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2004 ACVP Student Poster Award Winners

2005 ACVP Student Poster Award Abstract

METHYLATION OF THE E-CADHERIN TUMOR SUPRESSOR GENE IN CANINE MAMMARY CANCER.
L.M. Gooding*, J.L. Ford, and L.J. Rush.
Department of Veterinary Biosciences, The Ohio State University, Columbus, OH.

Loss of E-cadherin expression in epithelial neoplasms correlates with increased tumor invasiveness, metastasis and poor prognosis in humans and animals. In humans this loss is attributed to mutations, deletions, and/or DNA cytosine methylation – an epigenetic modification associated with transcriptional inactivation. However, little is known about the mechanisms for loss of E-cadherin expression in canine tumors. We hypothesized that promoter methylation of E-cadherin contributes to loss of gene expression in canine mammary tumors and this loss is correlated with tumor progression. To test this hypothesis we measured E-cadherin protein levels in normal, benign and malignant mammary tissue by immunohistochemistry, and compared this to the levels of promoter methylation determined by bisulfite sequencing. In order to restrict our methylation analyses to the epithelial component of the mammary tissues, epithelial cells were obtained using laser capture microdissection. The DNA was extracted and modified with sodium bisulfite which selectively converts unmethylated cytosine to thymine while leaving methylated cytosine unchanged. Subsequent PCR, cloning and sequence analysis allows the determination of the methylation status of each cytosine in our target region. Sixty-eight potential methylation sites in the 5’ region of E-cadherin were analyzed in 11 mammary tissue samples. Most samples showed a mixture of methylated and unmethylated alleles. Although the malignant tumors demonstrated decreased E-cadherin by immunohistochemistry, there was no correlation between the presence of methylation and the amount of protein seen with immunohistochemistry. In addition, there was no correlation between the overall amount of methylation and tumor progression. Based on our preliminary findings, it appears that DNA methylation in the region we examined does not play a major role in the transcriptional regulation of E-cadherin.

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