High expression of FBP1 and LDHB in fibroadenomas and invasive breast cancers.

BACKGROUND: The role of gluconeogenesis in cancer cells as the reverse pathway for glycolysis is not well known. Several studies of gluconeogenesis in cancer cells still show conflicting results. Expression of key enzymes such as FBP1 and LDHB in cancer tissues may explain the role of gluconeogenesis in tumor development. OBJECTIVE: This study aimed to analyze the expression of FBP1 and LDHB in fibroadenomas and invasive cancers of the breast. METHODS: The immunohistochemical staining technique was used to show the expression of FBP1 and LDHB in formalin-fixed, paraffin-embedded blocks of 32 fibroadenomas and 31 invasive breast cancer samples. RESULTS: FBP1 was expressed by the majority of fibroadenoma (68.7%) and invasive breast cancer (71%) samples. LDHB expression in fibroadenomas was significantly higher than in invasive breast cancers (P = 0.029). The expression of these two enzymes was found in invasive, lobular, and tubular breast carcinoma, and at well, moderately, and poorly differentiated breast malignancy. CONCLUSIONS: High expression of FBP1 and LDHB was found in fibroadenomas and invasive breast cancers. A higher level of LDHB expression was observed in fibroadenomas. These results may indicate the enzymes' role in the pathogenesis of both breast diseases.

Soluble guanylate cyclase isoenzymes: The expression of α1, α2, ß1, and ß2 subunits in the benign and malignant breast tumors.

Soluble guanylate cyclase (sGC) encompasses α and ß subunits. This study examined the expression of α1, α2, ß1, and ß2 subunits in the malignant and benign breast tumors using the Western blot analysis. Both benign and malignant tumors showed a significantly higher expression of the α1 subunit in comparison with normal tissues (p < 0.0001). In contrast, the expression of α2 and ß2 sGC were significantly lower in these tumors than normal tissues (p < .0015 and p < .001, p < .007 and p < .0001, respectively). The expression level of α1 sGC was significantly correlated with ER + PR+ (p < .0001). A significant correlation was also detected for sGC-α1 and -α2 expression with c-erbB2-negative status (p < .01). However, the expression level of sGC was not associated with tumor stage, tumor grade, or other clinicopathological features. In conclusion, as the expression of α1 sGC is upregulated and α2 and ß2 sGC are downregulated in malignant breast tumors. Variations in the expression of sGC isoenzymes may be suggested as an indicator to confirm the enzyme antitumor activity.

Carbonic anhydrase IX: a promising diagnostic and prognostic biomarker in breast carcinoma.

We examined the expression of carbonic anhydrase IX (CA IX) by immunohistochemical staining using monoclonal antibody M75 (Institute of Virology, Slovak Academy of Sciences, Bratislava) in a group of 38 fibroadenomas and 55 carcinomas of the breast. In each case, the intensity of staining, percentage of labeled cells and subcellular localization of CA IX were assessed. CA IX was detected in 11/38 fibroadenomas (28.9%). Weak cytoplasmic positivity was dominant in these positive cases. Immunohistochemical analysis of 55 carcinomas showed CA IX expression in 34 cases (61.8%). Membrane staining alone was observed in 27/55 carcinomas (49.1%), while cytoplasmic positivity was found in 4/55 cases (7.3%). Combined membrane and cytoplasmic immunostaining of CA IX was detected in 3/55 carcinomas (5.4%). The intensity of immunoreactivity varied from weak to strong. Under 50% of reactive cells were found in 9/38 fibroadenomas (23.6%) and in 29/55 carcinomas (52.7%). More than 50% of reactive cells were found in 2/38 fibroadenomas (5.3%) and in 5/55 carcinomas (9.1%). Statistical analysis confirmed significant differences in the subcellular localization, intensity of immunoreactivity and percentage of labeled cells in fibroadenomas and carcinomas (p<0.05). Our results confirmed the hypothesis that expression of CA IX may represent a valuable tumor biomarker as well as a promising diagnostic and prognostic parameter in breast cancer.

Prostate-specific antigen gene expression and telomerase activity in breast cancer patients: possible relationship to steroid hormone receptors.

