Department of Otolaryngology-Head and Neck Surgery, Johns Hopkins Medical Institutions, Baltimore, MD, USA.
PLoS One. 2012;7(9):e45534. doi: 10.1371/journal.pone.0045534. Epub 2012 Sep 24.
Although promoter hypermethylation has been an accepted means of tumor suppressor gene inactivation, activation of otherwise normally repressed proto-oncogenes by promoter demethylation has been infrequently documented.
In this study we performed an integrative, whole-genome analysis for discovery of epigenetically activated proto-oncogenes in head and neck cancer tumors. We used the 47K GeneChip U133 Plus 2.0 Affymetrix expression microarray platform to obtain re-expression data from 5-aza treated normal cell line and expression data from primary head and neck squamous cell carcinoma (HNSCC) tumor tissues and normal mucosa tissues. We then investigated candidate genes by screening promoter regions for CpG islands and bisulfite sequencing followed by QUMSP and RT PCR for the best candidate genes. Finally, functional studies were performed on the top candidate gene.
From the top 178 screened candidates 96 had CpG islands in their promoter region. Seven candidate genes showed promoter region methylation in normal mucosa samples and promoter demethylation in a small cohort of primary HNSCC tissues. We then studied the demethylation of the top 3 candidate genes in an expanded cohort of 76 HNSCC tissue samples and 17 normal mucosa samples. We identified MAGEB2 as having significant promoter demethylation in primary head and neck squamous cell carcinoma tissues. We then found significantly higher expression of MAGEB2 in tumors in a separate cohort of 73 primary HNSCC tissues and 31 normal tissues. Finally, we found that MAGEB2 has growth promoting effects on minimally transformed oral keratinocyte cell lines but not a definite effect on HNSCC cell lines.
In conclusion, we identified MAGEB2 as activated by promoter demethylation in HNSCCand demonstrates growth promoting effects in a minimally transformed oral keratinocyte cell line. More studies are needed to evaluate MAGBE2's exact role in HNSCC.
虽然启动子甲基化已被认为是肿瘤抑制基因失活的一种手段，但通过启动子去甲基化激活原本受抑制的原癌基因的情况很少见。
在这项研究中，我们对头颈部癌症肿瘤中被表观遗传激活的原癌基因进行了综合的全基因组分析。我们使用 47K GeneChip U133 Plus 2.0 Affymetrix 表达微阵列平台，从 5-aza 处理的正常细胞系中获得再表达数据，并从原发性头颈部鳞状细胞癌（HNSCC）肿瘤组织和正常黏膜组织中获得表达数据。然后，我们通过筛选启动子区域中的 CpG 岛并进行亚硫酸氢盐测序，然后进行 QUMSP 和 RT-PCR，对候选基因进行筛选。最后，对最佳候选基因进行功能研究。
从筛选出的前 178 个候选基因中，有 96 个候选基因在其启动子区域有 CpG 岛。在正常黏膜样本中，有 7 个候选基因的启动子区域发生甲基化，而在一小部分原发性 HNSCC 组织样本中发生去甲基化。然后，我们在一个扩大的 76 个 HNSCC 组织样本和 17 个正常黏膜样本队列中研究了前 3 个候选基因的去甲基化情况。我们发现 MAGEB2 在原发性头颈部鳞状细胞癌组织中存在显著的启动子去甲基化。然后，我们在一个独立的 73 个原发性 HNSCC 组织和 31 个正常组织的队列中发现 MAGEB2 的表达显著升高。最后，我们发现 MAGEB2 对最小转化的口腔角质形成细胞系有促进生长的作用，但对 HNSCC 细胞系没有明确的作用。
总之，我们发现 MAGEB2 在 HNSCC 中被启动子去甲基化激活，并在最小转化的口腔角质形成细胞系中表现出促进生长的作用。需要更多的研究来评估 MAGBE2 在 HNSCC 中的确切作用。
