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Breast cancer is the most frequently diagnosed malignant tumor in women. Due to the high heterogeneity and multiplicity of histological subtypes within this type of cancer, the expression of breast cancer markers is very diverse. Therefore, a biomarker with high sensitivity and specificity would be extremely important for the correct diagnosis and prognosis assessment in breast cancer patients. Mammaglobin A seems to be such a biomarker. Overexpression of this protein is closely related with carcinogenesis in the mammary gland and is observed in up to 80% of cases of the malignant breast cancers. According to many reports, it is postulated that mammaglobin A may be a promising tool in the diagnostics of cancers but also a prognostic, predictive and therapeutic factor. The information contained in this publication presents the current state of knowledge about the structure of mammaglobin A, its function, role in the tumorigenesis and the use of this protein as a diagnostic marker and therapeutic target in breast cancer.

Objective: Nasal Polyposis (NP) is a chronic inflammatory disease and genetic factors play an important role in the pathophysiology. Mammaglobin-A (MGA) gene expression was significantly higher in patients with NP and chronic rhinosinusitis compared to normal mucosa. In the present study, we investigated the relationship between single nucleotide polymorphisms (SNPs) in the MGA gene and nasal polyposis in the Turkish population. Materials and Methods: A total of 87 patients diagnosed with NP and 60 healthy volunteers were enrolled in the study. Genotypes of MGA promoter SNPs c38C>G, c.21C >T, c55+186G>A and c.243+230A>T were determined by light SNP ASSAY after real time PCR analysis using genomic DNA samples obtained from the peripheral blood samples of all participants. Results: A total of 87 NP patients, 51 male and 36 female, with a mean age of 38.18±9.5 years were included in the study. No significant difference was determined at all positions c38C>G, c.21C >T, c55+186G>A and c.243+230A>T in nasal polyp patients compared to controls with and without allergic rhinitis (AR). Conclusion: MGA gene c38C> G, c.21C> T, c55 + 186G> A, and c.243 + 230A> T genotypes did not appear to be associated with susceptibility to NP with and without AR in our study population.

Increased microvessel density (MVD) values in breast cancer correlate with tumor growth and progression while mammaglobin (MGB) expression in tumor cells is associated with a favorable prognosis. We aim to evaluate and correlate MVD values with MGB expression in molecular types of breast cancer specimens and to determine their utility as prognostic biological markers. A number of 52 breast cancer specimens were included in the study. Specimens were processed for routine histopathological diagnosis followed by the molecular classification by means of estrogen (ER), progesterone (PR) and HER2 immunohistochemical reactions. After performing immunohistochemistry for CD34 and MGB, MVD evaluation was made using the �hot spot� method for each case and MGB was scored between 0 (negative) and +3 (strong positive) depending on the intensity and distribution of the staining. MGB expression in tumor cells and MVD mean values were extremely variable. The greatest MVD mean values were obtained in luminal B followed by HER2, luminal A and triple negative breast cancer (TNBC) (95.33, 69, 62, and 40, respectively). MGB expression in the tumor cells generally ranged from mild to weak and was strong only in a few invasive ductal carcinoma cases. In cases with TNBCs the expression of MGB in tumor cells was weak and focal or negative. This variability was noticed between the molecular types of breast cancers and even within the same molecular type. In a restricted number of cases, MGB positive tumors were associated with low MVD values while the negative cases were characterized by increased MVD mean values. The variable results we obtained regarding the correlation between MVD and MGB in breast cancer specimens may indicate a rather restricted use of MVD/MGB in estimating breast cancer patients� prognosis.

We demonstrate high-sensitivity detection of metastatic breast cancer cells using terahertz (THz) chemical microscopy (TCM) with aptamers as ligands. For the aptamers, we use previously developed synthetic single stranded (ss) DNA aptamers mammaglobin B1 (MAMB1) and mammaglobin A2 (MAMA2) that bind to mammaglobin B and mammaglobin A proteins, respectively, which are overexpressed on the surface of MCF7 and MDA-MB-415 breast cancer cells. Each aptamer was immobilized on the surface of a sensing plate, and the amplitude of the THz signal was measured upon the binding of each aptamer to different number (10–106) of its target breast cancer cells. A change in the THz amplitude was observed when MAMB1 and MAMA2 bind to MCF7 and MDA-MB-415, respectively. We find that this change was linear as a function of the log number of breast cancer cells used. No change in the THz amplitude was observed when the same number of normal breast cells (MCF 10A) were used. Moreover, MAMB1 and MAMA2 did not show binding to the counter breast cancer cells, indicating high selectivity. We have demonstrated that the TCM using aptamers as ligands has a limit of detection as small as 1 breast cancer cell in 100 μL of sample. Results described here indicate that the TCM could be a powerful tool to detect metastatic breast cancer cells.

