Axillary lymph node status of operable breast cancers by combined steroid receptor and HER-2 status: triple positive tumours are more likely lymph node positive.

AIMS: To examine the frequency of axillary lymph node (ALN) invasion of operable breast cancers by their combined oestrogen receptor (ER), progesterone receptor (PR) and HER-2 status. METHODS: 2227 recently operated cases in one centre were retrieved from the Multidisciplinary Breast Centre database and stratified according to their combined immunohistochemical (IHC) expression of ER/PR/HER-2 status. An equivocal HER-2 status was further analysed by Fluorescence in situ Hybridisation (FISH). The following 6 groups were considered: ER(-)PR(-)HER-2(-) (NNN; triple negative), ER(-)PR(-)HER-2(+) (NNP), ER(+)PR(-)HER-2(-) (PNN), ER(+)PR(-)HER-2(+) (PNP), ER(+)PR(+)HER-2(- )(PPN), ER(+)PR(+)HER-2(+) (PPP; triple positive). For ALN, the following variables were tested in uni- and multivariate models: age at diagnosis (years), tumour size (mm), tumour grade, ER, PR, HER-2 and the combined steroid receptor and HER-2 status. Likelihood ratio chi(2)-tests were used for univariate analysis and logistic regression for multivariate analysis. RESULTS: Triple positive tumours had a higher likelihood of being ALN positive than others (56.2% versus 35.7%; P<0.0001). Univariate logistic regression also withheld age, size, grade and HER-2 as predictors of ALN involvement. Final multivariate logistic regression revealed age, size, grade and PPP versus non-PPP to be independent predictors of ALN involvement; the odds ratio (OR) and 95% CI for PPP versus non-PPP tumours was 2.169 (1.490-3.156). CONCLUSION: Our data provide insight into the natural history of triple positive breast carcinomas. Such tumours are more likely ALN positive than those with another steroid receptor and HER-2 status. How these findings correlate with breast cancer prognosis remains to be investigated.

Short-term outcome of primary operated early breast cancer by hormone and HER-2 receptors.

INTRODUCTION: Prognostic subgroup classification of operable breast cancers using cDNA clustering of breast cancer-related genes resembles the classification based on the combined immunohistochemical (IHC) expression of the hormone and HER-2 receptors. We here report the short-term disease-free interval (DFI) of operable breast cancers by their joint hormone receptor/HER-2 phenotype. PATIENTS AND METHODS: Short-term follow-up (FU) of a prospective cohort of 1,958 breast-cancer patients primary operated at our institution between 2000 and 2005. Receptors were evaluated using IHC. Steroid receptors were considered positive for any nuclear staining; HER-2 for strong (3+) membrane staining or positive fluorescence in situ hybridization (FISH). Kaplan-Meier (KM) DFI curves were calculated for any relapse defined as a local, regional, contralateral, or distant breast cancer event for the six predefined breast cancer subgroups: ER + PR + HER-2 - (PPN), ER + PR - HER-2 - (PNN), ER + PR + HER-2 + (PPP), ER - PR - HER-2 - (NNN), ER - PR - HER-2 + (NNP), and ER + PR - HER-2 + (PNP). P-values were calculated for comparison of the six different survival curves using two possible adaptations for multiple testing. A multivariate model for the receptors predicting DFI did incorporate local and systemic adjuvant therapy. RESULTS: Median patient age was 57 years (ranges 26-96) and median FU was 3.35 years. Overall, DFI at median FU was 91%; 94% for PPN, 89% for PNN, 86% for NNN, 81% for PPP, 80% for PNP, and 76% for NNP cases. Some receptor subgroups had a significantly better DFI than others based on multiple testing, especially when the PPN group was compared against the four most frequent subtypes. The multivariate model with local and systemic adjuvant therapy confirmed the prognostic value of ER, PR, and HER-2 for short-term DFI. CONCLUSION: It is possible to distinguish short-term prognostic breast cancer subgroups only on the basis of ER, PR, and HER-2 even when stratified for local and systemic adjuvant therapy. While gene expression profiles based on microarray data of over hundreds of genes will probably teach us much about breast cancer biology, heterogeneity, and prognosis, we emphasize the important short-term prognostic value of currently used IHC markers for ER, PR, and HER-2.

Quantity and quality of retrograde menstruation: a case control study.

