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Background: Gatipotuzumab is a fully humanized antibody which was designed to detect a cancer-specific glyco-modification of MUC1 (termed 'TA-MUC1') and which was optimized to effectively trigger antibody-dependent-cell-mediated cytotoxicity (ADCC) in cancer cells. Clinical trials investigating therapeutic efficacy of this antibody have been published recently. The current analysis aimed to determine whether TA-MUC1-as detected by Gatipotuzumab-is expressed in cervical cancer tissue and whether binding of Gatipotuzumab is associated with clinico-pathological variables including recurrence free (RFS) and overall survival (OS)., Methods: Cervical cancer tissue (n = 250) was stained for TA-MUC1 using Gatipotuzumab employing a standardized immunohistochemistry protocol. Staining was scored by applying the IR-score. Results were binarized and tested for association to clinico-pathological parameters., Results: TA-MUC1 as stained by Gatipotuzumab was detected in 188 (75.2%) out of the 250 cervical cancer cases investigated. Expression of TA-MUC1 was restricted to cancer cells and was positively correlated with viral oncoprotein E6. Membrane staining of TA-MUC1 predicted significantly reduced RFS and OS. Importantly, expression of TA-MUC1 at the cell surface identified a group of early stage cervical cancer patients with exceptional short RFS and OS., Conclusions: We report TA-MUC1-the antigen detected by Gatipotzumab-to be widely expressed in cervical cancer tissue and to localize to the cell membrane. The latter is seen as a pre-requisite to target this epitope by antibody-drug conjugates or antibodies eliciting ADCC. Since especially, membrane localization of TA-MUC1 predicted poor prognosis, evaluating Gatipotuzumab for its therapeutic efficacy in cervical cancer may turn attractive.

Background: Estrogen signalling is transmitted via various receptors and multiple intracellular signalling pathways. Estrogen receptor alpha (ERα)-mediated transcription of target genes has been demonstrated to be closely linked to human papilloma virus (HPV)-induced carcinogenesis in case of cervical cancer. So far, the role of non-genomic estrogen signals in cervical cancer, e.g. transmitted by the G protein-coupled estrogen receptor (GPER) remains to be rather elusive. Today's knowledge on the role of GPER in cervical cancer is sparse and-to the best of our knowledge-GPER has not been investigated in context with clinicopathological parameters or prognosis of cervical cancer. Therefore, the current study investigated whether GPER is expressed in cervical cancer tissue. Further, GPER was correlated to clinicopathological parameters, tissue markers of cervical carcinogenesis and to patient overall and recurrence-free survival., Materials and Methods: Cervical cancer tissue was collected from 156 patients during surgery between 1993 and 2002. GPER immunostaining was performed on all the cases and correlated to clinicopathological data. More than half of all patients were diagnosed at advanced stage (FIGO II-IV 93/156; 59.6%) of disease. The large majority of patients presented with tumours of intermediate or high grade (G2-3 140/152, 92.1%). 22 cervical cancer-related deaths (22/156, 14.1%) were documented during the follow-up period., Results: GPER was detected in various subcellular staining patterns. In 129/156 (82.7%) cases GPER was expressed in the tumour cell cytoplasm (GPER cyt ). GPER immunopositivity at the cell membrane (GPER mem ) was found in 114/156 (73.1%) cases. While co-occurrence of both membrane and cytoplasmic staining (GPER cyt  + GPER mem ) was detected in the majority of tissue samples (101/156; 64.7%), only few cases (14/156, 9.0%) were classified as not expressing GPER at all. GPER cyt was positively correlated with tumour grade. Statistical associations of GPER and both p16 and p53 were detected. Finally, immunopositivity of GPER cyt was predictive for favourable overall as well as recurrence-free survival in cervical cancer of early stage (FIGO I)., Conclusion: This retrospective study reports GPER cyt to be associated with improved overall and recurrence-free survival in early-stage cervical cancer. Further investigations are needed thus to determine whether this observation may be of clinical impact. Interestingly, Raloxifene-a GPER-activating selective estrogen receptor modulator-has recently been demonstrated to be preventive for cervical cancer relapse in mice. Whether this effect is only reliant on raloxifene blocking ERα or may also be related to activation of GPER remains to be determined.

Background: The detection of circulating tumour cells (CTCs) from peripheral blood of cancer patients can be carried out by real-time PCR approaches using different gene expression levels of tumour cells and surrounding blood cells., Materials and Methods: Potential marker genes were first analyzed in a model system and then applied to 20 blood samples of adjuvant breast cancer patients and gene expression levels were correlated to tumour characteristics., Results: The mean of gene expression levels was found elevated for the four genes analyzed in the adjuvant breast cancer patient group in comparison to the samples of the group of healthy donors, but no correlation between gene expression and tumour characteristics could be detected as being statistically significant., Conclusion: The results demonstrated, that the employed methodology is functional, but has to be refined by certain approaches like simultaneously running a state-of-the-art system of CTC-detection comparing the results, and by an enlargement of patient collective and number of marker genes., (Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.)

Altered glycosylation is a predominant feature of tumour cells; it serves for cell adhesion and detachment, respectively, and facilitates the immune escape of these cells. Therefore changes in the expression of glycosyltransferase genes could help to identify circulating tumour cells (CTCs) in the blood samples of cancer patients using a quantitative polymerase chain reaction (PCR) approach. Blood samples of healthy donors were inoculated with certain numbers of established breast cancer cell line cells, thus creating a model system. These samples were analysed by quantitative PCR for the expression of six different glycosyltransferase genes. The three genes with the best results in the model system were consecutively applied to samples from adjuvant breast cancer patients and of healthy donors. FUT3 and GALNT6 showed the highest increase in relative expression, while GALNT6 and ST3GAL3 were the first to reach statistically significant different ∆CT-values comparing the sample with and without addition of tumour cells. These three genes were applied to patient samples, but did not show any significant results that may suggest the presence of CTCs in the blood. Although the relative expression of some of the glycosyltransferase genes exhibited reasonable results in the model system, their application to breast cancer patient samples will have to be further improved, e.g. by co-analysis of patient blood samples by gold-standard methods.