Objectives: Breast cancer cells that are released into the bloodstream are called circulating tumor cells (CTCs). CTCs can express different genes, like TWIST-1 and mammaglobin A (MGA). The aims of this study were to analyze the expression of TWIST-1 and MGA in the blood of breast cancer patients to detect CTCs and to assess the association between the presence of CTCs and prognostic parameters of breast cancer. Methods: Prospective study. Total ribonucleic acid (RNA) from blood mononucleated cells was obtained from breast cancer patients (n = 36; age: 51.5 ± 12.5 years) and healthy donors (n = 14; age: 49.4 ± 9.4 years). Real-time polymerase chain reaction (RT-PCR) was performed to analyze the expression of TWIST-1 and MGA. Results: Patient carcinomas - ductal (86.7%), other types (13.3%). MGA gene expression was not detected in the donors’ samples, while it was detected in 14% of the patient samples. Overexpression of TWIST-1 gene was observed in 17% of the patient samples. The combined analysis of both markers allowed the detection of CTCs in 27.8% of the samples, resulting in a significant (p < 0.05) sensitivity increase of detection. No significant associations (p > 0.05) were found between expression of the analyzed genes and the breast cancer prognostic factors. Conclusion: Combined analysis of TWIST-1 and MGA increased the sensitivity of CTCs detection compared to the single analysis of each gene. The detection of CTCs was not associated with known prognostic factors, suggesting that it is able to provide clinical information in addition to routine breast cancer clinicopathological parameters. RESUMEN Objetivos: Las células de cáncer de mama liberadas al torrente sanguíneo se llaman células tumorales circulantes (CTCs). Las CTCs pueden expresar diferentes genes, como TWIST-1 y mamaglobina A (MGA). Los objetivos de este estudio fueron analizar la expresión de TWIST-1 y MGA en la sangre de pacientes con cáncer de mama (CM) para detectar CTCs y evaluar la asociación entre la presencia de CTCs y los parámetros pronósticos del CM. Métodos: Estudio prospectivo. Se obtuvo el ácido ribonucleico (ARN) de las células mononucleadas en la sangre de pacientes con CM (n = 36, edad: 51,5 ± 12,5 años) y donantes sanas (n = 14; edad: 49,4 ± 9,4 años). Se realizó reacción en cadena de la polimerasa en tiempo real (RT-PCR) para analizar la expresión de TWIST-1 y MGA. Resultados: Carcinoma ductal (86,7%), otros tipos (13,3%). No se detectó la expresión del gen MGA en las muestras de las donantes, pero en el 14% de las muestras de las pacientes. Se observó elevada expresión de TWIST-1 en el 17% de las muestras de pacientes con CM. El análisis combinado de ambos marcadores permitió detección de CTCs en el 27,8% de las muestras, resultando en un aumento significativo (p < 0,05) en la sensibilidad de detección. No se encontraron asociaciones significativas (p > 0,05) entre la expresión de los genes y los factores pronósticos. Conclusión: El análisis combinado de TWIST-1 y MGA aumentó la sensibilidad de detección de CTCs en comparación con el análisis de cada gen. La detección de CTCs no se asoció a factores pronósticos conocidos, sugiriendo que podría ofrecer informaciones clínicas adicionales a los parámetros clínico-patológicos de rutina del CM. RESUMO Objetivos: As células cancerígenas da mama liberadas na corrente sanguínea são chamadas de células tumorais circulantes (CTCs). As CTCs podem expressar diferentes genes, como TWIST-1 e mamaglobina A (MGA). Os objetivos deste estudo foram analisar a expressão de TWIST-1 e MGA no sangue de pacientes com câncer da mama (CM) para detectar CTCs e avaliar a associação entre a presença de CTCs e os parâmetros prognósticos do CM. Métodos: Estudo prospectivo. O ácido ribonucleico (RNA) das células mononucleadas no sangue foi obtido de pacientes com CM (n = 36, idade: 51,5 ± 12,5 anos) e doadoras saudáveis (n = 14; idade: 49,4 ± 9,4 anos). Reação da cadeia da polimerase em tempo real (RT-PCR) foi realizada para analisar a expressão de TWIST-1 e MGA. Resultados: Carcinoma ductal (86,7%), outros tipos (13,3%). A expressão do gene MGA não foi detectada nas amostras das doadoras, mas foi observada em 14% das amostras das pacientes. Superexpressão de TWIST-1 foi observada em 17% das amostras dos indivíduos com CM. A análise combinada de ambos os marcadores permitiu a detecção de CTCs em 27,8% das amostras, resultando em um aumento significativo (p < 0,05) na sensibilidade da detecção. Associações significativas (p > 0,05) entre a expressão dos genes e os fatores prognósticos não foram encontradas. Conclusão: A análise combinada de TWIST-1 e MGA aumentou a sensibilidade da detecção de CTCs em comparação com a análise de cada gene. A detecção de CTCs não foi associada a fatores prognósticos conhecidos, sugerindo que ela pode fornecer informações clínicas adicionais aos parâmetros clinicopatológicos de rotina do CM.

Although targeted therapy has been extensively investigated for breast cancers, a molecular target with broad application is currently unavailable due to the high heterogeneity of these cancers. Mammaglobin-A (Mam-A), which is overexpressed in most breast carcinomas, has been proposed as a promising target. However, the lack of specific targeting moieties due to uncertain binding epitopes hampers further translational study. Here, seven potential epitopes of Mam-A were disclosed, and a unique epitope was then identified in most types of breast cancers, despite the genotypic heterogeneity. With phage display technology, the epitope was determined to be N-terminal amino acids 42–51 of Mam-A (N42–51). Then, the N42–51 epitope-specific monoclonal antibody, mAb785, was conjugated to poly lactic-co-glycolic acid (PLGA) nanoparticles loaded with therapeutic agents, thereby enhancing the drug uptake and therapeutic efficacy in different genotypes of breast cancers. The computer simulation of the N42–51 epitope and the mAb785 structures, as well as their interactions, further revealed the specific targeting mechanism of the mAb785-conjugated nanoparticles to breast cancers.

