In 1982 we constructed a prognostic index for patients with primary, operable breast cancer. This index was based on a retrospective analysis of 9 factors in 387 patients. Only 3 of the factors (tumour size, stage of disease, and tumour grade) remained significant on multivariate analysis. The index was subsequently validated in a prospective study of 320 patients. We now present the results of applying this prognostic index to all of the first 1,629 patients in our series of operable breast cancer up to the age of 70. We have used the index to define three subsets of patients with different chances of dying from breast cancer: 1) good prognosis, comprising 29% of patients with 80% 15-year survival; 2) moderate prognosis, 54% of patients with 42% 15-year survival; 3) poor prognosis, 17% of patients with 13% 15-year survival. The 15-year survival of an age-matched female population was 83%.

We have examined basal and luminal cell cytokeratin expression in 1944 cases of invasive breast carcinoma, using tissue microarray (TMA) technology, to determine the frequency of expression of each cytokeratin subtype, their relationships and prognostic relevance, if any. Expression was determined by immunocytochemistry staining using antibodies to the luminal cytokeratins (CKs) 7/8, 18 and 19 and the basal markers CK 5/6 and CK 14. Additionally, assessment of alpha-smooth muscle actin (SMA) and oestrogen receptor status (ER) was performed. The vast majority of the cases showed positivity for CK 7/8, 18 and 19 indicating a differentiated glandular phenotype, a finding associated with good prognosis, ER positivity and older patient age. In contrast, basal marker expression was significantly related to poor prognosis, ER negativity and younger patient age. Multivariate analysis showed that CK 5/6 was an independent indicator for relapse free interval. We were able to subgroup the cases into four distinct phenotype categories (pure luminal, mixed luminal/basal, pure basal and null), which had significant differences in relation to the biological features and the clinical course of the disease. Tumours classified as expressing a basal phenotype (the combined luminal plus basal and the pure basal) were in a poor prognostic subgroup, typically ER negative in most cases. These findings provide further evidence that breast cancer has distinct differentiation subclasses that have both biological and clinical relevance.

Recent studies on gene molecular profiling using cDNA microarray in a relatively small series of breast cancer have identified biologically distinct groups with apparent clinical and prognostic relevance. The validation of such new taxonomies should be confirmed on larger series of cases prior to acceptance in clinical practice. The development of tissue microarray (TMA) technology provides methodology for high-throughput concomitant analyses of multiple proteins on large numbers of archival tumour samples. In our study, we have used immunohistochemistry techniques applied to TMA preparations of 1,076 cases of invasive breast cancer to study the combined protein expression profiles of a large panel of well-characterized commercially available biomarkers related to epithelial cell lineage, differentiation, hormone and growth factor receptors and gene products known to be altered in some forms of breast cancer. Using hierarchical clustering methodology, 5 groups with distinct patterns of protein expression were identified. A sixth group of only 4 cases was also identified but deemed too small for further detailed assessment. Further analysis of these clusters was performed using multiple layer perceptron (MLP)-artificial neural network (ANN) with a back propagation algorithm to identify key biomarkers driving the membership of each group. We have identified 2 large groups by their expression of luminal epithelial cell phenotypic characteristics, hormone receptors positivity, absence of basal epithelial phenotype characteristics and lack of c-erbB-2 protein overexpression. Two additional groups were characterized by high c-erbB-2 positivity and negative or weak hormone receptors expression but showed differences in MUC1 and E-cadherin expression. The final group was characterized by strong basal epithelial characteristics, p53 positivity, absent hormone receptors and weak to low luminal epithelial cytokeratin expression. In addition, we have identified significant differences between clusters identified in this series with respect to established prognostic factors including tumour grade, size and histologic tumour type as well as differences in patient outcomes. The different protein expression profiles identified in our study confirm the biologic heterogeneity of breast cancer and demonstrate the clinical relevance of classification in this manner. These observations could form the basis of revision of existing traditional classification systems for breast cancer. ' 2005 Wiley-Liss, Inc.Key words: breast cancer; classification; protein expression; tissue microarray Routine clinical management of breast cancer relies on traditional histopathologic classification including tumour grade, histologic tumour type, carcinoma size and lymph node stage. Despite the overall association of these variables with prognosis and outcome, 1 these systems remain relatively weakly predictive of behaviour in some circumstances. Tumours of apparently homogenous morphologic character vary in response to therapy and have divergent outc...