Objectives: Gestational trophoblastic disease (GTD) involves a spectrum of abnormal proliferations arising from the placental villous trophoblast. Although the incidence is low, a biomarker with short serum half-life would be a major clinical advance to monitor surgical and medical treatment reducing the socioeconomic burden of multiple control visits as well as patient's anxiety. Placental growth hormone (hGH-V) plays an important role in the regulation of normal placental growth and has shown angiogenic effects. We aimed to determine by immunohistochemistry (IHC) whether hGH-V is expressed in GTD and whether it can be detected in the patient's blood for potential monitoring of surgical or medical treatment procedures., Methods: Tissue and sera were collected from women undergoing treatment for GTD in a tertiary care university hospital. We evaluated partial and complete hydatidiform moles, invasive moles and choriocarcinoma, n=16. Trophoblast specimens were examined by a newly developed IHC set-up for hGH-V in addition to gross morphologic and histopathological examination. Serum samples were analyzed by a highly sensitive hGH-V specific immunoassay., Results: hGH-V was localized in all entities of GTD to the syncytiotrophoblast by immunohistochemistry. Serum hGH-V was detected for the first time in GTD and was present in a high percentage of all analyzed entities., Conclusions: hGH-V can be detected in all entities of GTD by IHC as well as by serum analysis and may therefore serve as a novel biomarker for the disease. Its clinical utility in diagnosis of GTD and monitoring surgical or medical treatment needs to be determined in further studies., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Background: Glycosylation is the most frequent posttranslational modification of proteins and lipids influencing inter- and intracellular communication and cell adhesion. Altered glycosylation patterns are characteristically observed in tumour cells. Normal and altered carbohydrate chains are transferred to their acceptor structures via glycosyltransferases. Here, we present the correlation between the presence of three different glycosyltransferases and tumour characteristics., Methods: 235 breast cancer tissue samples were stained immunohistochemically for the glycosyltransferases N-acetylgalactosaminyltransferase 6 (GALNT6), β-1,6-N-acetylglucosaminyltransferase 2 (GCNT2), and ST6 (α-N-acetyl-neuraminyl-2,3-β-galactosyl-1,3)-N-acetylgalactosamine α-2,6-sialyltransferase 1 (ST6GALNac1). Staining was evaluated by light microscopy and was correlated to different tumour characteristics by statistical analysis., Results: We found a statistically significant correlation for the presence of glycosyltransferases and tumour size and grading. Specifically smaller tumours with low grading revealed the highest incidences of glycosyltransferases. Additionally, Her4-expression but not pHer4-expression is correlated with the presence of glycosyltransferases. All other investigated parameters could not uncover any statistically significant reciprocity., Conclusion: Here we show, that glycosyltransferases can identify small tumours with well-differentiated cells; hence, glycosylation patterns could be used as a marker for early tumourigenesis. This assumption is supported by the fact that Her4 is also correlated to glycosylation, whereas the activated form of Her4 does not show such a connection with glycosylation.

The progesterone receptor (PR) has been increasingly well described as an important mediator of the pathogenesis and progression of breast cancer. The aim of this study was to assess the role of PR status as a prognostic factor in addition to other well-established prognostic factors. Data from five independent German breast cancer centers were pooled. A total of 7,965 breast cancer patients were included for whom information about their PR status was known, as well as other patient and tumor characteristics commonly used as prognostic factors. Cox proportional hazards models were built to compare the predictive value of PR status in addition to age at diagnosis, tumor size, nodal status, grading, and estrogen receptor (ER) status. PR status significantly increased the accuracy of prognostic predictions with regard to overall survival, distant disease-free survival, and local recurrence-free survival. There were differences with regard to its prognostic value relative to subgroups such as nodal status, ER status, and grading. The prognostic value of PR status was greatest in patients with a positive nodal status, negative ER status, and low grading. The PR-status adds prognostic value in addition to ER status and should not be omitted from clinical routine testing. The significantly greater prognostic value in node-positive and high-grade tumors suggests a greater role in the progression of advanced and aggressive tumors.

Purpose: Ovarian carcinoma is the third most common gynecological cancer and only short recurrence-free survival and overall survival times are achieved. The role of the estrogen receptor expression is well studied in breast cancer and breast cancer cell lines. Patients with positive estrogen receptor expression have a lower risk for recurrence and a better overall survival. Previous studies have shown that ESR1 methylation influences ovarian cancer development and might thus play a role regarding prognosis of ovarian carcinoma., Methods: A total of 75 patients were identified that were treated for ovarian carcinoma by debulking surgery and adjuvant standard chemotherapy. Isolation and bisulfite treatment of genomic DNA from serial sections of surgically resected ovarian carcinoma tissue was performed using commercially available kits. For the detection of methylated ESR1 promoter sequences, real-time methylation-specific PCR was used., Results: Promoter methylation did not show a correlation between clinical-pathological data for all patients. However, within the subgroup of low-grade ovarian carcinoma patients and patients with an ovarian tumor of low malignant potential methylation of the ESR1 promoter inversely correlated with survival (p = 0.031)., Conclusions: Although small numbers of ovarian carcinoma patients were analyzed, methylation status might be useful as a prognostic marker within the subgroup of low-grade ovarian carcinoma patients. Further studies should investigate a larger cohort and also address the use of demethylation agents with respect to improve patient's prognosis in this subgroup of ovarian carcinoma patients.

Ovarian granulosa cell tumors (GCTs) are thought to arise from cells of the ovarian follicle and comprise a rare entity of ovarian masses. We recently identified the G-protein-coupled estrogen receptor (GPER/GPR30) to be present in granulosa cells, to be regulated by gonadotropins in epithelial ovarian cancer and to be differentially expressed throughout folliculogenesis. Thus, supposing a possible role of GPER in GCTs, this study aimed to analyze GPER in GCTs. GPER immunoreactivity in GCTs (n = 26; n (primary diagnosis) = 15, n (recurrence) = 11) was studied and correlated with the main clinicopathological variables. Positive GPER staining was identified in 53.8% (14/26) of GCTs and there was no significant relation of GPER with tumor size or lymph node status. Those cases presenting with strong GPER intensity at primary diagnosis showed a significant reduced overall survival (p = 0.002). Due to the fact that GPER is regulated by estrogens, as well as gonadotropins, GPER may also be affected by endocrine therapies applied to GCT patients. Moreover, with our data supposing GPER to be associated with GCT prognosis, GPER might be considered as a possible confounder when assessing the efficacy of hormone-based therapeutic approaches in GCTs.

