Epigenetic gene silencing by aberrant DNA methylation leads to loss of key cellular pathways in tumorigenesis. In order to analyze the effects of DNA methylation on prostate cancer, we established LNCaP-derived human prostate cancer cells that can pharmacologically induce global reactivation of hypermethylated genes by the methyl-CpG targeted transcriptional activation (MeTA) method. The MeTA suppressed the growth of LNCaP-derived cells and induced apoptosis. Microarray analysis indicated that PYCARD (PYD and CARD domain containing) encoding an apoptosis-inducing factor was upregulated by 65-fold or more after treatment with MeTA. We analyzed DNA methylation statuses using 50 microdissected primary prostate cancer tissues and found an extremely high frequency of tumor-specific promoter hypermethylation of PYCARD (90%, 45/50). Moreover, DNA methylation status was significantly associated with Gleason score (); the frequency of tumor-specific hypermethylation was 96% (44/46) in tumors with Gleason , whereas that in tumors with Gleason score 6 was 25% (1/4). Immunohistochemical analyses using these 50 cases indicated that only 8% (4/50) of cancerous tissues expressed PYCARD, whereas 80% (40/50) of corresponding normal prostate epithelial and/or basal cells expressed PYCARD. In addition, there was no relationship between PYCARD immunostaining and the Gleason score in cancerous tissue and surrounding normal tissue. Inducible expression of PYCARD inhibited cell proliferation by induction of apoptosis. These results suggest that aberrant methylation of PYCARD is a distinctive feature of prostate cancers with Gleason and may play an important role in escaping from apoptosis in prostatic tumorigenesis.

中文翻译：

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异常高甲基化介导的 PYCARD 抑制在格里森前列腺癌中极为常见

由异常 DNA 甲基化引起的表观遗传基因沉默导致肿瘤发生中关键细胞途径的丢失。为了分析 DNA 甲基化对前列腺癌的影响，我们建立了 LNCaP 衍生的人前列腺癌细胞，该细胞可以通过甲基-CpG 靶向转录激活 (MeTA) 方法在药理学上诱导高甲基化基因的全局重新激活。MeTA 抑制 LNCaP 衍生细胞的生长并诱导细胞凋亡。微阵列分析表明，编码凋亡诱导因子的PYCARD（含有 PYD 和 CARD 结构域）在用 MeTA 处理后上调了 65 倍或更多。我们使用 50 个显微解剖的原发性前列腺癌组织分析了 DNA 甲基化状态，发现肿瘤特异性启动子高甲基化的频率极高皮卡德(90%, 45/50)。此外，DNA 甲基化状态与 Gleason 评分显着相关。); 使用 Gleason 的肿瘤中肿瘤特异性高甲基化的频率为 96% (44/46)，而在 Gleason 评分为 6 的肿瘤中为 25% (1/4)。使用这 50 个病例的免疫组织化学分析表明，只有 8% (4/50) 的癌组织表达 PYCARD，而 80% (40/50) 的相应正常前列腺上皮和/或基底细胞表达 PYCARD。此外，在癌组织和周围正常组织中，PYCARD 免疫染色与 Gleason 评分之间没有关系。PYCARD 的诱导表达通过诱导细胞凋亡抑制细胞增殖。这些结果表明PYCARD 的异常甲基化是格里森前列腺癌的一个显着特征 并且可能在前列腺肿瘤发生中逃避细胞凋亡中起重要作用。