Downregulation of EphA5 by promoter methylation in human prostate cancer.
Downregulation of EphA5 by promoter methylation in human prostate cancer.
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人前列腺癌中启动子甲基化下调 EphA5
DOI：
10.1186/s12885-015-1025-3
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发表时间：
2015-01-22
期刊：
影响因子：
3.8
通讯作者：
Guan M
中科院分区：
文献类型：
--
作者：
Li S;Zhu Y;Ma C;Qiu Z;Zhang X;Kang Z;Wu Z;Wang H;Xu X;Zhang H;Ren G;Tang J;Li X;Guan M
背景EphA5是Eph/ephrin家族的成员，在癌发生的调节中发挥着关键作用。转录组分析显示，与低级别前列腺癌组织相比，高级别前列腺癌组织中的 EphA5 转录物显着减少。由于对 EphA5 下调的机制和 EphA5 的功能知之甚少，因此我们研究了前列腺癌中 EphA5 的表达和表观遗传变化，并确定这些发现是否与前列腺癌的临床病理特征相关。 方法七种前列腺细胞系（RWPE-1、LNCap、LNCap-LN3、CWR22rv-1、PC-3、PC-3M-LN4 和 DU145）、三十九种通过 qRT-PCR、甲基化特异性 PCR、亚硫酸氢盐测序、免疫组织化学和蛋白质印迹检查 BPH、22 例原发性前列腺癌、23 对非癌性和癌性前列腺组织。通过伤口愈合和transwell实验检查EphA5在前列腺癌细胞迁移和侵袭中的作用。结果在45个前列腺癌中的28个（62.2％）、39个增生中的2个（5.1％）以及所有6个前列腺癌细胞系中检测到EphA5 mRNA或蛋白表达下调或缺失。 EphA5 启动子区域的甲基化存在于 45 个癌症样本中的 32 个 (71.1%)、39 个增生样本中的 3 个 (7.7%) 以及 6 个前列腺癌细胞系中。在23对前列腺癌组织中，16个肿瘤样本表现出EphA5的高甲基化，这16个样本中的15个（93.8%）显示出EphA5表达比各自匹配的非癌样本下调。免疫染色分析表明，13 个可用癌样本中的 10 个 (76.9%) 中 EphA5 蛋白缺失或下调，并且这 10 个样本中的 8 个 (80.0%) 表现出高甲基化。在格里森评分或 T3-T4 分期较高的癌症患者中，EphA5 甲基化的频率较高。用 5-aza-2'-deoxycytidine 处理 6 种前列腺癌细胞系后，EphA5 mRNA 水平显着增加。体外EphA5的异位表达显着抑制了前列腺癌细胞的侵袭和迁移。结论我们的研究提供了证据，证明EphA5是原发性前列腺癌表观遗传沉默的潜在靶标，并且是前列腺癌潜在有价值的预后预测因子和治疗标记物。
BackgroundEphA5 is a member of the Eph/ephrin family and plays a critical role in the regulation of carcinogenesis. A significant reduction of EphA5 transcripts in high-grade prostate cancer tissue was shown using a transcriptomic analysis, compared to the low-grade prostate cancer tissue. As less is known about the mechanism of EphA5 downregulation and the function of EphA5, here we investigated the expression and an epigenetic change of EphA5 in prostate cancer and determined if these findings were correlated with clinicopathologic characteristics of prostate cancer.MethodsSeven prostate cell lines (RWPE-1, LNCap, LNCap-LN3, CWR22rv-1, PC-3, PC-3M-LN4, and DU145), thirty-nine BPH, twenty-two primary prostate carcinomas, twenty-three paired noncancerous and cancerous prostate tissues were examined via qRT-PCR, methylation-specific PCR, bisulfite sequencing, immunohistochemistry and western blotting. The role of EphA5 in prostate cancer cell migration and invasion was examined by wound healing and transwell assay.ResultsDownregulation or loss of EphA5 mRNA or protein expression was detected in 28 of 45 (62.2%) prostate carcinomas, 2 of 39 (5.1%) hyperplasias, and all 6 prostate cancer cell lines. Methylation of the EphA5 promoter region was present in 32 of 45 (71.1%) carcinoma samples, 3 of 39 (7.7%) hyperplasias, and the 6 prostate cancer cell lines. Among 23 paired prostate carcinoma tissues, 16 tumor samples exhibited the hypermethylation of EphA5, and 15 of these 16 specimens (93.8%) shown the downregulation of EphA5 expression than that of their respectively matched noncancerous samples. Immunostaining analysis demonstrated that the EphA5 protein was absent or down-regulated in 10 of 13 (76.9%) available carcinoma samples, and 8 of these 10 samples (80.0%) exhibited hypermethylation. The frequency of EphA5 methylation was higher in cancer patients with an elevated Gleason score or T3-T4 staging. Following the treatment of 6 prostate cancer cell lines with 5-aza-2′-deoxycytidine, the levels of EphA5 mRNA were significantly increased. Prostate cancer cells invasion and migration were significantly suppressed by ectopic expression of EphA5 in vitro.ConclusionOur study provides evidence that EphA5 is a potential target for epigenetic silencing in primary prostate cancer and is a potentially valuable prognosis predictor and thereapeutic marker for prostate cancer.
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