Objective: Cytomorphologic differentiation of metastatic breast carcinoma from non breast metastases in cytological materials can be difficult. Current breast immunocytochemical markers have low sensitivities. Transcription factor GATA 3 is a promising marker for detecting breast differentiation in cytological materials. The aim of the study was to assess the diagnostic value of GATA 3 as a breast differentiation marker in metastatic cytological materials and to compare it with expression of mammaglobin and gross cystic disease fluid protein-15 (GCDFP-15). Material and Method: We retrospectively retrieved 133 cases of metastatic breast carcinoma from the archive the of Cytology Unit between December 2013 and June 2015. They included 77 fine needle aspiration and 56 serous effusion samples. Forty-five cytological materials from non mammary metastatic tumors were used as a control. Immunostaining was performed on cell blocks for the presence of GATA 3, mammaglobin and GCDFP-15. Results: GATA 3 nuclear staining was detected in 82.7% of metastatic breast carcinomas, and 11.1% of metastatic non mammary adenocarcinomas (p < 0.001). GATA 3 sensitivity, specificity, positive predictive value, negative predictive value and accuracy were 82.7%, 88.9%, 95.7%, 63.5% and 84.3%, respectively. Mammaglobin and GCDFP-15 staining of metastatic breast carcinoma cases was positive in 70.7% and 47.1%, respectively. GATA 3 staining was significantly higher compared with mammaglobin and GCDFP-15 (p< 0.001). Conclusion: GATA 3 is more sensitive marker than mammaglobin and GCDFP-15 for diagnosing metastatic breast carcinoma in cytological cell block materials. Adding mammaglobin to GATA 3 resulted in improvement in its sensitivity. GATA 3 was occasionally positive in some metastatic non mammary carcinoma that may cause misdiagnosis.

Mammaglobin-A (MamA) is overexpressed in 40&ndash, 80% of all human breast cancers. Recent phase I clinical trials of the MamA DNA vaccine showed encouraging safety outcomes. However, this vaccine elicited only a modest increase in MamA specific CD8+T lymphocyte (CTL) activation. As vaccine adjuvants play a critical role in enhancing the immunotherapeutic efficiency of vaccines, we tested the potential role of three synthetic CpG oligodeoxynucleotides (ODN2216&mdash, class A ODN, ODN2006&mdash, class B ODN, and ODN M362&mdash, class C ODN) to further enhance MamA specific CTL responses. Towards this, na&iuml, ve CD8+T cells were obtained from healthy HLA-A2+ human donors. The HLA-A2 specific immunodominant epitope of MamA, MamA2.1 (LIYDSSLCDL), was utilized to activate na&iuml, ve CD8+T cells. The THP-1 (HLA-A2+) cells were used as antigen presenting cells to stimulate na&iuml, ve CD8+T cells along with (or without) co-treatment of various ODNs mentioned above. Activation of na&iuml, ve CD8+T cells with the MamA2.1 peptide along with ODNs demonstrated enhanced MamA specific CTL mediated cytotoxicity on AU565 (HLA-A+/MamA+) breast cancer cells following co-treatment with ODN2006 and M362 compared to ODN2216 or MamA2.1 peptide alone. However, no significant cytotoxicity was noted upon treatment of MamA2.1 activated CTLs on MCF7 (HLA-A+/MamA&minus, ) cells, suggesting that the activation of CTLs is specific to the MamA antigen. Functional characterization studies demonstrated specific IL-12 mediated cross-talk between TLR-6 and -9 in THP-1 cells following stimulation with ODN2006 and M362, which was critical for the final cytotoxic activation of CD8+T lymphocytes. Based on these data, we conclude that ODN2006 and ODN M362 exerted a strong adjuvant effect through induction of the initial innate immune response through TLR9 upregulation followed by enhanced MamA specific CTL dependent adaptive immune responses. Our current data provide evidence for the application of Class-B/-C-CpG-ODNs as potential vaccine adjuvants towards enhancing the success of MamA based breast cancer vaccination.

The aim of the current study was to assess the prognostic value of circulating tumor cells (CTCs) and their related markers at different points of chemotherapy regimens in metastatic breast cancer (MBC) patients. The impact of CTCs on progression free survival (PFS) and overall survival (OS) rates were also assessed. Peripheral blood samples were obtained from 66 female patients with MBC at different time intervals for evaluation of CTCs by flow cytometry (FC). cytokeratin 19 (CK19), mammaglobin, prolactin inducible peptide (PIP), aldehyde dehydrogenase 1 (ALDH1) and human chorionic gonadotropin (hCG) were also assessed by qRT-PCR. Analysis of different CTC levels (at 4, 5, and 6 cells/7 ml), showed statistically significant values at 4 cells/7 ml blood. The presence of baseline CTCs

Mammaglobin A expression in peripheral blood (PB) of breast carcinoma patients has been evaluated by various studies, but the findings have been inconsistent. This meta-analysis aimed to clarify the prognostic value of mammaglobin A in the PB of breast carcinoma patients and define its relationships with clinicopathological features. PubMed, EMBASE, and the Cochrane Library databases were systematically searched for eligible studies through September 26, 2017. A total of 20 studies involving 2,323 patients were analyzed, and the data were independently extracted by two researchers. The combined hazard ratios (HRs) with 95% CI was used to assess the association between survival data and plasma mammaglobin A expression, and odds ratios (ORs) and 95% CIs were used to assess the associations between clinicopathological parameters and plasma mammaglobin A expression. The results indicated that plasma mammaglobin A expression was a predictor of poor prognosis for breast carcinoma patients, with an HR of 2.08 (95% CI=1.48-2.91; P

Background Mammaglobin A (MGA), mainly expressed in the breast epithelium, is overexpressed in breast cancer, and has been established as a tumor and promissory marker for the early detection of metastasis. Aim The main aim of this study was to evaluate the association between the presence of the MGA transcript in the peripheral blood of Brazilian breast cancer patients and healthy women and the development of breast cancer and tumor progression. Material and methods The expression of the MGA transcript in peripheral blood of 102 breast cancer patients and 102 healthy women was assessed by RT-PCR. Results MGA mRNA was expressed in the peripheral blood of 39 breast cancer patients and in none of the women from the control group. The presence of MGA was significantly associated with presence of metastasis and age at onset after 60 years. The presence of MGA mRNA in peripheral blood displayed a sensitivity of 38.2%, specificity of 100.0%, positive predictive value (PPV) of 100.0%, and negative predictive value (NPV) of 61.8% as a breast cancer marker. Conclusion This study provides additional evidence of the presence of MGA in the peripheral blood of breast cancer patients, and its applicability as an efficient biomarker for breast cancer (High specificity and PPV). To our knowledge, this is the first study to assess the expression of MGA mRNA in peripheral blood obtained from the Brazilian population.

