Methylation Patterns of , and Are Accompanied with Different Expression Levels in Human Astrocytoma.

In the present study, we investigated genetic and epigenetic changes and protein expression levels of negative regulators of Wnt signaling, , , and as well as glycogen synthase kinase 3 (GSK3β) and β-catenin in 64 human astrocytomas of grades II-IV. Methylation-specific PCR revealed promoter methylation of , , and in 38%, 43%, and 18% of samples, respectively. Grade IV comprised the lowest number of methylated cases and highest of . Evaluation of the immunostaining using H-score was performed for β-catenin, both total and unphosphorylated (active) forms. Additionally, active (pY216) and inactive (pS9) forms of GSK3β protein were also analyzed. Spearman's correlation confirmed the prevalence of β-catenin's active form (r = 0.634, < 0.001) in astrocytoma tumor cells. The Wilcoxon test revealed that astrocytoma with higher levels of the active pGSK3β-Y216 form had lower expression levels of its inactive form ( < 0.0001, Z = -5.332). Changes in exon 11 were observed in 44.44% of samples by PCR/RFLP. Astrocytomas with changes of had higher H-score values of total β-catenin compared to the group without genetic changes (t = -2.264, = 0.038). Furthermore, a positive correlation between samples with methylated promoter and the expression of active pGSK3β-Y216 (r = 0.356, = 0.011) was established. Our results emphasize the importance of methylation for the regulation of Wnt signaling. Large deletions of the gene associated with increased β-catenin levels, together with oncogenic effects of both β-catenin and GSK3β, are clearly involved in astrocytoma evolution. Our findings contribute to a better understanding of the etiology of gliomas. Further studies should elucidate the clinical and therapeutic relevance of the observed molecular alterations.