Nie Yan, Liao Jie, Zhao Xin, Song Yunlong, Yang Guang-yu, Wang Li-Dong, Yang Chung S
Susan Lehman Cullman Laboratory for Cancer Research, Department of Chemical Biology, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, NJ 08854-8020, USA.
Carcinogenesis. 2002 Oct;23(10):1713-20. doi: 10.1093/carcin/23.10.1713.
Abnormal hypermethylation of CpG islands associated with tumor suppressor genes can lead to repression of gene expression and contribute significantly to tumorigenesis. Esophageal squamous cell carcinoma (ESCC) is thought to be developed through a multi-stage process, which involves basal cell hyperplasia (BCH), dysplasia (DYS), carcinoma in situ (CIS) and carcinoma. In the present study, we studied the hypermethylation of 10 selected genes in biopsies from normal individuals and resected tissues from ESCC patients. Tumor and neighboring normal and precancerous tissues including BCH, DYS and CIS were microdissected from the resected tissues by laser capture microdissection. Hypermethylation of CpG islands was examined in these samples for 10 genes: p16(INK4a), p15(INK4b), p14(ARF), human leukocyte antigen (HLA)-A, -B, -C, hMLH1, E-cadherin (E-cad), fragile histidine triad and von Hippel-Lindau (VHL). Methylation of two Alu sequences, which neighbor E-cad and VHL, respectively, was used as control to verify the procedure of DNA extraction and chemical modification. In 48 biopsy samples with BCH or DYS, the most frequent hypermethylated genes were p16(INK4a) (18.8%) and p14(ARF) (14.6%). Seventeen out of these 48 samples (35.4%) contained hypermethylation of at least one gene. In the resected tissues, 52% of the BCH and 81% of the tumors showed hypermethylation of at least one gene. Genes hypermethylated in earlier stage lesions were always found hypermethylated at the later stage lesions in the same patient. All of the genes were methylated at some stages and they were clustered into four groups according to their frequencies. The first group of genes, which consisted of p16(INK4a) and p14(ARF), was most frequently hypermethylated in all stages, and the frequencies increased from normal epithelial (0%) to BCH, to displasia/carcinoma in situ and ESCC. Other genes were hypermethylated less frequently. Our results suggest that hypermethylation of key genes, such as p16(INK4a), p14(ARF) and hMLH1, may be used in combination with other molecular changes, such as p53 mutation, in the development of biomarkers for predicting the risk for ESCC.
与肿瘤抑制基因相关的CpG岛异常高甲基化可导致基因表达受抑，并在肿瘤发生过程中发挥重要作用。食管鳞状细胞癌（ESCC）被认为是通过一个多阶段过程发展而来的，该过程包括基底细胞增生（BCH）、发育异常（DYS）、原位癌（CIS）和癌。在本研究中，我们研究了10个选定基因在正常个体活检组织和ESCC患者切除组织中的高甲基化情况。通过激光捕获显微切割从切除组织中显微切割出肿瘤、邻近的正常组织和癌前组织，包括BCH、DYS和CIS。检测了这些样本中10个基因的CpG岛高甲基化情况：p16（INK4a）、p15（INK4b）、p14（ARF）、人类白细胞抗原（HLA）-A、-B、-C、hMLH1、E-钙黏蛋白（E-cad）、脆性组氨酸三联体和冯·希佩尔-林道（VHL）基因。分别邻近E-cad和VHL的两个Alu序列的甲基化用作对照，以验证DNA提取和化学修饰过程。在48个BCH或DYS活检样本中，最常发生高甲基化的基因是p16（INK4a）（18.8%）和p14（ARF）（14.6%）。这48个样本中有17个（35.4%）至少有一个基因发生高甲基化。在切除组织中，52%的BCH和81%的肿瘤至少有一个基因发生高甲基化。在早期病变中发生高甲基化的基因在同一患者的后期病变中也总是高甲基化。所有基因在某些阶段都发生了甲基化，并根据其发生频率分为四组。第一组基因由p16（INK4a）和p14（ARF）组成，在所有阶段最常发生高甲基化，其频率从正常上皮组织（0%）到BCH、发育异常/原位癌和ESCC逐渐增加。其他基因发生高甲基化的频率较低。我们的结果表明，关键基因如p16（INK4a）、p14（ARF）和hMLH1的高甲基化可能与其他分子变化如p53突变联合用于开发预测ESCC风险的生物标志物。
