Article
Methylation-specific multiplex ligation-dependent probe amplification and its possible impact on clinical findings in juvenile medulloblastomas
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Published: | May 21, 2013 |
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Objective: Gain of (proto-)oncogenes and loss of or promotor methylation of tumor suppressor gene (TSG) are considered to play an essential role in tumor genesis and progression. Methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) allows the simultaneous detection of both of these alterations. In a series of 20 medulloblastoma samples we used MS-MLPA as a screening for possible copy number changes in correlation with clinical and histological findings like age, sex, primary or recurrent tumor, CSF dissemination and the detection of necrosis and apoptosis as well as MRI findings, and the recurrence rate after one year.
Method: MS-MLPA was used in a series of 12 cryoconserved, and 8 paraffin embedded medulloblastoma samples. The three MS-MLPA probe sets used, could detect copy number changes in 77 unselected tumor suppressor genes and (proto-)oncogenes as well promotor methylation in a subset of 33 of these TSGs. For statistical analysis we used Mann-Whitney and exact Fisher’s test with p<0.05 as significant.
Results: Overall more gene amplifications (median 6.5) - than deletions (median 1.5) were found in the cryoconserved tumors. Furthermore, a median of 5.0 promotor hypermethylations per tumor were found. The most frequent single event was an amplification of ASC on 16p12 in 5 of the 12 tumors as well as gene amplifications on chromosome 17q (4/12 samples). A hypermethylation of MSH6 TSG on 2p16 was detected in 16 of all 20 specimen. There was a trend towards a negative correlation between the copy number alteration and the patients' age (coefficient of correlation R2=0.0132) as well as a direct correlation with a tumor cell CSF dissemination (p=0.055). Amplifications of the GATA5 gene were associated with a significant higher rate of tumor recurrences within one year (p=0.038), while hypermethylation of the CASP8 promotor were associated with a lower recurrence rate (p=0.036).
Conclusions: These findings confirm common aberrations like a gain on chromosome 17q but furthermore show possible new prognostic candidates on a genetic (like a gain of GATA5) and epigenetic level (like a hypermethylation of the CASP8 promotor). We conclude that a medulloblastoma-specific MS-MLPA probe set would be a valuable tool for both research and diagnostic approaches for this tumor entity.