Lactate dehydrogenase-B is silenced by promoter hypermethylation in human prostate cancer
Lactate dehydrogenase-B is silenced by promoter hypermethylation in human prostate cancer
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DOI：
10.1038/sj.onc.1209262
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发表时间：
2006-05-01
期刊：
影响因子：
8
通讯作者：
Rehman, I.
中科院分区：
文献类型：
--
作者：
Leiblich, A.;Cross, S. S.;Rehman, I.
为了确定与前列腺癌转移相关的新候选基因，我们比较了它们的蛋白质组学特征。对低转移性人前列腺癌细胞系LNCaP及其高转移性变体LNCaP- ln3进行了二维凝胶电泳。LNCaP细胞中有一个主要的蛋白斑点（pI为5.9，分子量为37 kDa）， LNCaP- LN3细胞中没有，通过串联质谱分析鉴定为乳酸脱氢酶- B （LDHB）。此外，酶动力学分析和酶谱分析显示，LNCaP细胞的LDH酶活性高于LNCaP- ln3。亚硫酸盐修饰的DNA测序显示LNCaP- LN3细胞中启动子超甲基化，而LNCaP、Du145、PC3、CWR22和BPH45细胞中没有。用5′-氮胞苷处理LNCaP- LN3细胞引起LDHB转录物的重新表达。在组织中，LDHB启动子超甲基化在前列腺癌中发生的频率更高，为14/ 31(45%)，而邻近的非恶性或良性组织为2/ 19 (11%)（P < 0.025）。免疫组化结果显示，LDHB在良性和非恶性组织中的表达频率为59/ 73(81%)，而在肿瘤组织中的表达频率为3/ 53 (6%)（P < 0.001）。在7/ 7（100%）的骨转移癌中，LDHB表达缺失。我们的数据首次显示了前列腺癌中LDHB表达的缺失，其机制似乎与启动子超甲基化有关。
In order to identify novel candidates associated with prostate cancer metastasis, we compared the proteomic pro. le of the poorly metastatic human prostate cancer cell line LNCaP, with its highly metastatic variant LNCaP-LN3, by two- dimensional gel electrophoresis. A major protein spot ( pI of 5.9 and molecular weight of 37 kDa) was seen in LNCaP cells, but not in LNCaP- LN3 cells and was identified as lactate dehydrogenase- B ( LDHB), by tandem mass spectrometry. Furthermore, enzyme kinetic assays and zymography showed a higher LDH enzyme activity in LNCaP cells compared with LNCaP-LN3. Bisulphite- modified DNA sequencing showed promoter hypermethylation in LNCaP- LN3 cells but not in LNCaP, Du145, PC3, CWR22 or BPH45 cells. Treatment of LNCaP- LN3 cells with 5'- azacytidine caused re-expression of LDHB transcripts. In tissues, LDHB promoter hypermethylation occurred at a higher frequency in prostate cancer, 14/ 31 ( 45%), compared to adjacent nonmalignant or benign tissue, 2/ 19 ( 11%) ( P < 0.025). Immunohistochemistry showed a higher frequency of LDHB expression in benign or nonmalignant tissues, 59/ 73 ( 81%), compared to cancer cases, 3/ 53 ( 6%) ( P < 0.001). Absent LDHB expression was also seen in 7/ 7 ( 100%) cases of metastatic cancer in bone. Our data are the first to show loss of LDHB expression in prostate cancer, the mechanism of which appears to involve promoter hypermethylation.