Researchers at The Ohio State University Comprehensive Cancer Center (OSUCCC) have identified minute genetic changes in normal colonic tissue that may signal later development of certain types of colon cancer. The changes, which occur through a process called hypermethylation, appear to be age-related and associated with a subset of colon cancer marked by microsatellite instability. The findings appear in the October 1 issue of Cancer Research.
Scientists have felt for some time that hypermethylation plays an important but poorly defined role in the development of cancer. Hypermethylation is a chemical process that essentially turns off the "brakes" of a cell, which normally keep it from growing out of control. Colon cancers marked by microsatellite instability (MSI+) represent about 15% of all colon cancer, and are structurally more vulnerable to environmental or epigenetic "hits," or changes, that may lead to carcinogenesis. In MSI+ colon cancer, hypermethylation is usually found in the promoter region of the MLH1 gene. Most previous work identifying patterns of hypermethylation has been done in cancerous tissue; this study is among the first to examine the phenomenon in normal tissue, and the first to use PCR in situ to examine the pattern of hypermethylation.
Led by Dr. Albert de la Chapelle, director of the Division of Human Cancer Genetics at OSUCCC, a team of investigators including Drs. Hidewaki Nakagawa, Gerard Nuovo, Emmanuel Zervos and Edward Martin, Jr., obtained tissue samples from 111 cases of MSI(-) colon cancer, 43 cases of MSI(+) cancer, and 10 cases of hereditary colon cancer and studied their normal colonic cells for methylation of MLH1. They also examined fresh tissue specimens from 17 patients with sporadic colon cancer who had recently undergone surgery at the Arthur G. James Cancer Hospital and Richard J. Solove Research Institute. Utilizing combined bisulfite restriction analysis, (COBRA), a chemical treatment that helps to isolate hypermethylated regions in a gene, and methylation-specific PCR in situ hybridization (MSP-ISH), the scientists analyzed the pattern of methylation throughout 133 of 164 available samples. Of 90 patients whose tumors were MSI(-), 20 percent revealed the presence of fully methylated alleles and 59 percent showed the presence of partial methylation. In contrast, in 33 patients who had MSI(+) tumors, 55 percent showed fully methylated alleles, and 7 percent revealed partially methylated alleles. This means that methylation is not an all-or-nothing phenomenon but can begin by affecting a few sites and then spread to cover the entire promoter region, resulting in a total turning off of the gene.
Through MSP-ISH, the researchers discovered methylation was present in a patchy pattern throughout the normal lining of the colon, but seemed to be particularly abundant in the lumen, or surface of the tissue as opposed to the crypts.
Additionally, the scientists found methylation to be an age-related phenomenon, discovering partial or full methylation in 15 of 34 or 44 percent of the tissue samples from patients younger than age 60, but in 20 of 24, or 83 percent of the samples from patients over age 80. This pattern appeared to be independent of the MSI status of the tumors. De la Chapelle says it is important to remember that colon cancer is a disease of the elderly. "If the increase in methylation can be causally linked to cancer predisposition, it might explain why this cancer is strongly age related."
De la Chapelle says the next logical step in further defining the role of methylation of MLH1 is to compare patterns in normal, healthy individuals to those from patients with colon cancer.
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