BACKGROUND: The purpose of this study was to test the hypothesis that menstruation is associated with a higher concentration of endometrial cells in peritoneal fluid(PF) and with increased white and red blood cell concentration in PF when compared to nonmenstrual phases of the cycle. METHODS: PF was obtained at laparoscopy from 107 women with endometriosis (n = 59) and controls with a normal pelvis (n = 48) during the luteal (n = 46), follicular (n = 38) or menstrual (n = 23) phase of the cycle. Endometriosis was classified according to the classification of the American Society for Reproductive Medicine (rAFS into minimal (n = 25), mild(n = 20), moderate(n = 6) and severe(n = 8) disease. Cell counts (leucocytes, erythrocytes, thrombocytes) were determined on a cell counter. In a subset of 32 patients (13 controls and 19 women with endometriosis), PF was fixed, processed and thinlayers were prepared and stained with Papanicolaou method and with immunocytochemistry using monoclonal antibodies against cytokeratin 7(CK 7), CK 8/18, Ber-Ep4, vimentin, calretinin and CD68. Ber-Ep4 is a marker for cells with epithelial origin (in some cases for mesothelial cells as well). CD68 is specific for cells from monocyte/macrophage lineage; CK7 and CK8/18 are markers for both endometrial epithelial and mesothelial cells, whereas calretinin and vimentin are markers for both endometrial stromal and mesothelial cells. RESULTS: In comparison with the nonmenstrual phase of the cycle, analysis of PF during menstruation showed an increased concentration of leucocytes (3.3 x 109/L vs 0.8 x 109/L, P = 0.03), erythrocytes (0.3 x 1012/L vs 0.02 x 1012/L, P = 0.006), hematocrit (0.03 L/L vs 0.003 L/L, P = 0.01) and hemoglobin (0.8 g/dL vs 0.1 g/dL, P = 0.01). Mesothelial cells stained positively with CK7, CK8/18, vimentin, and calretinin. Cells positive for Ber-Ep4 were not observed, except in 2 patients with endometriosis investigated during menses. In all patients 50-98% of single cells were strongly positive for both vimentin and CD68. CONCLUSION: When compared to nonmenstrual phases of the cycle, menstruation is associated with an increased concentration of red and white blood cells in PF. However, the presence of EM cells that are detectable by immunohistochemistry in PF is low during all phases of the cycle, including menstruation.

The E2F-regulated gene Chk1 is highly expressed in triple-negative estrogen receptor /progesterone receptor /HER-2 breast carcinomas.

We previously showed that checkpoint kinase 1 (Chk1) and Claspin, two DNA-damage checkpoint proteins, were down-regulated by 1,25-dihydroxyvitamin D(3), a known inhibitor of cell proliferation. In the present study, we aimed to investigate the transcriptional regulation of Chk1 and Claspin and to study their expression levels in human breast cancer tissue. Transient transfection experiments in MCF-7 breast cancer cells showed that promoter activities of Chk1 and Claspin were regulated by the E2F family of transcription factors. Subsequently, transcript levels of Chk1, Claspin, and E2F1 were determined by quantitative reverse transcriptase-PCR analysis in 103 primary invasive breast carcinomas and were compared with several clinicopathologic variables in breast cancer. A strong correlation was found between Chk1 and Claspin transcript levels. Transcript levels of Chk1, Claspin, and E2F1 were highest in histologic grade 3 tumors and in tumors in which the expression of estrogen receptor (ER) and progesterone receptor (PR) was lost. Moreover, Chk1 expression was significantly elevated in grade 3 breast carcinomas showing a triple-negative ER-/PR-/HER-2- phenotype compared with other grade 3 tumors. Further research is warranted to validate the use of Chk1 inhibitors in triple-negative breast carcinomas for which treatment strategies are limited at present.

Adenomyoepitheliomatous lesions of the mammary glands in transgenic mice with targeted PLAG1 overexpression.