Mammaglobin B (MGB2) and mammaglobin A (MGB1) are proteins expressed in metastatic breast cancers. The early detection of circulating tumor cells (CTCs) in breast cancer patients is crucial to decrease mortality rate. Herein, novel aptamers were successfully selected and characterized against MGB2 and MGB1 proteins using a hybrid SELEX approach. The potential use of the selected aptamers in breast CTC detection was studied using spiked breast cancer cells in whole blood lysate. The results obtained from this study showed that the selected aptamers (MAMB1 and MAMA2) bind to their target breast cancer cell lines with high affinity (low nanomolar Kd values) and specificity. They also bind to their free recombinant target proteins and show minimal non-specific binding to normal and other cancer cell lines. Additionally, they were able to distinguish a low number of breast cancer cells spiked in whole blood lysate containing normal blood cells. The results obtained in this study indicate the great potential for the use of aptamers to detect MGB1 and MGB2 protein biomarkers, expressed on the surface of breast CTCs.

Sentinel lymph node (SLN) micrometstasis detection improves outcome for breast cancer follow up procedure. The aim of the present study was to identify gene profiles that accurately predicted the outcome of breast cancer patients. Fifty tumor sample from breast cancer patients were analyzed for the expression of 3 genes using quantitative-PCR. Also clinical verification for recurrence to distant organs was performed. Three gene signature were confirmed based on tumor's stage, grade, ER status, using conditional logistic regression. Based on this findings, the negative reported lymph nodes for metastasis, had micro metastasis in significant values. There was a significant difference between normal and cancer samples in 3 gene expression marker and also there was meaningful relationship between three gene expression with tumor's grade, stage according to progression of tumor. A novel gene expression signature predictive of micro metastatic patients was evaluated. In this assessment, relationship between this gene with tumor's features that finding clear role for these genes with tumor's outcome, needs to be established.

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.

GATA-3 is a transcription factor that has recently been identified by immunohistochemistry to be highly expressed in urothelial and breast carcinomas (CAs). We sought to determine the potential utility of GATA-3 in identifying metastatic breast CA, and to compare its utility with the standard breast markers, GCDFP-15, and mammaglobin A. We identified an archival series of 338 formalin-fixed paraffin-embedded whole-tissue sections of various CAs. Using standard immunohistochemical (IHC) techniques we used mouse monoclonal antibodies to GATA-3 (clones L50-823, HG3-31), GCDFP-15 (23A3), and mammaglobin A (31A5). Both clones of GATA-3 showed positivity in 96% of non-triple-negative breast carcinomas (TNBCs), L50-823 and HG3-31, demonstrating expression in 87% and 63% of TNBCs, respectively; GCDFP-15 and mammaglobin A were expressed in 69% and 61% of non-TNBCs, respectively, and 10% and 17%, of TNBCs, respectively. The L50-823 clone manifested a lower specificity in identifying breast CAs (84%) than did the HG3-31 clone (97%). Both monoclonal antibodies to GATA-3 are very sensitive reagents for the identification of breast CA, surpassing antibodies to GCDFP-15 and mammaglobin A, and offer a significant improvement in identifying TNBCs. However, the L50-823 clone showed a lower level of specificity, which may qualify its utility in the setting of CAs of unknown primary.

Aims: In addition to oestrogen and progesterone receptors, gross cystic disease fluid protein-15 (GCDFP-15) and mammaglobin A (MAM) are the most common markers used to identify breast origin by immunohistochemistry. GCDFP-15 expression has been reported in approximately 60% of breast carcinomas and MAM expression in approximately 80%. Data on their expression in triple-negative breast cancer (TNBC) are very limited. The aim of this study was to examine the expression of these markers in TNBC to determine their utility in pathological diagnosis. Methods and results: We studied the immunohistochemical (IHC) expression of GCDFP-15 and MAM in 63 primary and 118 metastatic TNBCs. GCDFP-15 staining was present in 14% of primary and 21% of metastatic TNBCs. MAM staining was present in 25% of primary and 41% of metastatic TNBCs. The frequency of expression of GCDFP-15 and/or MAM was 30% in primary and 43% in metastatic TNBCs, and many positive tumours had only focal staining. Conclusions: Staining for GCDFP-15 and/or MAM in triple-negative carcinomas helps to confirm breast origin, but most tumours in this subgroup of breast carcinomas lack expression of either marker.

