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  • Title: Infiltrating leukocytes confound the detection of E-cadherin promoter methylation in tumors.
    Author: Lombaerts M, Middeldorp JW, van der Weide E, Philippo K, van Wezel T, Smit VT, Cornelisse CJ, Cleton-Jansen AM.
    Journal: Biochem Biophys Res Commun; 2004 Jun 25; 319(2):697-704. PubMed ID: 15178462.
    Abstract:
    Promoter hypermethylation is known to result in transcriptional downregulation of many genes including the CDH1 gene. In this study we set out to determine CDH1 promoter methylation in breast tumors with decreased or absent E-cadherin protein expression and without CDH1 gene mutations by methylation-specific PCR (MSP). Interestingly, some tumor samples with normal E-cadherin expression yielded a methylation-specific PCR product. We hypothesized that other cells than tumor cells contribute to these products. Since in normal breast tissue no CDH1 promoter methylation is detected, infiltrating leukocytes, often present in tumors, might account for these methylation-specific fragments. Indeed, a methylation-specific fragment is found in all twelve leukocyte samples tested. Furthermore, activated T-cells also yielded a methylation-specific fragment. Sequencing of these fragments reveals two distinct methylation profiles. Leukocytes have only partial methylation of some CpGs, while the tumor-associated methylation profile shows complete methylation of most CpGs. Therefore, to assess whether CDH1 methylation is tumor associated, sequencing of MSP products is a prerequisite. Here we show that out of six lobular tumors lacking E-cadherin protein expression, three have tumor-associated CDH1 promoter methylation while in three other tumors no methylation is detected.
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