Biallelic Inactivation of hMLH1 by Epigenetic Gene Silencing, a Novel Mechanism Causing Human MSI Cancers

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 95; no. 15; pp. 8698 - 8702
• Main Authors: Veigl, Martina L., Kasturi, Lakshmi, Olechnowicz, Joseph, Ma, AiHong, Lutterbaugh, James D., Periyasamy, Sumudra, Li, Guo-Min, Drummond, James, Modrich, Paul L., Sedwick, W. David, Markowitz, Sanford D.
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences of the United States of America 21-07-1998; National Acad Sciences; National Academy of Sciences; The National Academy of Sciences
• Subjects: Adaptor Proteins, Signal Transducing; Alleles; Azacitidine - pharmacology; Base Sequence; Biological Sciences; Cancer; Carrier Proteins; Cell lines; Colorectal cancer; Deoxyribonucleic acid; DNA; DNA Methylation; DNA Primers; Epigenetics; Genes; Humans; Methylation; Microsatellite Repeats; Mutation; MutL Protein Homolog 1; Neoplasm Proteins - genetics; Neoplasms - genetics; Nuclear Proteins; Promoter regions; Promoter Regions, Genetic; Protein synthesis; Tumor cell line; Tumors
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.95.15.8698

Mutations of DNA mismatch repair genes, including the hMLH1 gene, have been linked to human colon and other cancers in which defective DNA repair is evidenced by the associated instability of DNA microsatellite sequences (MSI). Germ-line hMLH1 mutations are causally associated with inherited MSI colon cancer, and somatic mutations are causally associated with sporadic MSI colon cancer. Previously however, we demonstrated that in many sporadic MSI colon cancers hMLH1 and all other DNA mismatch repair genes are wild type. To investigate this class of tumors further, we examined a group of MSI cancer cell lines, most of which were documented as established from antecedent MSI-positive malignant tumors. In five of six such cases we found that hMLH1 protein was absent, even though hMLH1-coding sequences were wild type. In each such case, absence of hMLH1 protein was associated with the methylation of the hMLH1 gene promoter. Furthermore, in each case, treatment with the demethylating agent 5-azacytidine induced expression of the absent hMLH1 protein. Moreover, in single cell clones, hMLH1 expression could be turned on, off, and on again by 5-azacytidine exposure, washout, and reexposure. This epigenetic inactivation of hMLH1 additionally accounted for the silencing of both maternal and paternal tumor hMLH1 alleles, both of which could be reactivated by 5-azacytidine. In summary, substantial numbers of human MSI cancers appear to arise by hMLH1 silencing via an epigenetic mechanism that can inactivate both of the hMLH1 alleles. Promoter methylation is intimately associated with this epigenetic silencing mechanism.