Jarrard D F, Kinoshita H, Shi Y, Sandefur C, Hoff D, Meisner L F, Chang C, Herman J G, Isaacs W B, Nassif N
Department of Surgery, University of Wisconsin Medical School, University of Wisconsin Comprehensive Cancer Center, Madison 53792, USA.
Cancer Res. 1998 Dec 1;58(23):5310-4.
Androgen-independent metastatic prostate cancer is characterized by a heterogeneous loss of androgen receptor (AR) expression among tumor cells. In this study, we evaluate DNA hypermethylation as a potential transcriptional regulatory mechanism in AR-negative prostate cancer cell lines. Nucleotide sequence analysis demonstrates an approximately 15-kb CpG island in the AR gene that encompasses the transcription start site and exon 1. Using Southern blotting with methylation-sensitive restriction enzymes and methylation-specific PCR, we find aberrant methylation in the AR expression-negative cell lines Du145, DuPro, TSU-PR1, and PPC1. Incomplete methylation in the AR CpG island is also seen in normal female breast and ovarian tissues consistent with the inactivation of one X chromosome by hypermethylation. In contrast, prostate cancer cell lines LNCaP and PC3 express AR and are unmethylated. Normal prostate epithelial cell strains demonstrate no methylation. Exposure of AR-negative prostate cancer cell lines to 5-aza-2' deoxycytidine, a demethylating agent, induces the reexpression of AR RNA in DuPro and TSU-PR1. This reexpression is associated with a demethylation of this region. Prostate-specific antigen, an androgen-responsive gene, is also specifically induced in these lines after AR reexpression. Therefore, in vitro DNA methylation of the 5' CpG AR island may be associated with the loss of AR expression. Furthermore, our results demonstrate that treatment with demethylating agents may engender the reexpression and function of the androgen receptor in AR-negative cell lines.
雄激素非依赖性转移性前列腺癌的特征是肿瘤细胞之间雄激素受体（AR）表达的异质性缺失。在本研究中，我们评估DNA高甲基化作为AR阴性前列腺癌细胞系中一种潜在转录调控机制的可能性。核苷酸序列分析显示AR基因中有一个约15 kb的CpG岛，其包含转录起始位点和外显子1。使用对甲基化敏感的限制性内切酶进行Southern印迹分析以及甲基化特异性PCR，我们发现在AR表达阴性的细胞系Du145、DuPro、TSU-PR1和PPC1中存在异常甲基化。在正常女性乳腺和卵巢组织中也可见AR CpG岛的不完全甲基化，这与一条X染色体通过高甲基化失活一致。相比之下，前列腺癌细胞系LNCaP和PC3表达AR且未发生甲基化。正常前列腺上皮细胞株未显示甲基化。将AR阴性前列腺癌细胞系暴露于脱甲基剂5-氮杂-2'-脱氧胞苷后，可诱导DuPro和TSU-PR1中AR RNA的重新表达。这种重新表达与该区域的去甲基化有关。雄激素反应性基因前列腺特异性抗原在AR重新表达后也在这些细胞系中被特异性诱导。因此，5' CpG AR岛的体外DNA甲基化可能与AR表达缺失有关。此外，我们的结果表明，用脱甲基剂处理可能会使AR阴性细胞系中的雄激素受体重新表达并发挥功能。