PLAG1 proto-oncogene overexpression has been causally linked to multiple tumors, highlighting its broad tumorigenic relevance. Here, the oncogenic potential of PLAG1 in mammary gland tumorigenesis was investigated in PLAG1 transgenic mice. To target mammary glands, mice of 2 independent PLAG1 transgenic strains, PTMS1 and PTMS2, in which PLAG1 expression can be modulated by Cre-mediation, were crossed with MMTV-Cre transgenic mice, resulting in P1-MCre and P2-MCre offspring, respectively. Hundred percentage of P1-MCre female mice showed mammary gland hyperplasia, caused by adenomyoepithelial adenosis, at 8 weeks. The tumorigenic process could not be studied further in P1-MCre mice, because concomitant fast-growing salivary gland tumors required euthanasia. Sixteen percentage of P2-MCre females developed mammary gland adenomyoepitheliomas within 30-45 weeks, and none displayed concomitant salivary gland tumors. To further study mammary gland tumorigenesis in PTMS1-derived mice, intercrossing with WAP-Cre transgenic mice, resulting in P1-WAPCre mice, was performed to target PLAG1 expression more specifically to mammary glands. Eighty percentage of such mice developed adenomyoepitheliomas within 53-88 weeks. All PLAG1-induced adenomyoepitheliomas revealed expression upregulation of Igf2/H19, Dlk1/Gtl2, Igfbps and Wnt signaling genes (Wnt6, Cyclin D1). Collectively, these results establish the oncogenic potential of PLAG1 in mammary glands of mice and point towards contributing roles of Igf and Wnt signaling.

Prophylactic mastectomy in familial breast carcinoma. What do the pathologic findings learn us?

OBJECTIVE: To evaluate the indications, techniques and pathologic findings of prophylactic mastectomy (pME). METHODS: Retrospective study of patients with a strong family history of breast cancer (and ovarian cancer) or with proven BRCA mutation. RESULTS: Between January 1995 and December 2005, sixty seven patients underwent pME: 15% had a BRCA1 mutation, 31% had a BRCA2 mutation and 33% had a strong family history but without proven BRCA mutation and in 21% mutation analysis was not performed. Fifty eight percent had a personal history of breast cancer of which 84% previously underwent a unilateral mastectomy as part of their treatment. The median time to decision from previous treatment for breast carcinoma to pME was 46 months. Mean age at pME was 43 years. Pathologic examination of the pME specimens revealed invasive and/or in situ carcinoma in 19% (13/67). Atypical ductal/lobular hyperplasia (ADH/ALH) or flat epithelial atypia (FEA) were present in another 3%. CONCLUSION: Twenty two percent of women at high risk for breast cancer presented at the time of pME with invasive carcinoma or intra-epithelial neoplasia undetected by imaging and clinical examination.

A thyroid thriller: acute transient and symmetric goiter after fine-needle aspiration of a solitary thyroid nodule.

OBJECTIVE: To report a case of a patient who developed an acute and transient, tender, and bilateral swelling of the thyroid that occurred during fine-needle aspiration (FNA) of a solitary nodule in the left thyroid lobe; to add accurate ultrasound measurements to support our clinical observation; and to analyze a possible underlying mechanism of this rare condition. RESULTS AND CLINICAL FOLLOW-UP: The calculated thyroid volume increased from 23 to 57 mL before and at 4 minutes, respectively, after the needle aspiration, but the thyroid volume returned to prediagnostic level after 4 hours. Cytology, serum calcitonin, and histology were concordant, and the nodule was diagnosed as a medullary thyroid carcinoma. Immunohistochemistry was positive for calcitonin, chromogranin, and the very potent vasodilator calcitonin gene-related peptide (CGRP). CONCLUSION: This is a rare case of acute and transient thyroid swelling during a common procedure as FNA of a thyroid nodule. This is the first case with documented acute volume expansion quantified by ultrasound measurements supporting our clinical observation, which is in accordance with two historical case reports. The clinical and ultrasound data support the hypothesis of vasodilation as the underlying mechanism, possibly evoked by the release of the vasodilator CGRP.

Sentinel lymph node biopsy and non-sentinel node involvement in special type breast carcinomas with a good prognosis.

This study aimed at identifying factors related to sentinel lymph node (SLN) involvement in patients with tubular, cribriform, mucinous or papillary breast carcinoma and those related to non-SLN metastases if an SLN was positive. Multivariate analyses involved logistic and stepwise regressions. The SLNs harboured metastases in 85 of 572 cases, 78 of whom underwent axillary dissection; 19 presented non-SLN positive disease. Lack of lymphovascular invasion, a tumour size < or = 10 mm and a single SLN removed were the factors predicting an SLN metastasis rate <10%, and patients with these features could be candidates for no surgical axillary staging. A positive SLN proportion of < or = 50% and no lymphovascular invasion were associated with a <10% rate of non-SLN invasion; patients with a positive SLN and these features could be candidates for the omission of completion axillary dissection. The opposite presentation of these factors would mandate SLN biopsy and axillary dissection, respectively.

