently described in several tumor types. These mutations lead to enhanced expression of telomerase, being responsible for telomere maintenance and allowing continuous cell division. Additionally, there are alternative telomere maintenance mechanisms, associated with histone H3 mutations, responsible for disrupting the histone code and affecting the regulation of transcription. Here, we investigated the clinical relevance of these mechanistically related molecules in medulloblastoma. Sixty-nine medulloblastomas, formalin fixed and paraffin embedded, from a cohort of patients aged 1.5-70 years, were used to investigate the hotspot mutations of the hTERT promoter region, i.e. H3F3A and HIST1H3B, using Sanger sequencing. We successfully sequenced hTERT in all 69 medulloblastoma samples and identified a total of 19 mutated cases (27.5%). c.-124:G>A and c.-146:G>A mutations were detected, respectively, in 16 and 3 samples. Similar to previous reports, hTERT mutations were more frequent in older patients (p < 0.0001), being found only in 5 patients <20 years of age. In addition, hTERT-mutated tumors were more frequently recurrent (p = 0.026) and hTERT mutations were significantly enriched in tumors located in the right cerebellar hemisphere (p = 0.039). No mutations were found on the H3F3A or HIST1H3B genes. hTERT promoter mutations are frequent in medulloblastoma and are associated with older patients, prone to recurrence and located in the right cerebellar hemisphere. On the other hand, histone 3 mutations do not seem to be present in medulloblastoma.

Reproducibility of the NanoString 22‐gene molecular subgroup assay for improved prognostic prediction of medulloblastoma

“…Hotspot h TERT somatic mutations have been reported in medulloblastomas with frequencies ranging 4–27% . In the present study, h TERT mutations were observed in 29% of the cases, and these mutations were more frequent in MB SHH as previously described Moreover, h TERT mutations were associated with older‐aged patients in accordance with previous studies .…”

“…Hotspot h TERT somatic mutations have been reported in medulloblastomas with frequencies ranging 4–27% . In the present study, h TERT mutations were observed in 29% of the cases, and these mutations were more frequent in MB SHH as previously described Moreover, h TERT mutations were associated with older‐aged patients in accordance with previous studies . Since h TERT ‐activating promoter mutations are reported to enhance telomerase expression, we included the h TERT gene in the CodeSet of the 22‐gene panel for medulloblastoma molecular subgrouping.…”

“…DNA was isolated from three to five sections each 10 μm thick using QIAmp DNA Mini kit (QIAgen) as previously reported . DNA concentrations were assessed by spectrophotometry (NanoDrop).…”

“…h TERT promoter hotspot mutations (−124 bp G > A and ‐146 bp G > A) were screened by PCR followed by Sanger sequencing as previously described . All samples with mutations were confirmed at least twice.…”

“…In addition to molecular signatures, gene alterations such as mutations in the promotor region of the gene for human telomerase reverse transcriptase (h TERT ) have been described in about 20% of medulloblastomas, and they correlate with a worse prognosis . Furthermore, the presence of h TERT promoter mutations is associated with a higher expression of h TERT gene in medulloblastomas, especially in non‐infant MB SHH.…”