Breast cancer, the most prevalent cancer among women, is a steroid hormone receptor-dependent cancer. Recently, it has been shown that telomerase and prostate-specific antigen (PSA) gene expressions are under control of steroid hormone receptors. The aim of this study was to investigate the relationship between telomerase activity and PSA gene expression with steroid hormone receptors in breast cancer patients. This study consisted of 50 women with breast benign tumors and 50 malignant (invasive) tumors. Telomerase activity was measured in tumor cytosol of samples by telomeric repeat amplification protocol (TRAP) assay. PSA protein and its mRNA expression were measured using ultrasensitive immunoassay and RT-PCR technique in all tumor tissues, respectively. Estrogen and progesterone receptors were stained using immunohistochemistry in tumor tissues. Telomerase activity was detected in all of the invasive breast cancer tissues. The difference of relative telomerase activity (RTA) values between stages and grades were statistically significant (p < 0.05). The PSA mRNA was detected only in benign tumors and stage I and grade I malignant tumor cytosol. Difference of tumor cytosol PSA levels between the cases and control groups and also between all grades and stages of diseases were significant (p < 0.05). There was an inverse significant correlation between the RTA and PSA protein levels in the case groups (r = -0.42, p < 0.05). There was a statistically significant difference between ER/PR with PSA level and telomerase activity in tumor tissues (p < 0.05). It is speculated that differential expression of PSA and telomerase genes in breast tumors are under control of steroid hormone receptors and could be used as a target for treatment in the future.

[Comparison of plasma matrix metalloproteases 2, 3, 9 in breast carcinomas and fibroadenomas]. / Confronto dei valori plasmatici delle metalloproteasi della matrice 2, 3, 9 in pazienti con carcinomi e fibroadenomi mammari.

Tumor cell infiltration causes the remodelling of peritumoral tissues, determined by an increased lytic activity of extracellular matrix exerted by the neoplastic invasive phenotype. Among the principal lytic enzymes produced by tumor cells and mainly involved in invasion process there are the matrix metalloproteases (MMPs). The Authors compared the plasmatic values of MMPs 2, 3, 9 from patients with breast carcinomas and fibroadenomas in order to evaluate whether there was a significant difference between the two groups of patients. MMPs 2, 3, 9 values were quantified by ELISA test from plasma collected 24 hours before surgery in 50 breast carcinomas and 30 fibroadenomas. MMP2 mean value from the patients with carcinomas resulted significantly higher as compared to that from the patients with fibroadenomas; while for MMP 3 and 9 mean values was not possible to find a significant difference between the two groups of malignant and benign breast tumors. These data confirm the main role played by MMPs during the tumor invasion process. Therefore, it is possible to propose the future inclusion of MMP2 test, together to other biological and clinical data, for prognostic evaluation of neoplastic breast lesions.

DNA oxidation and antioxidant status in breast cancer.

PURPOSE: : Oxidant/antioxidant balance has been suggested as an important factor for initiation and progression of cancer. The objective of this study was to determine 8-hydroxydeoxyguanosine (8-OHdG) level as a marker of oxidative DNA damage, glutathione peroxidase (G-Px), and superoxide dismutase (SOD) activities as antioxidant activity, in sera from women with breast cancer. METHODS: : Forty-nine patients with malign breast tumor were included in the study. Blood samples were collected before the surgical operation. Serum level of 8-OHdG was measured with a competitive enzyme-linked immunusorbent assay kit, SOD, and G-Px activities were measured by spectrophotometric kits. RESULTS: : 8-Hydroxydeoxyguanosine level and SOD activity were found to be increased in breast cancer group as compared with control group. Glutathione peroxidase activity in the breast cancer group was lower than those in the control group. The ratio of 8-OHdG/G-Px in breast cancer patients was found to be higher than those in the controls. There were correlations between 8-OHdG and CA19-9 (r = 0.77; P < 0.01); age and G-Px (r = -0.84; P < 0.05) in the breast cancer group. CONCLUSIONS: : Data show that serum levels of 8-OHdG and SOD activities are higher in patients with breast cancer. Glutathione peroxidase activity is lower in the breast cancer group. Increased ratio of 8-OHdG/G-Px in breast cancer patients is the evidence for impaired oxidant/ antioxidant balance in breast cancer.