Background human mammaglobin (hMAM) expression has been reported in pleural effusions (PE). The aim of this study was to assess the clinical relevance of hMAM mRNA in PE from patients who underwent thoracoscopy. Material and methods A total of 288 patients with PE were studied, 155 of which were diagnosed with malignant and 133 with non-malignant diseases by thoracoscopy. Cells from PE were analyzed by nested hMAM RT-PCR. Statistical analyses were performed to evaluate the diagnostic performance parameters (DPP), the association between hMAM expression and benign or malignant status and the relative risk of cancer for patients with negative thoracoscopy showing hMAM positivity. Results hMAM mRNA was found in 68/288 (23.6%) PE samples of which 51 were from the 155 patients diagnosed with malignant diseases and 17 were from the 133 patients diagnosed with non-malignant diseases. A significant correlation between hMAM expression and malignancy was found (OR=3.04) and the DPP were as follows: sensitivity=32.9%, specificity=87.2%, accuracy=58.0%, positive predictive value=75.0% and negative predictive value=52.7%. Among the patients with negative thoracoscopy (n=133), 5/17 (29.4%) hMAM-positive patients had or developed a tumor during the 18-month follow up period, as compared to 10/116 (8.6%) hMAM-negative patients (relative risk of 4.6 for developing a malignancy). Conclusion These findings suggest a possible application of hMAM RT-PCR detection in PE as to identify a false-negative thoracoscopy in non-specific pleuritis.

Background/Methods: This study determines the co-expression of mammaglobin-A, vascular endothelial growth factor receptor-3 (VEGFR3) and Ki67 by immunohistochemistry (IHC) in tissue samples from 80 patients undergoing breast surgery (cancer or benign disease). The tissue expression was compared with the tumour histopathology and Kaplan Meier 5-year survival analysis was performed. Results: Positive breast tissue expression was observed in 53% samples for mammaglobin, 41% Ki67 and 65% VEGFR3 with a significant positive correlation between Ki67 and VEGFR3 co-expression. Ki67 and VEGFR3 expression correlated with the breast tumour grade and Ki67 expression also correlated with oestrogen receptor (ER) status. At 5 years post-operatively, 6/80 patients had died and 3 patients were alive but had cancer recurrence. High Ki67 expression significantly correlated with poor survival (disease-free and overall). Conclusions: In this study, VEGFR3 and Ki67 expression but not mammaglobin-A correlated with breast tumour pathology. Positive Ki67 expression was also associated with a poor 5-year survival outcome.

UNLABELLED Mammaglobin may be a potential serum biomarker for the differential diagnosis of breast cancer. 260 serum samples were collected from 127 untreated breast cancer patients and 133 healthy volunteers to analyze the sera expression of mammaglobin and its implications for both. The expression vector of pGEX-4T-2-Mammaglobin and pBVIL1-Mammaglobin were constructed and transformed into E.coli.HB101 for expression. The mice were immunized with the purified recombinant protein to prepare monoclonal antibody and to detect by ELISA the serum of normal people and breast cancer patients. Recombinant mammaglobin antigen was effectively expressed in E.coli. Two hybridoma cell lines were obtained after the mice were immunized by pGEX-4T-2-mammaglobin. 133 cases of normal serum and 127 cases of breast cancer serum were analyzed by ELISA. The sera expression level of mammaglobin in breast cancer group (average OD value 0.645±0.223) was significantly (p KEYWORDS mammaglobin; cloning expression; monoclonal antibody; serologic study; breast cancer.

To investigate the prognostic relevance of disseminated tumor cells (DTCs) in bone marrow (BM) assessed by a multimarker mRNA panel consisting of TWIST1, cytokeratin 19 (CK19) and human mammaglobin A (hMAM) mRNA, in patients with early breast cancer.TWIST1 (gene name: TWIST1), CK19 (gene name: KRT19), and hMAM (gene name: SCGB2A2) mRNA was quantitated in BM samples from 191 operable breast cancer patients by real-time reverse transcriptase polymerase chain reaction (RT-PCR).Using the highest relative mRNA concentration of TWIST1 in the control population as a cut-off, 5 of the 191 breast cancer patients showed elevated TWIST1 mRNA levels in their BM by real-time RT-PCR. Two of these patients experienced a systemic relapse during a median follow-up of 98 months. Combining these results with previous hMAM and CK19 mRNA quantifications in the same BM samples, 12 (40%) of the 30 patients with BM positive for at least 1 marker (multimarker positive) experienced a systemic relapse as compared with 18 (11%) of the 161 patients with multimarker-negative BMs. The patients with multimarker-positive BM had significantly shorter systemic recurrence-free survival (P.001, log-rank test), breast cancer-specific survival (P.001), and overall survival (P = .03). The prognostic relevance of BM multimarker detection appeared to be independent of adjuvant treatment, although the difference was not statistically significant in the subgroup of patients who received adjuvant chemotherapy. Multivariate analysis demonstrated the BM multimarker panel status to be a strong independent predictor of clinical outcome.Our results demonstrated the prognostic relevance of BM DTCs assessed by a multimarker mRNA panel consisting of TWIST1, CK19, and hMAM in operable breast cancer patients.

