Bilateral breast cancers that develop at similar times in an individual are likely to have been subjected to similar hormonal, environmental and genetic influences during tumourogenesis compared with metachronous tumours. As such, it is possible that tumour phenotype in synchronous bilateral breast cancer may display similar biological characteristics. The aim of this study was to identify phenotypic similarities between synchronous and metachronous bilateral breast cancers which may suggest a common origin. Thirty-three cases of synchronous and 46 cases of metachronous bilateral breast cancer that displayed similar tumour type were analysed for concordance in relation to various histological and immunohistochemical parameters. A higher level of concordance was demonstrated for synchronous cases with the highest level seen for oestrogen receptor. It is likely that this is related to similar tumourogenic pathways occurring at equivalent exposure times to various environmental and hormonal influences, although, in a proportion of cases, inherited genetic factors may play a role.

The epidermal growth factor receptor (EGFR) family plays an important role in breast carcinogenesis. Much interest has been focused recently on its members because of their potential role as prognostic indicators in breast cancer and their involvement in cancer therapy. We have evaluated more than 1500 cases of invasive breast carcinoma immunohistochemically using tissue microarray technology to examine the expression of EGFR family receptor proteins. We have found that 20.1 and 31.8% of cases were positive for EGFR and c-erbB-2, respectively, and 45 and 45.1% of tumours overexpressed for c-erbB-3 and c-erbB-4, respectively. The expression of either EGFR or c-erbB-2 was associated with other bad prognostic features and with poor outcome. Neither c-erbB-3 nor c-erbB-4 had any association with survival. c-erbB-2 had an independent prognostic effect on overall and disease-free survival (DFS) in all cases, as well as in the subset of breast carcinoma patients with nodal metastases. Several hetero- and homodimeric combinations have been reported between the EGFR members. Those dimers can evoke diverse signal transduction pathways with variable cellular responses. We stratified cases according to their co-expression of receptors into distinct groups with different receptor-positive combinations. Patients whose tumours co-expressed c-erbB-2 and c-erbB-3, as well as those whose tumours co-expressed EGFR, c-erbB-2 and c-erbB-4 showed an unfavourable outcome compared with other groups, while combined c-erbB-3 and c-erbB-4 expression was associated with a better outcome. In cases showing expression of one family member only (homodimers), we found a significant association between c-erbB-4 homodimer-expressing tumours and better DFS. In contrast, patients with c-erbB-2 homodimer-expressing tumours had a significant poorer DFS compared with other cases. These data imply that the combined profile expression patterns of the four receptor family members together provide more accurate information on the tumour behaviour than studying the expression of each receptor individually.

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.

The diagnosis of metaplastic (sarcomatoid) carcinoma (MSC) of breast often requires immunohistochemistry with a cytokeratin (CK) panel to distinguish them from phyllodes tumors (PT), primary sarcomas, and fibromatoses. CK staining may be heterogeneous in metaplastic carcinomas. The aim of the study was to investigate the theory that MSCs show evidence of myoepithelial differentiation and to evaluate immunohistochemical markers that may be helpful in distinguishing MSCs from PT and fibromatosis. We reviewed histology and performed immunohistochemistry for AE1/AE3, 34betaE12, CK5 and CK14, Cam5.2, CK7 and CK19, epithelial membrane antigen (EMA) (B55), smooth muscle actin (SMA), S100, desmin, vimentin, CD31, CD34, and bcl-2 on paraffin-embedded tissue from 18 MSCs, 26 PTs, and 8 fibromatoses. We assessed staining by using a semiquantitative method. Sarcomatous areas in MSCs were positive for 34betaE12 in 11 cases; for SMA in 10; for CK5 in 7; for CK14 in 6; for Cam5.2, AE1/AE3, and S100 in 5; and for CK7 and CK19 in 3. No CK expression was seen in stromal areas in PT or in fibromatoses. CD34 and bcl-2 were more frequently expressed in spindle cell areas in PTs (18 and 12 of 26, respectively) than in MSCs (0 and 2 of 18, respectively). MSCs show strong evidence of myoepithelial differentiation. CD34 and, to a lesser extent, bcl-2 positivity in PTs may be helpful in differentiating these two lesions from MSCs, particularly in small biopsies, because CK staining in MSCs may be heterogeneous. In our hands, 34betaE12 was the CK most frequently expressed in sarcomatoid areas in MSCs.