Sentinel lymph node biopsy in staging small (up to 15 mm) breast carcinomas. Results from a European multi-institutional study.

Sentinel lymph node (SLN) biopsy has become the preferred method for the nodal staging of early breast cancer, but controversy exists regarding its universal use and consequences in small tumors. 2929 cases of breast carcinomas not larger than 15 mm and staged with SLN biopsy with or without axillary dissection were collected from the authors' institutions. The pathology of the SLNs included multilevel hematoxylin and eosin (HE) staining. Cytokeratin immunohistochemistry (IHC) was commonly used for cases negative with HE staining. Variables influencing SLN involvement and non-SLN involvement were studied with logistic regression. Factors that influenced SLN involvement included tumor size, multifocality, grade and age. Small tumors up to 4 mm (including in situ and microinvasive carcinomas) seem to have SLN involvement in less than 10%. Non-SLN metastases were associated with tumor grade, the ratio of involved SLNs and SLN involvement type. Isolated tumor cells were not likely to be associated with further nodal load, whereas micrometastases had some subsets with low risk of non-SLN involvement and subsets with higher proportion of further nodal spread. In situ and microinvasive carcinomas have a very low risk of SLN involvement, therefore, these tumors might not need SLN biopsy for staging, and this may be the approach used for very small invasive carcinomas. If an SLN is involved, isolated tumor cells are rarely if ever associated with non-SLN metastases, and subsets of micrometastatic SLN involvement may be approached similarly. With macrometastases the risk of non-SLN involvement increases, and further axillary treatment should be generally indicated.

Clinical relevance of thyroid fluorodeoxyglucose-whole body positron emission tomography incidentaloma.

Fluorodeoxyglucose (FDG) whole body positron emission tomography (PET) scan is increasingly used in the diagnostic work-up or follow-up of patients. In these conditions, positive PET scans with unexpected hot spots within the thyroid region could be defined as thyroid FDG-PET incidentaloma (in analogy with unexpected sonographic thyroid nodules). We describe eight consecutive patients referred to the endocrine department because of thyroid "hot spots," incidentally detected by whole body FDG-PET scan (September 1999 to March 2001). Using ultrasound, fine needle aspiration cytology (FNAC), and histology reports, we tried to identify the pathology underlying thyroid FDG-PET incidentaloma. FNAC showed an indication for surgery in all patients. Surgery has been performed in 7 patients. Malignancy was correctly identified in five patients: two medullary thyroid carcinomas, one with lymph node invasion, and three papillary thyroid carcinomas with invasion through the thyroid capsule in two of the PTC cases. In two patients with a positive FDG-PET scan, FNAC pointed to follicular neoplasms, and final histology reports showed follicular adenoma. In the remaining patient, FNAC revealed a follicular lesion, but surgery has not yet been performed. In conclusion, a small series of consecutive thyroid FDG-PET incidentaloma cases is presented and suggests a high rate of clinically relevant malignancies.

Use of liquid-based cytology in serous fluids: a comparison with conventional cytopreparatory techniques.

OBJECTIVE: To examine the performance of liquid-based cytology (LBC) on body cavity fluids as compared with conventional cytopreparatory techniques. STUDY DESIGN: Slides from a total of 592 serous fluids, effusions as well as intraoperative washings, were reviewed. Of these fluids, 301 were collected before and 291 after the introduction of the PrepStain LBC (TriPath Imaging, Burlington, North Carolina, U.S.A.) in our laboratory. RESULTS: PrepStain thin layers showed excellent morphology on a clear background, with preservation of 3-dimensional configurations and a sufficient amount of extracellular material to allow an accurate diagnosis. Cytopreparation, screening and interpretation of LBC were less time consuming. Moreover, when the performance of the various cytopreparatory techniques in malignant fluids was studied, PrepStain thin layers excelled over conventional cytopreparatory methods in showing a significantly lower false negative rate (P=.0414). CONCLUSION: Our findings indicate that in body fluid cytology, thin layers can safely replace other types of wet-fixed preparations, resulting in enhanced specimen quality and diminished false negative rates.