[Lactate dehydrogenase, adenosine deaminase and thymidine phosphorylase activity of blood and tissues in breast cancer].

Lactate dehydrogenase (LDH), adenosine deaminase and thymidine phosphorylase activity was analyzed in the blood serum, primary tumor and adjacent uninvolved breast tissues from 49 women with adenocarcinoma and from 10 ones with benign adenofibroma. The LDH activity was increased in both cancerous and adjacent tissues. Serum LDH level reflects cell membrane alterations not only in the tumor node cells but also to a greater extent--in the surrounding unmalignant tissues. The discovered changes in nucleosides catabolic enzyme's activity in patients with breast cancer are correlated with LDH activity and its level in the blood serum.

Expression of nm23, MMP-2, TIMP-2 in breast neoplasm in Zhengzhou Center Hospital, China.

BACKGROUND: In recent years, the carcinoma of the breast threatens to women's health heavily. Invasion and metastasis is the main reason that results in the patients death. OBJECTIVE: A prospective study is made in the center hospital of zhengzhou, in order to approach the expression of nm23, MMP-2 (Matrix metallo-proteinase-2), TIMP-2 (it's tissue-inhibitor of the metalloproteinase-2) in the breast neoplasm and the relationship with invasion and metastasis. METHODOLOGY: This study applied the immunohistochemistry technique SP method RESULTS: In fibroadenoma, ductal carcinoma in situ and invasive ductal carcinoma of the breast 4 groups, the positive immunostaining rate of nm23, MMP-2 and TIMP-2 have significant difference among 4 groups (p < 0.05). In the breast invasive ductal carcinoma, the expression of nm23 and TIMP-2 decreased or the expression of MMP-2 increased CONCLUSION: This suggested that invasion and metastasis is ability of the neoplasm. MMP-2 in the breast ductal carcinoma in situ appears of high expression and this suggested that the positive expression of this onco-proteins was the early incident in the genetic course of the breast cancer The unite detection of nm23, MMP-2 and TIMP-2 expression would contribute to the early diagnosis and prognostic assessment of the carcinoma of the breast.

Expression of the serine protease, matriptase, in breast ductal carcinoma of Chinese women: correlation with clinicopathological parameters.

Matriptase is a serine protease expressed by cells of surface epithelial origin, including epithelial breast tumor cells. Matriptase cleaves and activates proteins implicated in the progression of cancer and represents a potential prognostic and therapeutic target. The aim of this study was to examine matriptase expression in breast tumors of Chinese women and to identify its clinicopathological correlations. Immunohistochemical analysis of matriptase was performed in tissue microarrays of 251 breast tumors including 30 fibroadenomas, 59 ductal carcinomas in situ (DCIS), 38 grade I invasive ductal carcinomas (IDC), 79 grade II IDC, and 45 grade III IDC. The matriptase scores were significantly higher in the tumors than their non-tumor counterparts (178+/-12 for fibroadenoma; 275+/-11 for DCIS; 299+/-10 for grade I IDC; 251+/-10 for grade II IDC; and 314+/-11 for grade III IDC). In cases of IDC, matriptase scores were significantly correlated with tumor staging and nodal staging. Our findings demonstrate that matriptase is over-expressed in breast ductal carcinoma of Chinese women. It therefore may be a good biomarker for diagnosis and treatment of malignant breast tumors.

Focal adhesion proteins as markers of malignant transformation and prognostic indicators in breast carcinoma.

