Aberrant cytoplasmic expression of the p16 protein in breast cancer is associated with accelerated tumour proliferation

The p16 protein plays an important role in the transition of cells into the G(1) phase of the cell cycle. We have studied the prevalence of p16 protein expression in breast carcinomas in a prospective series of 368 invasive and 52 non-invasive malignancies, as well as in 88 locally recurring tumours and three tumour cell lines. p16 protein expression was evaluated immunohistochemically on paraffin sections using monoclonal and polyclonal anti-p16 antibodies, and by immunoblotting of tumour cell suspensions. Tumour cell lines were also subjected to polymerase chain reaction-single strand polymorphism (PCR-SSCP) analysis and direct DNA sequencing. The results were compared with established prognostic parameters, DNA flow cytometry and p53 protein expression. In 33 (9%) invasive and two (4%) intraductal carcinomas, a cytoplasmic accumulation of the p16 protein was seen. These cases were characterized by poor histological grade of differentiation, loss of of oestrogen receptors and progesterone receptors and frequent overexpression of the p53 protein. In addition, breast carcinomas with aberrant p16 expression demonstrated a high proliferative activity, with median S-phase fractions 74% higher than in the control group and the median Ki67 fractions elevated to 75%. A genetic alteration of the p16 gene was not detectable in three analysed cell lines with cytoplasmic p16 expression applying PCR-SSCP and direct DNA sequencing. These results indicate that cytoplasmic accumulation of the p16 protein identifies a subset of highly malignant breast carcinomas with accelerated tumour proliferation and other unfavourable parameters in breast cancer. The described protein accumulation is apparently not caused by an alteration of the p16 gene.