Qualitative assessment of the progesterone receptor and HER2 improves the Nottingham Prognostic Index up to 5 years after breast cancer diagnosis.

PURPOSE: To investigate whether the estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) can improve the Nottingham Prognostic Index (NPI) in the classification of patients with primary operable breast cancer for disease-free survival (DFS). PATIENTS AND METHODS: The analysis is based on 1,927 patients with breast cancer treated between 2000 and 2005 at the University Hospitals, Leuven. We compared performances of NPI with and without ER, PR and/or HER2. Validation was done on two external data sets containing 862 and 2,805 patients from Oslo (Norway) and Auckland (New Zealand), respectively. RESULTS: In the Leuven cohort, median follow-up was 66 months, and 13.7% of patients experienced a breast cancer-related event. Positive staining for ER, PR, and HER2 was detected, respectively, in 86.9%, 75.5%, and 11.9% of patients. Based on multivariate Cox regression modeling, the improved NPI (iNPI) was derived as NPI - PR positivity + HER2 positivity. Validation results showed a risk group reclassification of 20% to 30% of patients when using iNPI with its optimal risk boundaries versus NPI, in a majority of patients to more appropriate risk groups. An additional 10% of patients were classified into the extreme risk groups, where clinical actions are less ambiguous. Survival curves of reclassified patients resembled more closely those for patients in the same iNPI group than those for patients in the same NPI group. CONCLUSION: The addition of PR and HER2 to NPI increases its 5-year prognostic accuracy. The iNPI can be considered as a clinically useful tool for stratification of patients with breast cancer receiving standard of care.

Polysomy 17 in breast cancer: clinicopathologic significance and impact on HER-2 testing.

PURPOSE: Polysomy 17 is frequently found in breast cancer and may complicate the interpretation of HER-2 testing results. We investigated the impact of polysomy 17 on HER-2 testing and studied its clinicopathologic significance in relation to HER2 gene amplification. PATIENTS AND METHODS: In 226 patients with primary invasive breast carcinoma, HER2 gene and chromosome 17 copy numbers were determined by dual-color fluorescent in situ hybridization (FISH). The interpretation of FISH results was based on either absolute HER2 gene copy number or the ratio HER2/chromosome 17. Results were correlated with HER-2 protein expression on immunohistochemistry (IHC), HER2 mRNA expression by reverse transcriptase polymerase chain reaction (RT-PCR), and with various clinicopathologic parameters. RESULTS: All cases with an equivocal HER-2 result by FISH, either by absolute HER2 copy number (44 of 226 patients; 19.5%) or by the ratio HER2/chromosome 17 (three of 226 patients; 1.3%), displayed polysomy 17. On its own, polysomy 17 was not associated with HER-2 overexpression on IHC or increased HER2 mRNA levels by RT-PCR. Moreover, and in contrast with HER2 gene amplification, polysomy 17 was not associated with high tumor grade, hormone receptor negativity, or reduced disease-free survival. CONCLUSION: Polysomy 17 affects HER-2 testing in breast cancer and is a major cause of equivocal results by FISH. We show that tumors displaying polysomy 17 in the absence of HER2 gene amplification resemble more HER-2-negative than HER-2-positive tumors. These findings highlight the need for clinical trials to investigative whether polysomy 17 tumors benefit from HER-2-targeted therapy.

The complexity of genotypic alterations underlying HER2-positive breast cancer: an explanation for its clinical heterogeneity.

PURPOSE OF REVIEW: We discuss recent findings on the genotypic alterations associated with HER2-positive breast cancer in an attempt to clarify the clinical heterogeneity observed among these tumors. RECENT FINDINGS: Molecular genetic analysis supports the distinctive nature of HER2-positive breast cancer, which is primarily driven by HER2 gene amplification. Depending on the amplicon size, a variety of genes can be coamplified and overexpressed together with HER2, some of which may contribute to tumorigenesis; the amplicon size may even predict response to trastuzumab therapy. HER2 gene amplification may further destabilize the tumor genome, facilitating the generation of additional genomic aberrations including aneuploidy. The latter might imply polysomy 17, a phenomenon that should be discriminated from true HER2 gene amplification: polysomy 17 in the absence of HER2 gene amplification is not associated with HER2 overexpression nor with the clinical characteristics of HER2-positive breast cancer. SUMMARY: HER2 gene amplification is a complex event: it includes coamplification of other, potentially oncogenic genes and facilitates the generation of additional genomic aberrations. Further studies on these genotypic findings will be helpful to better identify the patients that might benefit from trastuzumab therapy.

