Methylation of CpG islands of p16INK4a and cyclinD1 overexpression associated with progression of intraductal proliferative lesions of the breast

SummaryP16INK4a is a tumor suppressor gene frequently inactivated by aberrant promoter hypermethylation. In this study, p16INK4a methylation was evaluated in intraductal proliferative lesions of the breast, using real-time quantitative polymerase chain reaction (MethyLight) and methylation-sensitive restriction endonuclease polymerase chain reaction. Immunohistochemistry was performed to compare and validate the methylation analysis. P16INK4a methylation associated with oncogene cyclinD1 expression, detected through the use of in situ hybridization and immunohistochemistry, was likewise characterized. P16INK4a methylation displayed varying significance among different types of intraductal proliferative lesions. Both the positive rate and the median quantitative methylation value increased with the evolution of intraductal proliferative lesions through the use of quantitative and qualitative assays. P16INK4a methylation was positively correlated to cyclinD1 overexpression. This study demonstrated that p16INK4a methylation served as the silencing mechanism of p16INK4a protein expression and played a crucial role in the intraductal proliferative lesions' progression. In the differential diagnosis of intraductal proliferative lesions, quantitative DNA methylation analysis of p16INK4a by MethyLight may be used as a surrogate, especially to distinguish atypical ductal hyperplasia from usual ductal hyperplasia and low-grade ductal carcinoma in situ. Furthermore, this study discovered that flat epithelial atypia do not share similar molecular profiles of p16INK4a epigenetic modification with atypical ductal hyperplasia and low-grade ductal carcinoma in situ.