Stage-specific methylome screen identifies that NEFL is downregulated by promoter hypermethylation in breast cancer

Aberrant hypermethylation of promoter regions in specific genes is a key event in the formation and progression of breast cancers, and an increasing number of marker genes have been identified. However, few genes which show methylation change in accordance with the progression of breast cancer have been identified. To identify genes which consistently undergo promoter methylation alterations as the tumor develops from a benign to a malignant form, genome-wide methylation databases of breast cancer cell lines from stage I to stage IV were analyzed. Heatmap and cluster analysis revealed that the genome-wide methylation changes showed a good accordance with tumor progression. Seven out of 14,495 genes were found to be consistently increased alongside the promoter methylation level through the normal cell line to the cancer stage IV cell lines. NEFL, one of the in silico hypermethylated genes in cancer, showed hypermethylation and lower expression in the cancer cell line MDA-MB-231, as well as in cancer tissues (methylation, p<0.05; expression, p<0.01). The expression was restored by inducing demethylation of the promoter in MDA-MB-231 cells. Our findings may lend credence to the possibility of using tumor stage-specific alterations in methylation patterns as biomarkers for estimating prognosis and assessing treatment options for breast cancer.