Validation of clinical prediction rules for a low probability of nonsentinel and extensive lymph node involvement in breast cancer patients.

BACKGROUND: Two recently developed clinical prediction rules aim to anticipate the lack of nonsentinel lymph node metastases and the involvement of less than 4 lymph nodes in breast cancer patients with positive sentinel lymph nodes (SLNs). METHODS: The University of Louisville Breast SLN Study clinical prediction rules were validated on an independent set of SLN-positive patients with tumors < or = 15 mm. RESULTS: The data on 475 and 473 patients, respectively, were used for the validation. The areas under the receiver operating characteristic curves were similar to the originals for both predictive tools (.70 and .76). The lowest score of 1 identified 5 of 7 patients with disease limited to the SLNs and 161 of 165 as having less than 4 involved lymph nodes. CONCLUSIONS: A subset of patients with SLN-only involvement and less than 4 metastatic lymph nodes can probably be identified by means of the Louisville clinical prediction rules, but prediction of the lack of non-SLN metastasis seems less reliable.

Comparative expressed sequence hybridization reveals differential gene expression in morphological breast cancer subtypes.

In this study, comparative expressed sequence hybridization (CESH) has been used to compare gene expression patterns in three morphologically different breast cancer subtypes: classic-type invasive lobular carcinoma (ILC), poorly differentiated ERBB2-negative invasive ductal carcinoma-not otherwise specified (IDC-NOS), and poorly differentiated ERBB2-positive IDC-NOS. CESH allows global detection of chromosomal regions with differential gene expression in a way similar to that of comparative genomic hybridization (CGH). Eight cases of each breast cancer subtype were included in the study. For each subtype, two pools of four cases each were constructed. CESH was used to compare both pools within the same morphological subtype, followed by a comparison of pools belonging to different subtypes. This revealed three chromosomal regions that were differentially expressed in ductal and lobular carcinomas, including relative overexpression at 8q13-q23 and 16q22, and relative underexpression at 8p21-p22. In addition, an expression signature characterized by relative overexpression at 3q24-q26.3, 14q23-31, 17q12, and 20q12-13 was identified for ERBB2-positive IDC-NOS. In summary, CESH analysis highlights chromosomal regions of differential gene expression that are associated with morphologically defined breast cancer subtypes and suggests that regions on chromosome 8 are of interest in the discrimination between ductal and lobular carcinomas. In addition, using CESH, it was possible to identify an ERBB2 expression signature, comprising four chromosomal regions with potential significance in the aggressive behaviour of ERBB2-positive IDC-NOS.

Association between HER-2/neu and the progesterone receptor in oestrogen-dependent breast cancer is age-related.

In oestrogen receptor-positive (ER+) breast cancer, HER-2/neu and the progesterone receptor (PR) are inversely associated. This explains a lower response to anti-oestrogens if ER+ breast cancers are HER-2/neu positive. One randomized study however, showed that premenopausal women with an ER+ breast cancer respond to anti-oestrogens independent of HER-2/neu. We therefore hypothesized an age-related association between HER-2/neu and PR in ER+ breast cancers. Receptors for ER, PR and HER-2/neu were analysed by immunohistochemistry (IHC). A uni- and multivariate analyses was carried out to assess this relationship in 1104 women with an ER+ breast cancer. We observed an inverse association between HER-2/neu and PR only after age 45. There were 173 women of age45 years. In multivariate analysis, only tumour grade (p=0.005) but not PR status was associated with HER-2/neu in women age45 years group, both PR status (p=0.001) and tumour grade (p=0.001) were independently associated with HER-2/neu. In ER+ breast cancers from women age>45, PR was positive in 76.9% if HER-2/neu negative but in 53.4% if HER-2/neu positive (p<0.001) and the median quantitative PR levels are 150 and 75 respectively in HER-2/neu negative and HER-2/neu positive tumours (p=0.002). This age effect of HER-2/neu on the PR status was not seen in women age45 years but not in women age