Cathepsin D is a protease involved in the metastasis and angiogenesis of mammary carcinomas. This review analyzes the significance of the tumor protease cathepsin D in mammary carcinomas as a tumor marker. We present a systematic overview based on a selective Medline search. Cathepsin D is expressed in mammary carcinomas and exhibits higher expression in invasive ductal carcinomas compared with lobular carcinomas. Nodal positive carcinomas showed reduced cathepsin D expression compared to lymph node metastases, and increased expression has been observed in hormone-receptor negative tumors. Thus, the expression of cathepsin varies between the two histological types. Increased cathepsin-D expression in acinar affection has also been described. The lack of an association of cathepsin D with known prognostic factors such as CA15-3, ERalpha and ERbeta does not prevent it from being using as a tumor marker. Cathepsin has already been used along with other genes as a prognostic parameter for carcinoma patients in gene arrays.

Background: Knowledge on immunosuppressive factors in the pathogenesis of endometrial cancer is scarce. The aim of this study was to assess Glycodelin (Gd) and its immunosuppressive isoform Glycodelin A (GdA) in endometrial cancer tissue and to analyze its impact on clinical and pathological features and patient outcome., Methods: 292 patients diagnosed and treated for endometrial cancer were included. Patient characteristics, histology and follow-up data were available. Gd and GdA was determined by immunohistochemistry and in situ hybridization was performed for Gd mRNA., Results: Endometrial cancer shows intermediate (52.2%) or high (20.6%) expression for Gd in 72.8%, and GdA in 71.6% (intermediate 62.6%, high 9.0%) of all cases. The glycosylation dependent staining of GdA is tumour specific and correlates with the peptide-specific Gd staining though neither of the two is associated with estrogen receptor, progesterone receptor or clinic-pathological features. Also Gd protein positively correlates with Gd mRNA as quantified by in situ hybridization. Gd positive cases have a favourable prognosis (p = 0.039), while GdA positive patients have a poor outcome (p = 0.003). Cox-regression analysis proofed GdA to be an independent prognostic marker for patient survival (p = 0.002), besides tumour stage, grade and the concomitant diagnosis of hypertension., Conclusion: Gd and GdA are commonly expressed in endometrial cancer tissue and seem to be of relevance in tumourigenesis. They differ not only in glycosylation but also in their biological activity, since only GdA holds prognostic significance for a poor overall survival in endometrial cancer patients. This finding might be explained by GdAs immunosuppressive capacity.

Introduction: Inhibins are important regulators of the female reproductive system. Recently, two new inhibin-subunits βC and βE have been described, although, their function is still quite unclear. Interestingly, there is an association between interferon and TGF-β expression. Therefore, the aim of this study was to determine expression changes of inhibin-βC and -βE subunits in endometrial Ishikawa carcinoma cell line after stimulation with interferon-β1a., Materials and Methods: The Ishikawa cell line was cultured until confluence was observed (after 2 days). After adding interferon-β1a (1,000 IE/ml), Ishikawa cells were analyzed for inhibin-βC and -βE subunits by RT-PCR. The fibroblast cell line BJ6 served as negative control. Experiments were performed in triplicates., Results: The endometrial adenocarcinoma cell line Ishikawa synthesized the inhibin- βC and -βE subunits. The fibroblast cells BJ6 did not demonstrate an inhibin -βC and -βE mRNA expression, while inhibin-βC subunit is down-regulated and inhibin-βE is up-regulated in Ishikawa carcinoma cell line after stimulation with interferon-β1a in Ishikawa., Discussion: We demonstrated for the first time a functional relationship between interferon and the novel inhibin-βC and -βE subunits. It might be possible that interferon exerts a possible apoptotic function through the βE-subunit, while, by down-regulating the βC isoform, cell proliferation is inhibited. However, the precise function of the novel βC- and βE-subunits are still not known in human endometrial tissue and a possible association with interferon is still unclear and warrants further research.

Follicle stimulating hormone receptor (FSHR) and luteinizing hormone receptor (LHCGR) were demonstrated to impact upon survival of patients suffering from epithelial ovarian cancer (EOC). Though structure wise the G-protein coupled estrogen receptor (GPER/GPR30) is related to FSHR/LHCGR, its prognostic impact in EOC remains controversial. We recently found that FSHR negative patients represent a specific EOC subgroup that may behave differently in respect to both treatment response and prognosis. Hence, the current study aimed to analyze how GPER may interact with the FSHR/LHCGR system in EOC and whether the prognostic significance of GPER in EOC cases (n=151) may be dependent on the FSHR/LHCGR immunophenotype of the tumor. Ovarian cancer cell lines were used to study how FSH and LH regulate GPER and whether GPER activation differentially affects in vitro cell proliferation in presence/absence of activated FSHR/LHCGR. In EOC tissue, GPER correlated with FSHR/LHCGR and was related to prolonged overall survival only in FSHR/LHCGR negative patients. Although GPER was found to be specifically induced by LH/FSH, GPER agonists (4-Hydroxy-Tamoxifen, G1) reduced EOC cell proliferation only in case of LH/FSH unstimulated pathways. To the same direction, only patients characterized as LHCGR/FSHR negative seem to gain from GPER in terms of survival. Our combined tissue and in vitro results support thus the hypothesis that GPER activation could be of therapeutic benefit in LHCGR/FSHR negative EOC patients. Further studies are needed to evaluate the impact of GPER activation on a clinical scheme.

Background: The aim of this study was to evaluate the expression of the cell adhesion-related glycoproteins MUC-1, β-catenin and E-cadherin in multicentric/multifocal breast cancer in comparison to unifocal disease in order to identify potential differences in the biology of these tumor types., Methods: A retrospective analysis was performed on the expression of MUC1, β-catenin and E-cadherin by immunohistochemistry on tumor tissues of a series of 112 breast cancer patients (total collective) treated in Munich between 2000 and 2002. By matched-pair analysis, 46 patients were entered into two comparable groups of 23 patients after categorizing them as having multicentric/multifocal or unifocal breast cancer. Matching criteria were tumor size, histology grade and lymph node status; based on these criteria, patients were distributed equally between the two groups (p = 1.000 each). Data were analyzed with the Kruskal-Wallis and the Mann-Whitney tests., Results: In the matched groups, we found a significantly down-regulated expression of E-cadherin in multicentric/multifocal breast cancer compared to unifocal disease (p = 0.024). The total collective showed even higher significance with a value of p < 0.0001. In contrast, no significant differences were observed in the expression of β-catenin between multicentric/multifocal and unifocal tumors (p = 0.636 and p = 0.914, respectively). When comparing the expression of MUC1, E-cadherin and β-catenin within the unifocal group, we found a significant positive correlation between E-cadherin and β-catenin (p = 0.003). In the multicentric/multifocal group we observed, in contrast to the unifocal group, a significant decrease of MUC1 expression with increased grading (p = 0.027)., Conclusion: This study demonstrates that multicentric/multifocal and unifocal breast cancers with identical TNM-staging clearly differ in the expression level of E-cadherin. We suggest that the down-regulation of E-cadherin in multicentric/multifocal breast cancer is causally connected with the worse prognosis of this tumor type.

