No abstract

Wilms tumour is a childhood tumour that arises as a consequence of somatic and rare germline mutations, the characterisation of which has refined our understanding of nephrogenesis and carcinogenesis. Here we report that germline loss of function mutations in TRIM28 predispose children to Wilms tumour. Loss of function of this transcriptional co-repressor, which has a role in nephrogenesis, has not previously been associated with cancer. Inactivation of TRIM28, either germline or somatic, occurred through inactivating mutations, loss of heterozygosity or epigenetic silencing. TRIM28-mutated tumours had a monomorphic epithelial histology that is uncommon for Wilms tumour. Critically, these tumours were negative for TRIM28 immunohistochemical staining whereas the epithelial component in normal tissue and other Wilms tumours stained positively. These data, together with a characteristic gene expression profile, suggest that inactivation of TRIM28 provides the molecular basis for defining a previously described subtype of Wilms tumour, that has early age of onset and excellent prognosis.

The placenta is a vital fetal exchange organ connecting mother and baby. Specialised placental epithelial cells, called trophoblasts, are essential for adequate placental function. Trophoblasts transform the maternal vasculature to allow efficient blood flow to the placenta and facilitate adequate nutrient uptake. Placental development is in part regulated by epigenetic mechanisms. However, our understanding of how DNA methylation contributes to human trophoblast differentiation is limited. To better understand how genome-wide methylation differences affect trophoblast differentiation, reduced representation bisulfite sequencing (RRBS) was conducted on four matched sets of trophoblasts; side-population trophoblasts (a candidate human trophoblast stem cell population), cytotrophoblasts (an intermediate progenitor population), and extravillous trophoblasts (EVT, a terminally differentiated population) each isolated from the same first trimester placenta. Each trophoblast population had a distinct methylome. In line with their close differentiation relationship, the methylation profile of side-population trophoblasts was most similar to cytotrophoblasts, whilst EVT had the most distinct methylome. In comparison to mature trophoblast populations, side-population trophoblasts exhibited differential methylation of genes and miRNAs involved in cell cycle regulation, differentiation, and regulation of pluripotency. A combined methylomic and transcriptomic approach was taken to better understand cytotrophoblast differentiation to EVT. This revealed methylation of 41 genes involved in epithelial to mesenchymal transition and metastatic cancer pathways, which likely contributes to the acquisition of an invasive EVT phenotype. However, the methylation status of a gene did not always predict gene expression. Therefore, while CpG methylation plays a role in trophoblast differentiation, it is likely not the only regulatory mechanism involved in this process.

BackgroundChildhood acute lymphoblastic leukaemia (ALL) is the most common malignancy in children. Despite high cure rates, side effects and late consequences of the intensive treatments are common. Unquestionably, the identification of new therapeutic targets will lead to safer, more effective treatments. We identified TES promoter methylation and transcriptional silencing as a very common molecular abnormality in childhood ALL, irrespective of molecular subtype. The aims of the present study were to demonstrate that TES promoter methylation is aberrant, to determine the effects of TES re-expression in ALL, and to determine if those effects are mediated via TP53 activity.MethodsNormal fetal and adult tissue DNA was isolated and TES promoter methylation determined by Sequenom MassARRAY. Quantitative RT-PCR and immunoblot were used to confirm re-expression of TES in ALL cell lines after 5’-aza-2’-deoxycytidine (decitabine) exposure or transfection with TES expression plasmids. The effects of TES re-expression on ALL cells were investigated using standard cell proliferation, cell death and cell cycle assays.ResultsIn this study, we confirm that the TES promoter is unmethylated in normal adult and fetal tissues. We report that decitabine treatment of ALL cell lines results in demethylation of the TES promoter and attendant expression of TES mRNA. Re-expression of TESTIN protein in ALL cells using expression plasmid transfection results in rapid cell death or cell cycle arrest independent of TP53 activity.ConclusionsThese results suggest that TES is aberrantly methylated in ALL and that re-expression of TESTIN has anti-leukaemia effects which point to novel therapeutic opportunities for childhood ALL.

Epigenetic abnormalities at the IGF2/H19 locus play a key role in the onset of Wilms tumor. These tumors can be classified into three molecular subtypes depending on the events occurring at this locus: loss of imprinting (LOI), loss of heterozygosity (LOH), or retention of imprinting (ROI). As IGF2 LOI is a consequence of aberrant methylation, we hypothesized that this subtype of Wilms tumors might display global abnormalities of methylation. We therefore analyzed the methylation status of satellite DNA, as a surrogate for global methylation in 50 Wilms tumor patients. Satellite methylation was quantified by a methylation-sensitive quantitative PCR. We confirmed hypomethylation of both satellite α (Sat α) and satellite 2 (Sat 2) DNA in Wilms tumor samples compared with normal kidney. In addition, we found that LOI tumors, unlike ROI or LOH ones, showed concordant hypomethylation of both Sat α and Sat 2 DNA. This would suggest that the LOI subtype of Wilms tumor, which unlike other subtypes results from an epimutation, has a global deregulation of methylation mechanisms.