Conflicting results on the prevalence of cyclin D1 ovexpression and its correlation with CCND1 amplification and outcome of breast cancer patients have been reported. Owing to limited sensitivity and specificity of most antibodies against cyclin D1, evaluation of cyclin D1 immunoexpression is reported to be problematic. The aims of this study were to assess the prevalence of cyclin D1 expression in breast carcinomas using the SP4 rabbit monoclonal antibody; to correlate cyclin D1 expression with amplification, assessed using chromogenic in situ hybridisation (CISH); and to analyse the relationship between CCND1 amplification and overexpression with clinicopathological parameters and outcome in a tissue microarray containing replicate tumour samples from 245 breast cancer patients. Immunohistochemistry for cyclin D1 was performed using the SP4 and the results were scored according to the Allred scoring system. CISH was carried out using the Zymed CCND1 SpotLight probe. CISH signals were counted in 60 morphologically unequivocal neoplastic cells. Amplification was defined as >5 signals per nucleus in more than 50% of cancer cells, or when large gene copy clusters were seen. Strong cyclin D1 expression and CCND1 amplification were found in 67.4 and 14.5% of the cases, respectively. A strong correlation between cyclin D1 overexpression and CCND1 amplification was demonstrated (P<0.0001). Cyclin D1 expression showed a positive correlation with hormone receptor expression (both ER and PgR, P<0.0001). An inverse correlation was observed between an immunohistochemical panel of ‘basal-like' markers and both cyclin D1 overexpression (P<0.0001) and CCND1 amplification (P<0.0001). On univariate analysis cyclin D1 expression showed a correlation with longer overall survival (OS). Neither cyclin D1 nor CCND1 were independent prognostic factors for disease-free survival or OS. The results of this study confirm the association between cyclin D1 overexpression and positivity for hormone receptors and the lack of CCND1 amplification in basal-like breast carcinomas.
