title: Methylation-specific multiplex ligation-dependent probe amplification and its impact on clinical findings in medulloblastoma.
abstract: Gain of (proto-)oncogenes and loss or promoter hypermethylation of tumor suppressor genes (TSGs) play essential roles in tumorigenesis. Methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) allows simultaneous detection of both these alterations. MS-MLPA was performed on 20 medulloblastoma samples (n = 12 cryoconserved; n = 8 formalin-fixed paraffin-embedded, FFPE) in order to screen for copy number changes in 77 unselected TSGs and (proto-)oncogenes as well as for promoter hypermethylation in a subset of 33 TSGs. In all specimens, determination of promoter methylation status was possible, whereas robust data concerning copy number changes could be obtained on cryopreserved material only. We found a median of 1.5 deletions and 6.5 amplifications in the 12 cryopreserved medulloblastoma and a median of 5 promoter hypermethylation per tumor. Frequent copy number changes included amplification of ASC on 16p12 (5/12) and amplification of several adjacent genes on 17q (3/12) including IGFBP4. Hypermethylation of MSH6 on 2p16 was found in 16 samples. MS-MLPA findings were also correlated with clinical and histological characteristics. The number of promoter hypermethylation was significantly associated with presence of necrosis (p = 0.004). Tumors which recurred within 1 year were more likely to show amplification of the GATA5 gene (p = 0.038), while hypermethylation of CASP8 was associated with a lower tumor recurrence rate (p = 0.036). There was also a trend towards a correlation between total number of aberrations and CSF dissemination (p = 0.055). Our findings confirm frequent presence of certain aberrations and reveal novel candidates for improving prognosis based on genetic and epigenetic tumor features. A medulloblastoma-specific MS-MLPA probe set seems a potentially valuable tool for further investigations on larger sample series.
authors.affiliation: Department of Neurosurgery, Jena University Hospital, Friedrich-Schiller-University Jena, Erlanger Allee 101, 07747, Jena, Germany, denise.feierabend@med.uni-jena.de.
authors.forename: Denise
authors.initials: D
authors.lastname: Feierabend
authors.forename: Jan
authors.initials: J
authors.lastname: Walter
authors.forename: Susanne
authors.initials: S
authors.lastname: Grube
authors.forename: Christian
authors.initials: C
authors.lastname: Herbold
authors.forename: Christian
authors.initials: C
authors.lastname: Beetz
authors.forename: Rolf
authors.initials: R
authors.lastname: Kalff
authors.forename: Christian
authors.initials: C
authors.lastname: Ewald
mesh_terms: D000328:Adult; D000368:Aged; D002528:Cerebellar Neoplasms / Q000175:diagnosis* / Q000235:genetics*; D002648:Child; D002675:Child, Preschool; D002874:Chromosome Mapping; D056915:DNA Copy Number Variations / Q000235:genetics; D015254:DNA Modification Methylases / Q000235:genetics; D005260:Female; D016147:Genes, Tumor Suppressor; D006801:Humans; D007223:Infant; D018974:Insulin-Like Growth Factor Binding Protein 4 / Q000235:genetics; D008297:Male; D008527:Medulloblastoma / Q000175:diagnosis* / Q000235:genetics*; D008875:Middle Aged; D060885:Multiplex Polymerase Chain Reaction; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*
chemical_list: D018974:Insulin-Like Growth Factor Binding Protein 4; D015254:DNA Modification Methylases