Purpose: To investigate the prognostic value of the molecular detection of circulating tumor cells (CTCs) using three markers [cytokeratin 19 (CK19), mammaglobin A (MGB1), and HER2] in early breast cancer. Experimental Design: CK19mRNA+, MGB1mRNA+, and HER2mRNA+ cells were detected using real-time (CK19) and nested (MGB1 and HER2) reverse transcription-PCR in the peripheral blood of 175 women with stage I to III breast cancer before the initiation of adjuvant chemotherapy. The detection of CTCs was correlated with clinical outcome. In 10 patients, immunofluorescence staining experiments were done to investigate the coexpression of cytokeratin, MGB1, and HER2 in CTCs. Results: CK19mRNA+, MGB1mRNA+, and HER2mRNA+ cells were detected in 41.1%, 8%, and 28.6% of the 175 patients, respectively. Patients had one of the following molecular profiles: CK19mRNA+/MGB1mRNA+/HER2mRNA+ (n = 8), CK19mRNA+/MGB1mRNA+/HER2mRNA− (n = 1), CK19mRNA+/MGB1mRNA−/HER2mRNA+ (n = 42), CK19mRNA+/MGB1mRNA−/HER2mRNA− (n = 21), CK19mRNA−/MGB1mRNA+/HER2mRNA− (n = 5), and CK19mRNA−/MGB1mRNA−/HER2mRNA− (n = 98). Double-immunofluorescence experiments confirmed the following CTC phenotypes: CK+/MGB1+, CK+/MGB1−, CK−/MGB1+, CK+/HER2+, CK+/HER2−, MGB1+/HER2−, and MGB1+/HER2+. In univariate analysis, the detection of CK19mRNA+, MGB1mRNA+, and HER2mRNA+ cells was associated with shorter disease-free survival (DFS; P < 0.001, P = 0.001, and P < 0.001, respectively), whereas the detection of CK19mRNA+ and MGB1mRNA+ cells was associated with worse overall survival (P = 0.044 and 0.034, respectively). In multivariate analysis, estrogen receptor–negative tumors and the detection of CK19mRNA+ and MGB1mRNA+ cells were independently associated with worse DFS. Conclusion: The detection of peripheral blood CK19mRNA+ and MGB1mRNA+ cells before adjuvant chemotherapy predicts poor DFS in women with early breast cancer.

Background The causes of nasal polyposis remain unclear. Mammaglobins have been implicated in its pathogenesis. However, their association with the occurrence of nasal polyps in the presence of allergic rhinitis (AR) has not been explored. The aim of this study was to compare the expression levels of mammaglobins A and B with the nasal polyps of patients with and without AR. Methods Thirty-one patients with bilateral nasal polyposis underwent skin-prick tests to specific aeroallergens. Nasal polyp tissues were obtained from all patients and divided into two groups as nasal polyps with and without AR depending on clinical history and the skin-prick test results. All polyp tissues were analyzed for the levels of mammaglobin A and mammaglobin B by using real-time quantitative polymerase chain reaction technique. Results Of the 16 samples from patients having nasal polyps with AR, only 1 sample expressed a detectable level of mammaglobin A (1/16). There was no detectable expression of mammaglobin A in tissues from the group of nasal polyps without AR (0/15). Expression of mammaglobin B was detected in all nasal polyp tissues from both groups. The expression of mammaglobin B was not significantly different between nasal polyps with AR (median, 25th-75th percentiles; 0.023, 0.013-0.046) and nasal polyps without AR (0.032, 0.007-0.16). Conclusion Expression levels of mammaglobins A and B in nasal polyps are not different between patients with and without AR. Our findings suggest that mammaglobins’ implication in the pathogenesis of nasal polyps is independent of an underlying AR.

Existing serum markers for breast cancer such as CA 15-3, BR 27.29 and CEA lack sensitivity and specificity. The aim of this study was to evaluate the value of new putative breast-specific markers for differentiating breast cancer from non-breast tissues. Expression of mammaglobin A (MGA), B726P, small breast epithelial mucin (SBEM) and MUC1 was measured by RT-PCR. MGA mRNA was detected in 86/162 (60%) breast cancers but in only 1/32 (3%) non-breast tissues; B726P was detected in 44/108 (41%) breast cancers but in none of 20 non-breast tissues, while SBEM was present in 52/103 (51%) breast cancers but in only 1/26 non-breast cancer tissues. In contrast to these novel markers, the established breast cancer marker MUC1 was detected in 72/99 (73%) breast cancers and in 22/32 (59%) of non-breast tissues. Combining MGA with B726P separated breast cancer from non-breast tissue with a sensitivity of 71% and a specificity of 95% while combining MGA with SBEM differentiated breast cancer from non-breast tissues with a sensitivity of 76% and a specificity of 89%. Genes such as MGA, B726P and SBEM that are expressed relatively exclusively in breast tissue are potential new markers for breast cancer.

Looking inside the human body fascinated mankind for thousands of years. Current diagnostic and therapy methods are often limited by inadequate sensitivity, specificity and spatial resolution. Raman imaging may bring revolution in monitoring of disease and treatment. The main advantage of Raman imaging is that it gives spatial information about various chemical constituents in defined cellular organelles in contrast to conventional methods (liquid chromatography/mass spectrometry, NMR, HPLC) that rely on bulk or fractionated analyses of extracted components. We demonstrated how Raman imaging can drive the progress on breast cancer just unimaginable a few years ago. We looked inside human breast ducts answering fundamental questions about location and distribution of various biochemical components inside the lumen, epithelial cells of the duct and the stroma around the duct during cancer development. We have identified Raman candidates as diagnostic markers for breast cancer prognosis: carotenoids, mammaglobin, palmitic acid and sphingomyelin as key molecular targets in ductal breast cancer in situ, and propose the molecular mechanisms linking oncogenes with lipid programming.

Objectives The distinction between metastatic breast carcinomas (BCs) and primary lung carcinomas (PLCs) can be difficult. This study tested the utility of trefoil factor 1 (TFF1) for this purpose and compared it with mammaglobin and GATA protein binding 3 (GATA-3). Methods Tissue microarrays containing 365 BCs and 338 PLCs were stained with TFF1, mammaglobin, and GATA-3, and an H-score was calculated. Sensitivity, specificity, and accuracy were calculated, and logistical regression analysis was performed. Results Accuracy of correctly classifying the tumor type was 81.9%, 71.3%, and 64.0% for GATA-3, mammaglobin, and TFF1, respectively. Odds ratios for selecting BCs were 25.69, 93.15, and 4.17, respectively, with P values less than .001. With a single exception, the best immunopanel included GATA-3 and mammaglobin in all comparisons. Conclusions TFF1 demonstrated breast specificity but was inferior to mammaglobin and GATA-3. Therefore, its routine clinical use may not be justified. TFF1 showed little benefit when added to an immunopanel.

