Methylation of the androgen receptor promoter CPG island is associated with loss of androgen receptor expression in prostate cancer cells

Androgen-independent metastatic prostate cancer is characterized by a heter ogeneous lass of androgen receptor (AR) expression among tumor cells. In th is study, we evaluate DNA hypermethylation as a potential transcriptional r egulatory mechanism in AR-negative prostate cancer cell lines. Nucleotide s equence analysis demonstrates a similar to 1.5-kb CpG island in the AR gene that encompasses the transcription start site and exon 1, Using Southern b lotting dth methylation-sensitive restriction enzymes and methylation-speci fic PCR, we find aberrant methylation in the AR expression-negative cell li nes Du145, DuPro, TSU-PR1, and PPC1. Incomplete methylation in the AR CpG i sland is also seen in normal female breast and ovarian tissues consistent w ith the inactivation of one X chromosome by hypermethylation. In contrast, prostate cancer cell lines LNCaP and PC3 express AR and are unmethylated. N ormal prostate epithelial cell strains demonstrate no methylation. Exposure of AR-negative prostate cancer cell lines to 5-aza-2' deoxycytidine, a dem ethylating agent, induces the reexpression of AR RNA in DuPro and TSU-PR1. This reexpression is associated with a demethylation of this region. Prosta te-specific antigen, an androgen-responsive gene, is also specifically indu ced in these lines after AR reexpression. Therefore, in vitro DNA methylati on of the 5' CpG AR island may be associated with the loss of AR expression . Furthermore, our results demonstrate that treatment with demethylating ag ents may engender the reexpression and function of the androgen receptor in AR-negative cell lines.