Cunningham J M, Christensen E R, Tester D J, Kim C Y, Roche P C, Burgart L J, Thibodeau S N
Department of Laboratory Medicine and Pathology, Mayo Foundation, Rochester, Minnesota 55905, USA.
Cancer Res. 1998 Aug 1;58(15):3455-60.
Recent studies have demonstrated the presence of microsatellite instability (MSI) in tumors from patients with hereditary nonpolyposis colon cancer and in a subset of patients with sporadic colorectal cancer (CRC). In sporadic CRC, three tumor phenotypes have been defined: microsatellite stable (MSS), low-frequency MSI, and high-frequency MSI (MSI-H). Although defective mismatch repair, consisting primarily of alterations in hMSH2 and hMLH1, is believed to be responsible for the MSI phenotype in the majority of patients with hereditary nonpolyposis colon cancer, the genetic defect responsible for this phenotype in sporadic CRC has yet to be clearly delineated. Somatic or germ-line alterations in these two genes have been identified in only a minority of these cases. Analysis of the protein expression patterns of hMSH2 and hMLH1 in unselected CRC, however, suggests that alterations in hMLH1 may account for a majority of the MSI-H cases. In an effort to explore the underlying molecular basis for these findings, we have examined the methylation status of the presumptive hMLHI promoter region in 31 tumors that vary in regard to their MSI status (MSI-H or MSS), their hMLH1 protein expression (MLH- or MLH+), and their gene mutation (Mut+ or Mut-) status. Hypermethylation of the hMLH1 promoter occurred in all 13 MSI-H/ MLH- tumors that did not have a detectable mutation within the hMLH1 gene. Of those MSI-H tumors containing germ-line or somatic alterations in hMLH1 (n = 7, including 3 frameshift, 1 nonsense, 2 missense mutations, and 1 tumor containing multiple mutations: missense, splice-site alteration, and a frameshift), four had a normal methylation pattern, whereas three others demonstrated hypermethylation of the hMLH1 promoter region. Two of these cases had a missense alteration, the other a frameshift alteration. The single MSI-H/Mut+ tumor that had normal hMLH1 and hMSH2 expression, as well as 9 of the 10 MSS cases, lacked methylation of the hMLH1 promoter. Hypermethylation of the hMSH2 promoter was not observed for any of the cases. These results suggest that hypermethylation of the hMLH1 promoter may be the principal mechanism of gene inactivation in sporadic CRC characterized by widespread MSI.
近期研究已证实，遗传性非息肉病性结直肠癌患者的肿瘤以及部分散发性结直肠癌（CRC）患者的肿瘤中存在微卫星不稳定性（MSI）。在散发性CRC中，已定义了三种肿瘤表型：微卫星稳定（MSS）、低频MSI和高频MSI（MSI-H）。尽管主要由hMSH2和hMLH1改变组成的错配修复缺陷被认为是大多数遗传性非息肉病性结直肠癌患者MSI表型的原因，但散发性CRC中导致该表型的基因缺陷尚未明确界定。在这些病例中，仅少数病例发现了这两个基因的体细胞或种系改变。然而，对未经选择的CRC中hMSH2和hMLH1的蛋白质表达模式分析表明，hMLH1的改变可能占大多数MSI-H病例。为了探究这些发现的潜在分子基础，我们检测了31个肿瘤中假定的hMLHI启动子区域的甲基化状态，这些肿瘤在MSI状态（MSI-H或MSS）、hMLH1蛋白表达（MLH-或MLH+）以及基因突变（Mut+或Mut-）状态方面存在差异。hMLH1启动子的高甲基化发生在所有13个MSI-H/MLH-肿瘤中，这些肿瘤在hMLH1基因内未检测到突变。在那些hMLH1中含有种系或体细胞改变的MSI-H肿瘤（n = 7，包括3个移码突变、1个无义突变、2个错义突变以及1个含有多种突变的肿瘤：错义、剪接位点改变和移码突变）中，4个具有正常的甲基化模式，而另外3个则显示hMLH1启动子区域高甲基化。其中2例有错义改变，另一例有移码改变。唯一的MSI-H/Mut+肿瘤具有正常的hMLH1和hMSH2表达，以及10例MSS病例中的9例，均未检测到hMLH1启动子的甲基化。所有病例均未观察到hMSH2启动子的高甲基化。这些结果表明，hMLH1启动子的高甲基化可能是以广泛MSI为特征的散发性CRC中基因失活的主要机制。