“…we are convinced that analysis of the immune response in the primary tumor following vaccination is likely to be significantly more informative than studies of the immune response in the peripheral blood.”

Objective. To identify novel molecular biomarkers useful for the early diagnosis and therapy of ovarian cancer by gene expression profiling. To compare the genetic fingerprints of flash-frozen ovarian serous carcinomas to those of matched highly purified primary tumor cell cultures. Methods. Gene expression profiles of 19 flash-frozen ovarian serous papillary carcinoma (OSPC) were analyzed and compared to 15 controls (highly purified human ovarian surface epithelium short-term cultures, HOSE) using oligonucleotide microarrays complementary to >14,500 human genes. In addition, gene expression profiling of 5 highly purified primary OSPC cultured in vitro for less than 2 weeks was compared to flash-frozen ovarian carcinoma biopsies obtained from matched samples. Quantitative RT-PCR and IHC staining techniques were used to validate microarray data at RNA and protein levels for some of the differentially expressed genes. Results. Unsupervised analysis of gene expression data readily distinguished normal tissue from flash-frozen OSPC and identified 901 and 557 genes that exhibited >3-fold up-regulation or down-regulation, respectively, in OSPC when compared to HOSE. Mammaglobin 2 , an ovarian secreted protein, was identified as the top differentially expressed gene in OSPC (19 out 19 OSPC versus 0 out of 15 HOSE) with over 827-fold up-regulation relative to HOSE. The claudin and kallikrein family of proteins including the clostridium perfringens enterotoxin receptors claudin 3 and 4 , kallikreins 6 , 7 , 8 , 10 , 11 and the immunomodulatory molecule B7-H4 were found among the most highly overexpressed genes in OSPC when compared to HOSE. Genetic fingerprints of flash-frozen OSPC were found to have high correlation with those of purified primary OSPC short-term in vitro cultures with only 31 out of 8637 genes (0.35%) differentially expressed between the two groups. Conclusions. Short-term in vitro culture of primary ovarian carcinomas may greatly increase the purity of ovarian tumor RNA available for gene expression profiling without causing major alteration in OSPC fingerprints. Mammaglobin 2 , kallikreins 6 , 7 , 8 , 10 , 11 , claudin 3 and 4 and B7-H4 gene expression products represent candidate biomarkers endowed with great potential for early screening and therapy of OSPC patients.

Existing serum-based markers for breast cancer all lack organ specificity. Mammaglobin A (MGA) is a 93 amino acid protein expressed almost exclusively in breast tissue. The aim of our study was to investigate the different forms of MGA protein in fibroadenomas and breast carcinomas. MGA protein was measured by Western blotting in 132 breast cancers, 29 fibroadenomas and 14 nonbreast tissues. MGA protein in breast tissue was found to exist in 2 main forms. These forms migrated with approximate molecular masses of 18 and 25 kDa. Both forms of MGA were detected more frequently in breast carcinomas compared to fibroadenomas. The high molecular weight form of MGA but not the low molecular weight form was found more frequently in hormone receptor-positive than in receptor-negative cancers. Furthermore, an inverse relationship was found between the high molecular weight form of MGA and both tumour grade and proliferation index. No significant correlation was found between the MGA proteins and either tumor size or nodal status. Our results show that MGA protein exists in 2 main forms in breast tissue. As the high molecular weight form correlated positively with hormone receptors and negatively with tumor grade and proliferation rate, its presence is likely to be associated with a favourable prognosis for breast cancer. As expression of MGA is almost breast specific, it is a promising marker for breast cancer. Its most immediate use is likely to be in detecting micrometastases from breast cancer. © 2004 Wiley-Liss, Inc.

Purpose The identification of malignant cells in effusions by conventional cytology is hampered by its limited sensitivity. The aim of this study was to improve tumor cell detection in effusions by molecular approaches. Materials and Methods A total of 157 effusions from patients with tumors and 72 effusions from patients without a history or evidence of malignancy were included in this study. All effusion specimens were evaluated in parallel by cytology, fluorescence in situ hybridization (FISH) for aneuploidy, and reverse-transcriptase polymerase chain reaction (RT-PCR) for expression of human mammaglobin (hMAM) and mammaglobin B (hMAM-B). Results In effusions from patients with tumors, the sensitivities of tumor cell detection by cytology, FISH, and hMAM and hMAM-B detection were 46.2%, 53.3%, 36.4%, and 57.7%, respectively. The corresponding specificities were 94.4%, 97.0%, 87.1%, and 88.6%. Notably, a high percentage of effusions containing malignant cells were in fact transudates, indicating the necessity for molecular diagnostic work-up of transudates collected from patients with tumors. Dependent on the tumor type, the use of appropriate marker combinations improved tumor cell detection in effusions significantly. By combining all four diagnostic tests, a positive test result indicating the presence of malignancy was achieved in 81.1%, with a fairly good specificity of 70.1%. Conclusion Molecular techniques are definitely useful to detect malignancy in cytologically negative effusions. Tumor cell detection in effusions can be significantly improved by FISH and PCR techniques applying appropriate molecular markers. This finding should help to improve tumor staging, prognostic assessment, and treatment monitoring.

