Aberrant DNA methylation is deeply involved in the development and progression of human breast cancers, but its inducers and molecular mechanisms are still unclear. To reveal such inducers and analyze molecular mechanisms of methylation induction, animal models are indispensable. However, genes methylation-silenced in the animal models of breast cancer are unknown, and identification of inducers and analysis of mechanisms are hampered.

Here, to identify genes methylation-silenced in rat primary mammary carcinomas, we took a combined approach of methylated DNA immunoprecipitation (MeDIP)-CpG island (CGI) microarray and chemical genomic screening. MeDIP-CGI microarray analysis of a carcinoma cell line induced by 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) revealed that, among 4,680 promoter CGIs analyzed, 664 were methylated in the cell line. Sixty-two of the 664 genes were re-expressed by treatment of the cell line with 5-aza-2’-deoxycytidine and trichostatin A. An unmethylated status in normal mammary glands was confirmed for 31 of the 62 genes by MeDIP-CGI microarray analyses of a pool of 4 normal mammary glands. Among 13 primary mammary carcinomas induced by PhIP, five of the 31 genes were methylated in one carcinoma or more. Among 12 primary carcinomas induced by 7,12-dimethylbenz[a]anthracene, methylation of all the five genes was also observed. RT-PCR analysis revealed that one of the five genes had high expression in normal mammary glands, but low expression in carcinomas, showing that this gene was methylation-silenced in rat primary mammary carcinomas.

In humans, the gene was expressed in mammary epithelial cells that had unmethylated DNA, and was down-regulated in three cancer cell lines that had methylated DNA. The gene was methylated in about 9% of 80 primary human breast cancers, and it was suggested that aberrant DNA methylation of this gene was a common event between rat and human breast cancers.

This is the first study of genome-wide DNA methylation analysis using rat mammary carcinomas, and the methylation-silenced gene, along with the other methylated genes, is expected to be useful as a marker in identifying inducers of aberrant DNA methylation and analyzing its molecular mechanisms.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 141.