PankoMab-GEX is a novel humanized and glycooptimized antibody, which recognizes a novel specific tumour epitope of MUC1 (TA-MUC1). The aim of this study was to evaluate PankoMab-GEX binding to a variety of ovarian cancer specimens (n=156) and to normal ovarian tissue. In addition, PankoMab-GEX staining was compared to that of the well-known anti-MUC1 antibodies HMFG-1 and 115D8. PankoMab-GEX showed positive reactivity in serous (100% of cases, mean IRS 8.23), endometrioid (95% of cases, mean IRS 6.40), mucinous (58% of cases, mean IRS 4.17), and clear cell (92% of cases, mean IRS 7.58) carcinomas. In contrast to HMFG-1, healthy ovarian tissue was not recognized by PankoMab-GEX. Staining with antibody 115D8 was increased with staging. Cytoplasmic PankoMab-GEX staining increased with tumour grade, but no correlation was found with staging. Univariate Kaplan-Meier analysis revealed a tendency of reduced survival of patients with high expression of TA-MUC1. The findings are encouraging with respect to a potential use of PankoMab-GEX as a new therapeutic antibody for the treatment of ovarian cancer patients.

Unlabelled: There is evidence that breast cancer patients suffer from thyroid disorders. However, the relation between thyroid receptor (TR) expression and breast cancer remains unknown so far. Therefore, the aim of this study was an immunohistochemical analysis of TR expression in breast cancer patients., Materials and Methods: The expression of the combined antibody TRalpha1 and 2 and TRalpha1 or 2 alone as well as the expression of combined TRbeta1 and 2 and TRbeta1 or 2 alone was investigated with specific monoclonal or polyclonal antibodies in 82 patients. All patients presented with a first diagnosis of sporadic breast cancer. The ABC method was used for staining and staining intensities were analyzed using the IRS score., Results: Both TRalpha and TRbeta were expressed in the nuclei of breast cancer cells. In 24% (28/78) of the slides TRalpha1 and 2 IRS was positive. Immunopositivity for TRalpha1 was found in 55/78 slides, for TRalpha 2 in 54/79 slides (71 and 68%, respectively). The expression of TRbeta1 and 2 showed a positive detection in 33/77 (43%) of the slides, for TRbeta1 it was 43/79 (54%), for TRbeta2 60/76 (79%). Significant correlations of the expression of TRs - especially TRalpha2 - were found with further prognostic histopathological parameters such as tumor size, axillary lymph node involvement, grading and hormone receptor status. Multivariate analysis showed a trend for TRalpha2 as an independent predictor of disease-free and overall survival., Discussion: Our results revealed specific alterations in the expression of TRs - especially of TRalpha2 - in breast cancer patients, suggesting it as a marker with possible prognostic validity.

It is widely known that cells from epithelial tumors, e.g., breast cancer, detach from their primary tissue and enter blood circulation. We show that the presence of circulating tumor cells (CTCs) in samples of patients with primary and metastatic breast cancer can be detected with an array of selected tumor-marker-genes by reverse transcription real-time PCR. The focus of the presented work is on detecting differences in gene expression between healthy individuals and adjuvant and metastatic breast cancer patients, not an accurate quantification of these differences. Therefore, total RNA was isolated from blood samples of healthy donors and patients with primary or metastatic breast cancer after enrichment of mononuclear cells by density gradient centrifugation. After reverse transcription real-time PCR was carried out with a set of marker genes (BCSP, CK8, Her2, MGL, CK18, CK19). B2M and GAPDH were used as reference genes. Blood samples from patients with metastatic disease revealed increased cytokine gene levels in comparison to normal blood samples. Detection of a single gene was not sufficient to detect CTCs by reverse transcription real-time PCR. Markers used here were selected based on a recent study detecting cancer cells on different protein levels. The combination of such a marker array leads to higher and more specific discovery rates, predominantly in metastatic patients. Identification of CTCs by PCR methods may lead to better diagnosis and prognosis and could help to choose an adequate therapy.

Background: Glycodelin is a cell surface glycoprotein offering a unique gender specific carbohydrate configuration. Sialylated carbohydrate structures, which are unusual for mammals, characterize Glycodelin isolated from amniotic fluid (Glycodelin A, GdA). Glycodelin in general exerts multiple, partly opposing functions ranging from immunosuppression to cell differentiation. As these markedly influence tumorigenesis, this study aimed to clarify whether expression of different Glycodelin isoforms is related to clinicopathological characteristics and prognosis of ovarian cancer patients. Further the use of Glycodelin as a serum marker in benign and malignant ovarian diseases was evaluated., Methods: Ovarian cancer specimens (n = 152) were stained for Glycodelin with carbohydrate and peptide specific antibodies. Associations between Glycodelin expression and histological grading, FIGO stage as well as patient's prognosis were examined. Glycodelin was correlated to expression of gonadotropin receptors and mucin-1, which are discussed as ovarian cancer tissue markers. In addition, Glycodelin serum concentrations were analyzed in patients suffering from benign (n = 73) or malignant (n = 38) ovarian neoplasias., Results: Glycodelin A was found to be an independent prognostic marker for poor prognosis in advanced ovarian cancer patients. GdA staining correlated with gonadotropin receptor (FSHR and LHCGR) and with hCG expression. Gd expression showed a positive correlation with a tumour-associated epitope of mucin 1 (TA-MUC1). Further, compared to ovarian cancer, serum Gd was increased in patients with benign ovarian tumors., Conclusion: Glycodelin A might be related to tumor aggressiveness and poor clinical outcome in advanced epithelial ovarian cancer. Glycodelin serum levels found in patients suffering from benign ovarian tumors, might contribute to a more global attenuation during progression of these precursor lesions.

