Methylation Profile of Small Breast Cancer Tumors Evaluated by Modified MS-HRM

The DNA methylation profile of breast cancer differs from that in healthy tissues and can be used as a diagnostic and prognostic biomarker. Aim of this study: To compare the levels of gene methylation in small malignant breast cancer tumors (<2 cm), in healthy tissue, and in fibroadenoma, and to evaluate the effectiveness of the modified Methylation Sensitive-High Resolution Melting (MS-HRM) method for this analysis. Analysis was performed using the modified MS-HRM method. For validation, the methylation levels of five genes were confirmed by pyrosequencing. The main study group included 96 breast cancer samples and the control group included 24 fibroadenoma samples and 24 healthy tissue samples obtained from patients with fibroadenoma. Breast cancer samples were divided into two subgroups (test set and validation set). The methylation of the following 15 genes was studied: MAST1, PRDM14, ZNF177, DNM2, SSH1, AP2M1, CACNA1E, CPEB4, DLGAP2, CCDC181, GCM2, ITPRIPL1, POM121L2, KCNQ1, and TIMP3. Significant differences in the validation set of samples were found for seven genes; the combination of the four genes GCM2, ITPRIPL1, CACNA1E, DLGAP2 (AUC = 0.99) showed the highest diagnostic value based on logistic regression for all breast cancer samples. Our modified MS-HRM method demonstrated that small breast cancer tumors have a specific DNA methylation profile that distinguishes them from healthy tissues and benign proliferative lesions.