Methylation of the thyroid-stimulating hormone receptor gene in epithelial thyroid tumors: a marker of malignancy and a cause of gene silencing.

Thyroid-stimulating hormone receptor (TSHR) expression is frequently silenced in epithelial thyroid cancers associated with decreased or absent TSH-promoted iodine uptake. To study the underlying molecular mechanism of decreased TSHR expression, we examined the methylation status of the TSHR gene promoter by sequencing bisulfite-treated DNA from thyroid tumors. After identification of methylated sites by sequencing bisulfite-treated DNA, we used methylation-specific polymerase chain reaction and found frequent CpG methylation in papillary thyroid cancer (23 of 39 patients; 59%) and follicular thyroid cancers (7 of 15 patients; 47%). In contrast, we saw no methylation in normal thyroid tissues and benign adenomas (0 of 8 patients; 0%). In human thyroid tumor cell lines, we observed that TSHR was normally expressed at the protein and mRNA level in cells where the TSHR gene was unmethylated, whereas it was silenced in cell lines where the TSHR promoter was hypermethylated. Treatment of the latter cells with a demethylating agent partially restored TSHR expression. We thus demonstrate aberrant methylation of human TSHR as a likely molecular pathway responsible for the silencing of this gene in thyroid cancers. We propose that methylation of TSHR may provide a novel diagnostic marker of malignancy and a basis for potential use of demethylating agents in conjunction with TSH-promoted radioiodine therapy for epithelial thyroid cancers.