Aim: Mucin 1 (MUC1) is a high molecular weight transmembrane glycoprotein with unique properties which is used as a tumour marker in sera of ovarian cancer patients. The common test kit for the cancer antigen 15-3 (CA15-3) is not sufficient for the discrimination between sera from healthy individuals and sera from patients with benign changes of the ovaries. In this study, the newly developed anti-MUC1 antibody PankoMab was tested in normal and patient sera with an ELISA, and the obtained data were compared against data from experiments using the commercial kits for CA15-3 and CA27.29., Materials and Methods: Sera of 123 patients diagnosed with benign or malignant changes of the ovaries were obtained before surgery. CA15-3 was analysed with an automated ELISA system (Immulite 2000). CA27.29 was measured with the ST AIA-PACK CA27.29 for the AIA-600II-Analyzer (Tosoh Bioscience, Belgium). The release of MUC1 fragments carrying the TA-MUC1 epitope was analysed with an ELISA using the PankoMab antibody., Results: Using the already established markers CA15-3 and CA27.29, significant differences between benign and malignant changes of the ovaries were found. The same result was obtained with the newly developed TA-MUC1 test. In contrast to CA15-3 and CA27.29, however, the median of TA-MUC1 was lower in sera from patients with ovarian cancer compared to sera from patients with benign diseases of the ovary. However, sera of patients with benign ovarian diseases had significantly higher TA-MUC1 values compared to sera of healthy individuals. The risk score of TA-MUC1 achieved an area under the curve (AUC) of 78.4% in receiver operating characteristic (ROC) curves and a sensitivity of 37% for the prediction of ovarian disease, at 95% specificity., Conclusion: In this study we employed an additional marker for MUC1 which recognizes a more tumour-specific MUC1 epitope (TA-MUC1). We obtained results showing significant differences between detection in benign and malignant ovarian diseases. Although the mean MUC1 values were elevated in sera of patients with ovarian cancer compared to values of patients with benign cysts, by using all three test systems, a different result was found by analysing the median TA-MUC1 values. PankoMab could be a useful, additional tool for obtaining conclusive information on the transformation process from benign to malignant state in ovarian tissues.

Aim: PankoMab is a novel antibody that recognizes a tumor-specific epitope of Mucin 1 (MUC1). The aim of this study was the evaluation of PankoMab as a potential diagnostic tool and its comparison with two established antibodies against MUC1 in human ductal breast cancer., Materials and Methods: Breast carcinomas were obtained from 82 patients. MUC1 expression and hormone receptor status were determined by immunohistochemistry of paraffin-embedded material., Results: PankoMab revealed strong correlation to hormone receptor expression. DF3 showed no correlation with grading, lymph node involvement and/or estrogen receptor (ER) expression. In the subgroup of lymph node-positive and ER-negative tumors, we saw a significantly reduced DF3 staining in G3 tumors compared to G2 tumors. VU-4-H5 showed increased staining intensity in correlation with increased grading. In addition, we also identified a significantly higher expression of the VU-4-H5 epitope in lymph node-positive carcinomas compared to carcinomas without lymph node involvement., Conclusion: PankoMab revealed strong correlation to hormone receptor expression in ductal carcinoma of the breast. VU-4-H5 showed increased staining intensity in correlation with increased grading and lymph node involvement. PankoMab and VU-4-H5 staining could be a useful combination in ductal breast cancer prognosis by immunohistochemistry., (Copyright (c) 2009 S. Karger AG, Basel.)

Inhibins are dimeric glycoproteins, composed of an alpha-subunit (inhibin-alpha) and one of two possible beta-subunits (betaA or betaB), with substantial roles in human reproduction and in endocrine-responsive tumours. The aims of this study were to determine the distribution of inhibin-alpha, -betaA and -betaB subunits in malignant human endometrial tissue and the assessment of an association with specific clinicopathologic tumour features and clinical outcome. A series of 302 endometrial cancer tissue samples were immunohistochemically analysed with monoclonal antibodies against inhibin subunits. The inhibin-alpha subunit showed a significant association with histological grading, surgical staging, lymph node status and diabetes in patients with endometrial cancer. Interestingly, loss of inhibin-alpha expression resulted in a poorer survival of endometrial cancer patients. Additionally, survival analysis demonstrated that inhibin-alpha immunoreactivity was an independent prognostic factor for progression-free survival, cause-specific survival as well as for overall survival. In contrast, although inhibin-betaA- and -betaB subunits showed a significant association between endometrial histological subtypes and histological grading, both subunits were not found to be associated with survival in endometrial cancer patients. Therefore, inhibin-alpha immunostaining might be used as a simple and efficient marker to identify high-risk patients leading to the selection of patients for an aggressive adjuvant therapy.

Glycodelin (Gd), previously known as placental protein 14 (PP 14), acts as an immuno-suppressive glycoprotein by suppressing the cytolytic capacity of human natural killer (NK) cells and T-cells in vitro. Glycodelin is expressed in normal glandular epithelium of the endometrium as well as in normal and malignant glandular cells in and outside of the reproductive tract. Recently, Gd expression was demonstrated in normal and cancerous human breast tissue. Paraffin-embedded breast cancer tissue blocks (n=121) were examined for Gd expression. No part of the specimens contained carcinoma in situ. Gd expression was present in lobular and ductal breast carcinoma. We observed expression of Gd in breast cancer independent of grading. With regard to nodal status, no significant differences in the expression of Gd between cancer tissue from patients with or without axillary lymph node metastases were present. However, Gd expression was found to be significantly higher in breast cancer tissue when the staining reaction for steroid receptors was also positive. These results implicate that Gd might be an additional marker for the differentiation of breast cancer tissue. To which extent Gd could serve as an additional indicator for breast cancer survival is part of our ongoing research.