Purpose The tumor mRNA expression levels of mammaglobin, a novel breast-specific and breast cancer-associated marker, were correlated with disease outcome in 280 patients with primary breast cancer. Patients and Methods Mammaglobin expression levels were assessed by quantitative reverse transcriptase polymerase chain reaction in frozen tumor tissue from breast cancer patients with a median age of 60 years (range, 30 to 88 years) and a median follow-up of 85 months (range, 2 to 169 months). Results High expression levels were associated with low-grade tumors (P = .018), with positive estrogen and progesterone receptor status (P < .001), and postmenopausal status (P = .010). In the analysis of all patients, low tumor mammaglobin expression levels predicted an early relapse both in Cox univariate (hazard ratio [HR], 0.52; 95% CI, 0.34 to 0.79; P = .002) and multivariate regression analyses corrected for the traditional prognostic factors (HR, 0.55; 95% CI, 0.35 to 0.88; P = .012). The association of mammaglobin expression with the rate of relapse was particularly favorable in patients who received adjuvant tamoxifen treatment (HR, 0.35; 95% CI, 0.17 to 0.71; P = .004). Conclusion These results demonstrate that the assessment of the tumor mRNA expression level of the breast-specific protein mammaglobin could be useful to stratify patients for individual adjuvant treatment strategies.

Aims We have demonstrated previously that gross cystic disease fluid protein-15 (GCDFP-15) and mammaglobin A (MAM) are of limited utility in triple-negative breast cancer (TNBC). GATA-binding protein 3 (GATA-3) is an emerging breast-associated immunohistochemical (IHC) marker with limited data in TNBC. Here, we examined GATA-3 expression in TNBC in comparison with GCDFP-15 and MAM. Methods and results We studied GATA-3, GCDFP-15 and MAM IHC expression in 62 primary and 68 metastatic TNBCs. In primary TNBCs, GATA-3 staining was observed in 25 cases (40%), including 16 cases that were negative for GCDFP-15 and MAM. In metastatic TNBCs, GATA-3 staining was observed in 30 cases (44%), including 16 cases that were negative for GCDFP-15 and MAM. The expression frequency of any of the markers was 56% in primary and 62% in metastatic TNBCs. However, when focal staining was excluded, the expression frequency of any marker dropped to 31% and 44%, respectively. Conclusion GATA-3 is expressed at a higher frequency by IHC in TNBC compared to GCDFP-15 and MAM, although the tissue specificity of the latter markers may be superior. When evaluating a triple-negative tumour, including GATA-3 in a panel of markers may increase the diagnostic accuracy for tissue origin in the appropriate clinical setting.

Background Mammary analog secretory carcinoma (MASC) of the salivary gland has been recently described according to morphological, immunohistochemical, and molecular (ETV6-NTRK3 translocation) similarities with the mammary secretory carcinoma. The most important differential diagnostic considerations of MASC are low-grade adenocarcinoma not otherwise specified (NOS), cystadenocarcinoma, and acinic cell carcinoma (AciCC). These tumors may share an overlapping morphology with MASC, and additional immunohistochemical studies are required to reinforce the diagnosis. Mammaglobin, GCDFP-15, and p63 staining have been reported in MASC. Our study was designed to check the specificity of these antibodies in MASC compared to other frequent tumors of salivary glands. Methods A series of 62 salivary gland tumors [10 MASCs, 5 adenocarcinomas NOS and 2 cystadenocarcinomas with MASC features and without ETV6 rearrangement, one low-grade cribriform cystadenocarcinoma (LGCCC), 9 AciCCs, 10 MECs, 10 adenoid cystic carcinomas (AdeCCs), 5 polymorphous low-grade adenocarcinomas (PLGAs), and 10 pleomorphic adenomas (PAs)] was analyzed by immunohistochemistry with mammaglobin, GCDFP-15, and p63 antibodies. Results Positivity for mammaglobin was observed in all MASCs, cystadenocarcinomas, LGCCC, and PLGAs, in some adenocarcinomas NOS, PAs, and MECs, rarely in AciCCs and never in AdeCCs. Positivity for GCDFP-15 was observed in most of the tumor types except in AdeCCs. Interestingly, cytoplasmic positivity for p63 was observed in most of MASCs and PLGAs while rarely in adenocarcinomas NOS and PAs, and never in the other tumor types. Conclusion Our study revealed the usefulness of mammaglobin and p63 cytoplasmic staining to define which tumors are worth to be screened for ETV6 rearrangement.

The mammaglobin gene has been shown to be preferentially expressed in breast tissue. Few genes match its specificity. Mammaglobin has generated much interest, and studies are ongoing to develop diagnostic tests for breast cancer based on the detection of mammaglobin. While searching the Incyte Genomics Lifeseq database for tissue-specific markers, we observed a second secretoglobin, BU101, also known as lipophilin B. We report here that mammaglobin, in breast tissue, is found as a complex with BU101. The complex was isolated from breast cancer tissue and was characterized as the biologically relevant form of mammaglobin.

Background: We used a combination of genetic subtraction, silicon DNA microarray analysis, and quantitative PCR to identify tissue- and tumor-specific genes as diagnostic targets for breast cancer. Methods and Results: From a large number of candidate antigens, several specific subsets of genes were identified that showed concordant and complementary expression profiles. Whereas transcriptional profiling of mammaglobin resulted in the detection of 70% of tumors in a panel of 46 primary and metastatic breast cancers, the inclusion of three additional markers resulted in detection of all 46 specimens. Immunomagnetic epithelial cell enrichment of circulating tumor cells from the peripheral blood of patients with metastatic breast cancer, coupled with RT-PCR-based amplification of breast tumor-specific transcripts, resulted in the detection of anchorage-independent tumor cells in the majority of patients with breast cancer with known metastatic disease. Conclusion: Complementation of mammaglobin with three additional genes in RT-PCR increases the detection of breast cancers in tissue and circulating tumor cells.