Abstract
BackgroundIn this study we aimed to quantify tumor suppressor gene (TSG) promoter methylation densities levels in primary neuroblastoma tumors and cell lines. A subset of these TSGs is associated with a CpG island methylator phenotype (CIMP) in other tumor types.MethodsThe study panel consisted of 38 primary tumors, 7 established cell lines and 4 healthy references. Promoter methylation was determined by bisulphate Pyrosequencing for 14 TSGs; and LINE-1 repeat element methylation was used as an indicator of global methylation levels.ResultsOverall mean TSG Z-scores were significantly increased in cases with adverse outcome, but were unrelated to global LINE-1 methylation. CIMP with hypermethylation of three or more gene promoters was observed in 6/38 tumors and 7/7 cell lines. Hypermethylation of one or more TSG (comprising TSGs BLU, CASP8, DCR2, CDH1, RASSF1A and RASSF2) was evident in 30/38 tumors. By contrast only very low levels of promoter methylation were recorded for APC, DAPK1, NORE1A, P14, P16, TP73, PTEN and RARB. Similar involvements of methylation instability were revealed between cell line models and neuroblastoma tumors. Separate analysis of two proposed CASP8 regulatory regions revealed frequent and significant involvement of CpG sites between exon 4 and 5, but modest involvement of the exon 1 region.Conclusions/significanceThe results highlight the involvement of TSG methylation instability in neuroblastoma tumors and cell lines using quantitative methods, support the use of DNA methylation analyses as a prognostic tool for this tumor type, and underscore the relevance of developing demethylating therapies for its treatment.
Highlights
In this study we aimed to quantify tumor suppressor gene (TSG) promoter methylation densities levels in primary neuroblastoma tumors and cell lines
Overall mean TSG Z-scores were significantly increased in cases with adverse outcome, but were unrelated to global LINE-1 methylation
Similar involvements of methylation instability were revealed between cell line models and neuroblastoma tumors
Summary
In this study we aimed to quantify tumor suppressor gene (TSG) promoter methylation densities levels in primary neuroblastoma tumors and cell lines. A subset of these TSGs is associated with a CpG island methylator phenotype (CIMP) in other tumor types. A number of clinical features and tumor phenotypes influence the disease outcome; the most important are age at diagnosis, tumor stage, amplification of the MYCN. Hypermethylation of tumor suppressor gene promoters were shown to be frequent in neuroblastoma with possible prognostic implications [9,10]. DNA methylation most commonly refers to methylation of the C in a CpG dinucleotide motif; a modification of fundamental importance for epigenetic regulation [11]. DNA methylation is central in processes such as expressional regulation, parental imprinting, and
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