Background: The aim of this study was to assess the prognostic value of cancer antigen (CA) 125 and CA 72-4 in patients with ovarian borderline tumor (BOT)., Methods: All women diagnosed and treated for BOT at our institution between 1981 and 2008 were included into this retrospective study (n=101). Preoperatively collected serum samples were analyzed for CA 125 (Architect, Abbott and Elecsys, Roche) and CA 72-4 (Elecsys, Roche) with reference to clinical data and compared to healthy women (n=109) and ovarian cancer patients (n=130)., Results: With a median of 34.7 U/mL (range 18.1-385.0 U/mL) for CA 125 and 2.3 U/mL (range 0.2-277.0 U/mL) for CA 72-4, serum tumor markers in BOT patients were significantly elevated as compared to healthy women with a median CA 125 of 13.5 U/mL (range 4.0-49.7 U/mL) and median CA 72-4 of 0.8 U/mL (range 0.2-20.6 U/mL). In addition, there was a significant difference compared with ovarian cancer patients who showed a median CA 125 of 401.5 U/mL (range 12.5-35,813 U/mL), but no difference was observed for CA 72-4 (median 3.9 U/mL, range 0.3-10,068 U/mL). Patients with a pT1a tumor stage had significantly lower values of CA 125 but not of CA 72-4 compared with individuals with higher tumor stages (median CA 125 29.9 U/mL for pT1a vs. 50.9 U/mL for >pT1a; p=0.014). There was a trend for increased concentrations of CA 125 but not of CA 72-4 in the presence of ascites, endometriosis or peritoneal implants at primary diagnosis. With respect to the prognostic value of CA 125 or CA 72-4, CA 125 was significantly higher at primary diagnosis in patients who later developed recurrence (251.0 U/mL vs. 34.65 U/mL, p=0.012)., Conclusions: Serum CA 125 and CA 72-4 concentrations in BOT patients differ from healthy controls and patients with ovarian cancer. CA 125, but not CA 72-4, at primary diagnosis correlates with tumor stage and tends to be increased in the presence of ascites, endometriosis or peritoneal implants. Moreover, CA 125 at primary diagnosis appears to have prognostic value for recurrence.

Background: The aim of the present prospective case-control study was to evaluate the putative relevance of circulating microparticles (MP) as a biomarker in breast cancer patients., Materials and Methods: Endothelial cell-(EMP) and leukocyte-derived MP (LMP) were determined by flow cytometry in breast cancer patients (n = 41) and healthy controls (n = 25) and compared to carcinoembryonic antigen (CEA), cancer antigen (CA)15-3 and von Willebrand factor antigen (vWF) levels by specificity-sensitivity profiles., Results: LMP, CEA and CA15-3 levels differed significantly between breast cancer patients and controls, whereas EMP and vWF did not. The specificity-sensitivity profiles of LMP and CA15-3 were similar., Conclusion: Increasing levels of circulating LMP (CD45+), CEA and CA15-3 correlated with increasing tumor size, thus reflecting disease stage. LMP showed an equal specificity-sensitivity profile to the established marker CA15-3 and therefore might have the potential to become a new biomarker in breast cancer patients.

Purpose: The presence of disseminated tumor cells in the bone marrow (DTC-BM) of breast cancer patients has shown independent prognostic impact. Immunomagnetic enrichment of such cells is an approach to increase the number of detected cells with limited sample volume, especially for circulating tumor cells (CTCs) in blood. The Thomsen-Friedenreich (TF) antigen (CD 176) is a specific oncofetal carbohydrate epitope (Galbeta1-3GalNAcalpha-O) expressed on the surface of various carcinomas. Own studies demonstrated a nearly complete TF expression on DTC-BM, indicating its suitability as marker for immunomagnetic enrichment., Methods: BM samples of 65 and peripheral blood samples of 11 breast cancer patients were examined immunocytochemically by staining with the anti-Cytokeratin antibody A45-B/B3 before and after immunomagnetic enrichment. Enrichment was done by incubation with the primary antibody TF 2 (IgM), followed by secondary magnetically labelled rat-anti mouse IgM. Cytospin slides were screened manually by bright-field microscopy., Results: 15/65 pts (23%) showed DTC-BM in primary screening with a median of 2/2 mio cells (range 1-10). By enrichment, a median of 23.3 mio cells (0.8-218) could be analysed, increasing positivity to 72% (47/65 pts) with a med. of 4 DTCs (1-105, P < .0001). Blood from 1/11 pts before and 5/11 (45%) after enrichment showed CTCs (med. 2, 1-20), at a med. of 12.4 mio (2.6-38.5) cells analysed. Comparing BM and blood of the same patients after enrichment, 5 were positive in both compartments, 4 showed DTC-BM without presence of CTCs., Conclusion: The positive immunomagnetic enrichment technique with TF-antibodies enables to analyse larger sample volumes and increase tumor cell detection rate. This could allow monitoring and characterisation of CTCs as targets for therapies.

The expression of the classic steroid receptors ERalpha and PR-A has been correlated with stage, histological grade and survival in endometrial cancer. Endometrial cancer samples (293) were immunohistochemically analysed with monoclonal antibodies against the four steroid receptors. The loss of ERalpha, PR-A and PR-B resulted in a poorer survival in endometrial cancer patients, while ERbeta expression did not demonstrate any correlations with several analysed clinicopathological characteristics and did not affect survival. Additionally, multivariate survival analysis demonstrated that PR-B was a significant independent prognostic factor for cause-specific survival. In contrast, although ERalpha and PR-A showed a significant association between different endometrial histological subtypes and grading, both receptors were not independent factors with survival in endometrial carcinoma patients. Therefore, the PR-B immunostaining might be used as an easy, simple and highly efficient marker to identify high-risk patients and may aid in the selection of patients for a more aggressive adjuvant therapy.

Background: Breast cancer cells can invade and generate metastasis via either lymphatic or blood vessels. E-cadherin mediates tumor cell-cell adhesion. Partial or complete loss of E-cadherin expression correlates with poor prognosis in breast cancer patients. In this study, the expression of E-cadherin was examined in mammary ductal carcinoma in situ, invasive breast carcinomas without metastasis, invasive carcinomas with their lymph node and distant metastases and invasive carcinomas with local recurrence in breast cancer tissue., Materials and Methods: Paraffin-embedded slides of carcinoma in situ (8 DCIS), invasive carcinomas without lymph node metastases (9 invasive ductal carcinomas), invasive carcinomas (7 invasive ductal carcinomas) with corresponding lymph node metastases, invasive carcinomas (8 invasive ductal carcinomas) with corresponding recurrence and invasive carcinomas (5 invasive ductal carcinomas) with corresponding distant metastases were investigated for E-cadherin expression. Tissue slides were incubated with monoclonal antibodies against E-cadherin and stained with the ABC-elite kit. Staining intensities were analyzed by using a semi-quantitative score., Results: A strong expression of E-cadherin in carcinoma in situ was demonstrated. Expression of E-cadherin was moderate in invasive carcinomas without metastases. However, very weak expression of E-cadherin in primary carcinoma with lymph node metastases was detected. E-cadherin expression was elevated in lymph node metastases compared to the primary tumor., Conclusion: Analysis of a tumor antigen involved in adhesion of breast cancer cells showed that there are significant differences of expression of E-cadherin between primary breast cancer cells and their metastases. Evaluation of this marker involved in cell adhesion could be a useful method for evaluating the metastatic risk in breast cancer patients.