p27kip1 is a member of the KIP/CIP family of cyclin-dependent kinase inhibitors and is a negative cell-cycle regulator that is thought to play a role in tumour suppression. Reduced levels of this protein have been observed in a number of human cancers. However, evidence is conflicting as to whether p27kip1 has a role to play in breast cancer, including predicting behaviour and prognosis. The present investigation aimed to provide a definitive study of 830 breast cancer cases with median patient follow-up of 104 months to determine the true prognostic significance, if any. Immunohistochemical analysis of tissue microarrays and three scoring methods were used to assess p27kip1 expression. Univariate analysis showed a significant relationship between reduced p27kip1 expression and increasing tumour grade, nuclear pleomorphism, mitosis, and decreasing tubule formation (all p<0.001). Significant associations between reduced p27, negative oestrogen receptor status, and ductal/no special type tumours were also observed. Survival analysis demonstrated that patients with tumours with high p27kip1 levels had an improved survival compared with those with cancers with low expression. On multivariate analysis, when compared with existing factors, p27kip1 was not, however, an independent prognostic factor. It is concluded that the inverse relationship between p27kip1 levels and histological grade and individual grade components suggests a role for p27kip1 in both cell proliferation and differentiation, but is not clinically useful.

BRCA1 is a tumour suppresser gene frequently mutated in familial breast cancer and thought to influence the progression of sporadic breast cancer. Decreased BRCA1 mRNA and protein expression has been identified in breast cancer cell lines and sporadic breast tumours. Here the prognostic significance of reduced BRCA1 protein expression is investigated in primary operable breast cancer. Immunohistochemical analysis was used to determine the level of BRCA1 protein expression in 100 breast cancers. BRCA1 expression was compared with known prognostic factors and survival to investigate its prognostic significance. BRCA1 nuclear expression was reduced by varying amounts in breast carcinomas. A progressive loss of BRCA1 expression correlated well with higher histological grade (p = 0.002) and an excess of medullary/atypical medullary/grade 3 ductal carcinomas (p = 0.0001). When adjusted for grade, patients with loss of BRCA1 expression had a significantly longer disease-free survival time. Loss of BRCA1 expression associated with high-grade breast tumours suggests that BRCA1 may play an important role in the pathogenesis of sporadic breast cancer.

Many studies to date have examined epidermal growth factor receptor (EGFR) expression but in relatively small numbers of breast cancer patients, with only short follow-up. We have investigated the correlation between EGFR expression and tumour characteristics, patient features and outcome in a series of women treated before 1980. 255 tumour samples were stained immunohistochemically with a mouse monoclonal. subclass IgGl antibody, E 30 raised against purified, denatured EGFR (Biogenex, CA). Results were classified according to the degree of tumour membrane staining and tumour cytoplasmic reactivity into scores 0. 1. 2. 3; 0 being no staining, lvery faint reactivity, 2distmct reactivity and 3strong reactivity. For analysis, 0 and 1 were regarded as negative, and 2 and 3 as positive. 56 (20%) tumours showed positive immunoreactivity for EGFR. An inverse relationship between expression of EGFR and ER status was seen (p < 0.001). EGFR was also statistically associated with histologic grade (p < 0.021). Lymph node disease, vascular invasion (p = 0.515). tumour size (p = 0.925). NPI, menopausal status (p = 0.216) and age (p = 0.456) were not related to EGFR expression. Life table analysis showed no correlation with disease free interval or overall survival. Despite longer-term follow-up and its association with histological grade, we have found no prognostic significance in EGFR expression in invasive breast cancer patients.

