In situ detection of the hypermethylation-induced inactivation of the p16 gene as an early event in oncogenesis
In situ detection of the hypermethylation-induced inactivation of the p16 gene as an early event in oncogenesis
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DOI：
10.1073/pnas.96.22.12754
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发表时间：
1999-10-26
影响因子：
11.1
通讯作者：
Herman, JG
中科院分区：
文献类型：
--
作者：
Nuovo, GJ;Plaia, TW;Herman, JG
我们已经开发了一种技术，甲基化特异性PCR原位杂交（MSP-ISH），它允许在单个细胞中可视化特定DNA序列的甲基化状态。我们使用MSP-ISH监测肺癌和宫颈癌进展过程中p16肿瘤抑制基因异常甲基化的时间和后果。在原位病变和浸润性癌中，p16的高甲基化仅定位于肿瘤细胞，并且与p16蛋白表达的丧失相关。MSP-ISH使我们能够剖析宫颈癌中发生的p16高甲基化这一令人惊讶的发现。这种肿瘤与致癌的人乳头瘤病毒感染有关，该病毒表达一种蛋白质E7，使视网膜母细胞瘤（Rb）蛋白失活。因此，同时Rb和p16失活将不需要废除关键的细胞周期蛋白D-Rb途径。MSP-ISH揭示了p16超甲基化在与表达病毒E7转录物的细胞群分开的早期宫颈肿瘤细胞群中不均匀地发生。在晚期宫颈癌中，大多数细胞具有超甲基化的p16，缺乏p16蛋白，但不再表达E7，这些数据表明，p16失活被选为阻断细胞周期蛋白D的最有效机制。Rb信号通路在癌前病变演变为浸润性癌过程中的作用。这些研究表明，MSP-ISH是一个强大的方法来研究在肿瘤演变过程中的关键肿瘤抑制基因的异常甲基化的动力学。
We have developed a technique, methylation-specific PCR in situ hybridization (MSP-ISH), which allows for the methylation status of specific DNA sequences to be Visualized in individual cells. We use MSP-ISH to monitor the timing and consequences of aberrant hypermethylation of the p16 tumor suppresser gene during the progression of cancers of the lung and cervix. Hypermethylation of p16 was localized only to the neoplastic cells in both in situ lesions and invasive cancers, and was associated with loss of p16 protein expression. MSP-ISH allowed us to dissect the surprising finding that p16 hypermethylation occurs in cervical carcinoma. This tumor is associated with infection of the oncogenic human papillomavirus, which expresses a protein, E7, that inactivates the retinoblastoma (Rb) protein. Thus, simultaneous Rb and p16 inactivation would not be needed to abrogate the critical cyclin D-Rb pathway. MSP-ISH reveals that p16 hypermethylation occurs heterogeneously within early cervical tumor cell populations that are separate from those expressing viral E7 transcripts, In advanced cervical cancers, the majority of cells have a hypermethylated p16, lack p16 protein, but no longer express E7, These data suggest that p16 inactivation is selected as the most effective mechanism of blocking the cyclin D-Rb pathway during the evolution of an invasive cancer from precursor lesions. These studies demonstrate that MSP-ISH is a powerful approach for studying the dynamics of aberrant methylation of critical tumor suppressor genes during tumor evolution.