Background and Aim: Endometrial carcinoma is the most common female pelvic genital malignancy. The strong association between the development of endometrial cancer and influence of steroid hormones (especially estrogen) was demonstrated in many studies. Mucinous carcinoma is an uncommon type of endometrial carcinoma. Most cancers are low grade and have a relatively good prognosis. The expression of one type of estrogen (ER) and progesterone receptor (PR) has been well documented. Recently, two new types of receptors (ER-beta and PR-B) were demonstrated. The aim of this study was to determine the expression of all four steroid receptors (ER-alpha, ER-beta, PR-A and PR-B) in human mucinous carcinoma of the endometrium., Patients and Methods: An immunohistochemical hormone receptor assay using specific monoclonal antibodies against estrogen receptors (ER-alpha, ER-beta) and progesterone receptors (PR-A and PR-B ) was used to study formalin-fixed and paraffin-embedded slides of 12 patients, diagnosed with primary endometrial mucinous carcinoma of different histological grades (G1 n = 9; G2 n = 3; G3 = 0)., Results: Three types of steroid receptors (ER-alpha, PR-A and PR-B) were frequently expressed in mucinous adenocarcinoma. ER-beta was weakly expressed in only one analyzed case. The immunohistochemical expression of PR-B demonstrated a statistically significant decrease in G1 neoplasms in comparison to G2 (p < or = 0.001)., Conclusion: We demonstrated the expression of four different steroid receptors in mucinous endometrial carcinoma. No significant differences between different histological grades of tumor with respect to ER-alpha and ER-beta expression were observed. Interestingly, a statistically significant increase in expression of PR-B in G2 neoplasms compared to G1 was demonstrated. The higher expression of PR-B in G2 tumors suggests a substantial function of progesterone, and thus progesterone receptor, in the malignant transformation of mucinous endometrial cancer. Therefore, PR-B expression might be utilized as a tumor marker to distinguish between G1 and G2 mucinous tumors. However, additional studies are necessary to evaluate whether these parameters could be used as tumor markers for endometrial cancer.

Aim: The aim of the study was to determine possible pathogenetic factors and molecules which may be used as tumor markers of adenosquamous endometrial carcinoma., Materials and Methods: Eight adenosquamous endometrial carcinomas were immunohistochemically tested with specific monoclonal antibodies for HPV (polyclonal anti-HPV and monoclonal anti-HPV 18), estrogen receptors ER-alpha and ER-beta, progesterone receptors PR-A and PR-B and the inhibin/activin subunits inhibin-alpha, -betaA and -betaB., Results: HPV 18 and the polyclonal HPV antibody was detected in all adenosquamous endometrial carcinomas, both in the endometrioid (n = 7/8) and squamous (n = 8/8) parts of the tumor. Neither ER-alpha or ER-beta were detectable in any tumor, in contrast to PR-A and PR-B which were detected in about half of these tumors (PR-A: n = 5/8 and PR-B: n = 2/8). Inhibin-alpha and -betaB were not detected, while inhibin-betaA was expressed in all adenosquamous carcinomas., Conclusion: The carcinogenesis of adenosquamous endometrial carcinomas was associated with HPV infection. Adenosquamous endometrial carcinomas seem not to be controlled by estrogens. The absence of the expression of the inhibin-alpha subunit suggests a tumor-suppressive function in adenosquamous endometrial tumors. The absence of the expression of the inhibin-betaB subunit, which is probably a marker of differentiation, points to the malignancy of these tumors. The other inhibin subunit, inhibin-betaA, was expressed in all adenosquamous tumors. It remains to be clarified if these parameters can be used as tumor markers.

Background: Cathepsin D (Cath-D), a lysosomal protease, is considered to be involved in the breakdown of the extracellular matrix during the process of tumour metastasis. Its expression in breast cancer in association with known factors of prognosis was investigated in this study., Material and Methods: 120 breast cancer tumours were analysed immunohistochemically and evaluated with the immunoreactive score (IRS). Statistical evaluation was performed using the Mann-Whitney test (p < 0.05)., Results: For nodal-positive tumors we found a statistically significant increase of Cath-D immunohistochemical reaction. Ductal carcinomas showed a significantly higher immunohistochemical reaction compared to lobular carcinomas. We found a non-significant increase from G1 to G2 and G1 to G3, and a non-significant increase of Cath-D expression of all receptor-positive over all receptor-negative tumours., Conclusion: Cath-D expression was found to be associated with known prognostic factors. The clinical relevance of the observed difference depending on the histological type needs further investigation. Cath-D might be a useful marker to discriminate between ductal and lobular subtypes of breast cancer.

Background: Sialyl Lewis x (SLeX), sialyl Lewis a (SLeA), Lewis Y (LeY) and the Thomsen-Friedenreich (TF) antigen are carbohydrate motifs that mediate the adhesion between tumour cells and the endothelium. These antigens are usually not expressed in non-malignant tissue. Overexpression of SLeX and SLeA is combined with poor prognosis and malignant relapse. In this study, we analysed the combined expression of SLeX, SLeA, LeY and TF in normal squamous epithelium tissue of the penis shaft, glans and foreskin and in addition of the vagina and vulva., Materials and Methods: Paraffin-embedded slides of vaginal tissue (8), vulva tissue (8) and penis shaft (8) and glans tissue (8) were fixed and incubated with monoclonal antibodies against SLeX (IgM), SLeA (IgM), LeY (IgM) and TF (IgM). Staining reaction was performed with ABC reagent. The intensity of immunohistochemical reaction on images of the slides was analyzed using a semiquantitative score., Results: Strong focal expression of both sialyl Lewis antigens was found in the uretra of the penis shaft and on epithelial tissue of the glans, and permanent moderate expression of SLeX and SLeA in squamous epithelial tissue of the vagina. Moderate expression of TF was observed in male squamous epithelial tissues of the glans and foreskin and faint expression of TF was found in vulval epithelial tissue. Faint expression of Le Y was observed in female vulval epithelial tissue., Conclusion: Expression of SLeX, SLeA, LeY and especially of the TF antigen in normal non malignant epithelial tissue is surprising and can be explained by the function of this tissue in human reproduction. In addition, moderate TF expression seems to be restricted to epithelial tissue of the penis glans and foreskin.

