Methylation analysis of the glypican 3 gene in embryonal tumours

We have previously shown that the glypican 3 (GPC3) gene was expressed in neuroblastoma (NB) and Wilms' tumour (WT), two embryonal tumours. GPC3 is an X-linked gene that has its peak expression during development and that is downregulated in all investigated tissues after birth. GPC3 expression could be involved in the aetiology of embryonal tumours such as NB and WT. Methylation is known to play a role in gene silencing, notably in chromosome X inactivation. Southern blot- and PCR-based methylation assays were used to assess the methylation status of the GPC3 promoter on genomic DNA from both normal and embryonal tumour cells. In normal cells, the promoter was not methylated in males and partially methylated in females. Our results suggest that DNA methylation of the promoter region is not essential for the transcriptional repression of the GPC3 gene and that the methylation observed in females is probably linked to the inactive X chromosome. In tumour samples, methylation abnormalities have been found exclusively in female NB samples (loss of methylation) and mainly in male WT samples (gain of methylation). Overall, methylation did not significantly correlate with the expression status of GPC3. Although promoter methylation is likely to affect the expression status of the gene, our results suggest that the deregulation of GPC3 transcriptional expression seen in NB and WT involves other regulatory levels.