Epithelial cadherin (E-CD) is a member of the cadherin family of cell adhesion molecules and has been implicated as an invasion suppressor molecule in vitro and in vivo. We analysed 174 breast tumours from the Nottingham/Tenovus Breast Cancer Series immunohistochemically for E-CD expression using the mouse monoclonal antibody HECD-1 (Zymed Laboratories Inc.). In normal epithelial cells E-CD was strongly expressed at cell–cell boundaries. 66% of the breast cancers examined had reduced intensity of E-CD expression with 74% having significant reductions in the proportion of E-CD-positive tumour cells. Using a combined intensity/proportion score, significant associations were found between E-CD expression and tumour type (P ≤ 0.001). ER status (P = 0.026) and histological grade (P = 0.031). Expression of E-CD was not found to be related to recurrence, distant metastases, lymph node stage, vascular invasion, primary tumour size, prognostic group or survival. Thus E-CD expression in human breast cancer appears to have minimal prognostic value, but may have a role as a phenotypic marker. © 2001 Cancer Research Campaign http://www.bjcancer.com

Neuroendocrine differentiation and prognosis in breast adenocarcinoma Aims : Neuroendocrine differentiation has been detected, and its prognostic value studied, in a number of common human carcinomas. To date there are few detailed studies examining its relevance in breast carcinoma. In this study we evaluate the frequency and prognostic importance of neuroendocrine differentiation in breast adenocarcinoma. Methods and results : The presence of neuroendocrine differentiation, defined as positive reactivity for three markers, neuron‐specific enolase (NSE), chromogranin A and/or synaptophysin, has been evaluated in 99 patients with primary operable breast cancer using standard immunocytochemical techniques. A consecutive cohort of patients were selected from the Nottingham/Tenovus series. Comprehensive patient and tumour records have been maintained, and patients were followed up according to a defined protocol. Eighteen cases were positive for NSE, 10 for chromogranin A and 13 for synaptophysin. Eleven percent were positive with more than one neuroendocrine marker. No significant association was found between neuroendocrine differentiation and tumour size, grade, stage or the prevalence of vascular invasion. There was no significant difference in either overall or disease‐free survival between patients with or without neuroendocrine differentiation. Conclusions : In this study we confirm that neuroendocrine differentiation can be identified in a subset (10–18%) of human breast carcinomas. This phenomenon appears to have no relationship to established prognostic factors or patient outcome.

The Type 1 family of growth factor receptors includes epidermal growth factor receptor (EGFR), c-erbB-2, c-erbB-3 and c-erbB-4. Overexpression of the first two members is associated with poorer prognosis in patients with breast carcinoma. In this study we examined the expression of c-erbB-4 protein using the monoclonal antibody HFR-1. A total of 127 consecutive cases of primary operable invasive breast carcinoma presenting between 1975 and 1977 were studied. All patients were managed by simple mastectomy or conservation surgery with radiotherapy and no adjuvant therapy given. Long-term follow-up was maintained. Routine, formalin-fixed, paraffin-embedded tumour samples were used and sections were stained immunohistochemically using the Duet StreptABC method. Immunoreactivity was classified using a simple semi-quantitative scoring method. Protein expression was generally low but definite positive cytoplasmic, membranous and nuclear reactivity was identified in 58%, 41% and 25% of cases respectively. Expression at all three sites demonstrated significant inverse associations were histological grade. In addition, membrane accentuation correlated inversely with the Nottingham Prognostic Index (NPI), while cytoplasmic reactivity showed a positive association with c-erbB-3 expression. No significant associations were found with disease-free interval or survival. The results of this study demonstrate that higher levels of c-erbB-4 protein expression are associated with a more differentiated histological phenotype in contrast to the other members of the Type 1 family. Larger series with extended follow-up will be required to ascertain definitively the prognostic value of c-erbB-4 expression in breast carcinoma. © 2000 Cancer Research Campaign