DNA Methylation and Estrogen Receptor in Dog Mammary Tumors

Abstract

Estrogen receptor &#x3b1; (ER&#x3b1;) has an important role in mammary carcinogenesis, prognosis, and treatment. In human and canine mammary cancer, the most aggressive tumors show loss of ER&#x3b1; expression, which in human breast cancer has been attributed to methylation of the cytosine followed by guanine (CpG) island within the estrogen receptor &#x3b1; gene ( ESR1) promoter. This study aimed to investigate the role of ESR1 CpG island (CGI) methylation in ER&#x3b1; expression in canine mammary tumors. Twenty-one canine mammary samples were sorted into three groups: malignant tumor (n = 9), benign tumor (n = 8), and normal gland (n = 4). Immunohistochemical analysis and reverse-transcription quantitative real-time PCR were performed to assess ER&#x3b1; expression and ESR1 mRNA levels. The methylation status was determined using sodium-bisulfite-treated DNA sequencing. All normal mammary glands and benign tumors showed high ER&#x3b1; expression (score range, 5-8). Six of the nine malignant tumors did not show ER&#x3b1; expression (score 0), two had score 2, and one had score 4. Lower ER&#x3b1; ( P < .005) and ESR1 mRNA levels ( P < .005) were found in malignant mammary tumors than in the other two groups. Canine ESR1 has an intragenic and non-promoter-associated CGI, different from humans. No significant variation in methylation percentage was observed among the groups, suggesting that ESR1 is not regulated by DNA methylation, unlike that in humans. This difference should be considered in further research using ER&#x3b1; as a biomarker for mammary tumors in canine studies on ER&#x3b1;-targeting therapy.