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Basic and Translational Investigations |
Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, Braga, Portugal (M.V.-P., I.A., R.M.R.); Institute of Molecular Pathology and Immunology, University of Porto, Porto, Portugal (S.S., R.S.); Histopathology Department, Addenbrooke's Hospital, University of Cambridge, Cambridge, United Kingdom (B.M.-A.); Laboratory of Neuropathology, Hospital de Santa Maria, Institute of Molecular Medicine, Lisbon, Portugal (J.P.)
Address correspondence to Rui Manuel Reis, Life and Health Sciences Research Institute, School of Health Sciences, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal (rreis{at}ecsaude.uminho.pt).
Medulloblastoma is the most common malignant brain tumor in children. The presence of microsatellite instability (MSI) in brain tumors, particularly medulloblastomas, has not been properly addressed. The aim of the present study was to evaluate the role of MSI in medulloblastoma carcinogenesis. MSI status was determined in 36 patients using a pentaplex PCR of quasimonomorphic markers (NR27, NR21, NR24, BAT25, and BAT26). Methylation status of mismatch repair (MMR) genes was achieved by methylation-specific multiplex ligation-dependent probe amplification (MLPA). In addition, MutS homolog 6 (MSH6) expression was determined by immunohistochemistry. Mutations of 10 MSI target genes (TCF4, XRCC2, MBD4, MRE11, ATR, MSH3, TGFBR2, RAD50, MSH6, and BAX) were studied by pentaplex PCR followed by analysis with GeneScan 3.7 software. Mutation analysis of hotspot regions of β-catenin (CTNNB1) and BRAF (v-raf murine sarcoma viral oncogene homolog B1) oncogenes was performed by PCR single-strand conformation polymorphism analysis followed by direct sequencing. Among the 36 tumors, we found four (11%) cases with instability, one with high MSI and three with low MSI. Methylation analysis of MMR genes in cases presenting shifts on the MSI markers revealed mild hypermethylation of MSH6 in 75% of cases, yet MSH6 was expressed in all the tumors. The MSI target genes MBD4 (methyl-CpG binding domain protein 4) and MRE11 (meiotic recombination 11 homolog A) were mutated in two different tumors. No CTNNB1 or BRAF mutations were found. This study is the most comprehensive analysis of MSI in medulloblastomas to date. We observed the presence of MSI together with mutations of MSI target genes in a small fraction of cases, suggesting a new genetic pathway for a role in medulloblastoma development.
Key Words: medulloblastoma • microsatellite instability • mismatch repair • target genes
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