Inhibins (INH) are dimeric glycoproteins, composed of an alpha-subunit (INH-alpha) and one of two possible beta-subunits (INH-betaA or -betaB), with substantial roles in human reproduction and in endocrine-responsive tumors. The aims of this study were to determine the frequency and tissue distribution of INH-alpha, -betaA and -betaB in normal and malignant endometria. Samples were obtained from normal (n=46), atrophic (n=8) and endometrioid carcinoma tissue (EC; G1=93; G2=32; G3=14). INH-alpha was significantly higher in normal compared to malignant endometrial tissue, showing a cyclical variation throughout the menstrual cycle. EC G3 did not express this subunit. INH-betaA and -betaB showed specific staining reactions within the tumor cells. The highest intensity of INH-betaA was observed in the normal secretory phase compared to adenocarcinomas (p<0.05). For INH-betaB, the significantly highest expression was noted in EC G3 compared to EC G2 (p<0.05) and atrophic endometrial tissue. In conclusion, INH-alpha, -betaA and -betaB were immunolabeled in normal and malignant endometria. INH-alpha was expressed in a declining relationship in the transition from normal to tumor tissue, suggesting a tumor suppressive function in EC. A high expression of INH-betaB was observed in EC G3 compared to G2, suggesting an important role in the progression of endometrial carcinogenesis. However, the utilization of these subunits as specific tumor markers still remains unclear.

Background: Sertoli-Leydig cell tumours of the ovary account for only 0.2% of malignant ovarian tumours. Two-thirds of all patients become apparent due to the tumour's hormone production., Methods: A 41-year-old patient (gravida 4, para 4) presented with dyspnoea, enlarged abdominal girth and melaena. Diagnostic imaging was suspicious for an ovarian cancer. The standard tumour marker for ovarian cancer (CA 125) was elevated to 984 U/mL., Results: Surgical exploration of the abdomen revealed a mouldering tumour of both adnexes extending to the level of the navel. Frozen sections showed an undifferentiated carcinoma of unknown origin. Radical surgery was performed. The final histological report described a malignant sex-cord stroma tumour, a Sertoli-Leydig cell tumour, emanating from both ovaries. Analysis of preoperative blood serum showed elevated levels of CYFRA 21-1 (10.4 ng/mL), neuron-specific enolase (36.2 ng/mL), oestradiol (485 pg/mL) and CA-125 (984 U/mL). Adjuvant chemotherapy and regional hyperthermia were performed due to the malignant potential and incomplete resection of the tumour., Conclusions: Undifferentiated Sertoli-Leydig cell tumours show a poor clinical course. As only two-thirds of patients with this rare disease present with elevated hormone levels, new markers deserve further investigation to offer more specific, individualised tumour monitoring.

Glycodelin, previously known as PP14, has been localized in endometrial, ovarian and cervical carcinoma cells. Recently, glycodelin was demonstrated to be expressed in cancerous human breast tissue. In this study, paraffin-embedded slides of carcinoma in situ, invasive carcinomas without metastases, invasive carcinomas with corresponding lymph node metastases, invasive carcinomas with corresponding recurrence and invasive carcinomas with corresponding distant metastases were investigated for glycodelin protein and mRNA expression. Protein expression was found in all cases of carcinoma in situ, in invasive carcinoma without lymph node metastases in 90% of cases, in breast cancer with lymph node metastases in 50% of cases, in breast cancer with recurrence in 38% of cases and in breast cancer with distant metastases in 40% of cases. Results were confirmed by in situ hybridization showing reduced glycodelin expression as lymph node metastasis progressed, compared to carcinoma in situ. Glycodelin mRNA expression is not further reduced in carcinomas with distant metastasis and recurrence compared to carcinoma in situ. Results demonstrate that invasive breast carcinomas without metastases are more likely to express glycodelin. In contrast, cases of breast cancer with metastatic infiltration and recurrence show weak expression of glycodelin. On the basis of these results, we speculate that glycodelin could be used as a prognostic marker for breast cancer.

Objectives: Determination of the frequency and tissue distribution of inhibin-alpha (INH-alpha) in normal, hyperplastic and malignant endometrium., Materials and Methods: Endometrial tissue was obtained from normal, hyperplastic (glandular-cystic hyperplasia), adenomatous hyperplasia (AH grade I to III) and endometroid adenocarcinoma and immunohistochemically characterized with INH-alpha antibody., Results: INH-alpha was expressed in normal, hyperplastic and malignant endometrium. Highest expression was observed during secretory phase and glandular-cystic hyperplasia compared to all groups. A continuous decline was noted from AH grade I to III with a significance between AH I and II., Discussion: A menstrual cycle associated expression of INH-alpha in glandular endometrial epithelium was observed. Since AH grade II and III can be considered as a precursor of endometrial cancer, INH-alpha could be a marker of cell transformation and an endometrial tumor-suppressor agent.

Background: The objective of the present study was to examine the prognostic and predictive significance of the apoptosis-related marker Fas ligand (FasL):Fas ratio in breast cancer., Methods: Tumour biopsies from 215 primary invasive breast cancer patients were examined for the expression of FasL and Fas mRNA transcripts by quantitative real-time RT-PCR. Their prognostic and predictive impact on patient survival was determined in univariate and multivariate survival analyses., Results: Using a cutoff value of 1, a FasL:Fas ratio greater than 1 was found to have significant prognostic value for disease-free survival among the total population (median follow up 54 months). It was associated with a significantly decreased disease-free survival (P = 0.022) and with a tendency toward increased mortality (P = 0.14) in univariate analysis. Hormone receptor positive women exclusively treated with tamoxifen (n = 86) and with a FasL:Fas ratio greater than 1 had a significantly decreased disease-free survival (P = 0.008) and overall survival (P = 0.03) in univariate Kaplan-Meier analysis. Furthermore, tumour size and FasL:Fas ratio were of independent predictive significance in the multivariate model for disease-free and overall survival in that subgroup. Among postmenopausal patients (n = 148) both of those factors retained independent prognostic significance in the multivariate model for disease-free survival. In contrast, FasL:Fas ratio had no significant predictive value in patients exclusively treated with chemotherapy., Conclusion: The data presented indicate that FasL:Fas ratio may be useful not only as a prognostic factor but also as a predictive factor for projecting response to the antioestrogen tamoxifen. The results strongly support a correlation between FasL:Fas ratio greater than 1 and lack of efficacy of tamoxifen in hormone receptor positive patients.