Evidence of epigenetic changes affecting the chromatin state of the retinoic acid receptor β2 promoter in breast cancer cells

Retinoic acid (RA)-resistance in breast cancer cells has been associated with irreversible loss of retinoic acid receptor beta, RAR beta, gene expression. Search of the causes affecting RAR beta gene activity has been oriented at identifying possible differences either at the level of one of the RAR beta promoters, RAR beta 2, or at regulatory factors. We hypothesized that loss of RAR beta 2 activity occurs as a result of multiple factors, including epigenetic modifications, which can pattern RAR beta 2 chromatin state. Using methylation-specific PCR, we found hypermethylation at RAR beta 2 in a significant proportion of both breast cancer cell lines and primary breast tumors. Treatment of cells with a methylated RAR beta 2 promoter, by means of the DNA methyltransferase inhibitor 5-Aza-2'-deoxycytidine (5-Aza-CdR), led to demethylation within RAR beta 2 and expression of RAR beta indicating that DNA methylation is at least one factor, contributing to RAR beta inactivity. However, identically methylated promoters can differentially respond to RA, suggesting that RAR beta 2 activity may be associated to different repressive chromatin states. This supposition is supported by the finding that the more stable repressive RAR beta 2 state in the RA-resistant MDA-MB-231 cell line can be alleviated by the HDAC inhibitor, trichostatin A (TSA), with restoration of RA-induced RAR beta transcription. Thus, chromatin-remodeling drugs might provide a strategy to restore RAR beta activity, and help to overcome the hurdle of RA-resistance In breast cancer.