Focal adhesion kinase (FAK) is one of the central signaling molecules found at focal adhesion sites, which are specific areas on the cell membrane where cells attach to extracellular matrix proteins. Focal adhesion kinase interacts with multiple signaling and adaptor molecules and effects several signaling pathways. Overexpression of FAK and its substrate c-Src has been implicated in malignant transformation and acquisition of an invasive tumor phenotype of different tissues. Overexpression of the multidomain protein paxillin, which is also a FAK ligand and a c-Src substrate, has been associated with less malignant tumor behavior. The purpose of this study was to analyze the involvement of integrin signaling molecules FAK, c-Src, and paxillin in malignant transformation of the breast epithelium. Using phosphospecific antibodies FAK-pY(397) and Src-pY(416), we demonstrated that neither activation of FAK nor activation of c-Src correlates with development of invasive tumor properties. However, activation of both FAK and c-Src correlates with malignant transformation. We further demonstrated that overexpression of paxillin also correlates with malignant transformation and is a marker of a less invasive tumor phenotype. Using tissue microarray, we demonstrated that expression and activation of paxillin inversely correlated with lymph node metastases and lymphovascular invasion, respectively. No correlation between paxillin expression and activation and tumor grade, estrogen, progesterone, and Her2/Neu receptor expression was found. In summary, focal adhesion proteins FAK and c-Src can be used as markers of malignant transformation in epithelial cells but not invasive phenotype, whereas expression and activation of paxillin may represent a good prognosticator in breast carcinoma.

Significance of inducible nitric oxide synthase expression in benign and malignant breast epithelium: an immunohistochemical study of 151 cases.

The role of calcium independent inducible nitric oxide synthase (iNOS) in breast carcinoma is controversial, and the implications of iNOS expression on prognosis are not known. In this study, we aimed to investigate the significance of immunohistochemical iNOS expression in 100 invasive ductal carcinomas. In addition, 11 normal breast tissues, 20 cases of usual ductal hyperplasias (UDHs) and 20 fibroadenomas were included. We found that 78% of malignant and 75% of benign cases showed iNOS immunoreactivity. However, the intensity and the quantity of iNOS expression were significantly higher in the cancer group when compared with benign breasts (P<0.001), suggesting a role of iNOS in breast carcinogenesis. We were unable to show a correlation between iNOS expression and tumor grade, axillary lymph node status, and estrogen receptor expression. In 50 axilla negative cases having 5--12 years follow-up, disease free survival (DFS) rate was significantly lower in cases showing strong iNOS expression (P=0.05). As strong iNOS expression was correlated with short DFS, we concluded that further studies would be necessary to elucidate if iNOS expression might be a useful prognostic marker in breast carcinoma, especially in the axilla negative group.

[Expression of estrogen sulfotransferase (EST) gene and bridging integrator protein-1 (BIN1) gene and their significance in breast tissues].

BACKGROUND & OBJECTIVE: Estrogen sulfotransferase (EST) is an enzyme which metabolizes estrogen into inactive estrogen sulphate. Bridging integrator protein-1 (BIN1) is a novel c-myc-interacting protein with the features of tumor suppressor. EST and BIN1 may be protective in tumorigenesis. This study was to detect the gene expression of EST and BIN1 in breast cancer tissues,and further investigate the carcinogenesis mechanism of breast cancer. METHODS: The mRNA levels of EST and BIN1 in 12 specimens of normal human breast tissue, and 24 specimens of breast cancer were measured by reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: EST mRNA and BIN1 mRNA were expressed in all normal human breast tissues, while EST mRNA was decreased in 75.0% (18/24), and lost in 25.0% (6/24) breast cancer specimens; BIN1 mRNA was decreased in 25.0%(16/24), and lost in 66.7.0% (16/24) breast cancer specimens. CONCLUSION: Down-regulation of EST mRNA and BIN1 mRNA may play an important role in the molecular mechanisms of breast carcinogenesis.

Augmented release of matrix metalloproteinase-9 by PKC activation in organotypic cultures of human breast cancer and adjacent normal breast tissue and fibroadenoma.

The organotypic culture technique and quantitative gelatin zymography were used to determine the expression of matrix metalloproteinase (MMP)-9 and MMP-2 in human breast cancer and adjacent normal breast tissue and fibroadenoma. MMP-9 and MMP2 were constitutively expressed in all cultures. The release of these two enzymes in breast cancer was higher than that in adjacent normal breast tissue and fibroadenoma. Administration of 12-o-tetradecanoyl- phorbol-13-acetate (TPA) increased the release of MMP-9 but not of MMP-2. This response was inhibited by protein kinase C (PKC) inhibitor (H7), transcription inhibitor (actinomycin D) and translation inhibitor (cycloheximide). Moreover, the increased level of MMP-9 by TPA in breast cancer was also higher than that in adjacent normal breast tissue and fibroadenoma. These phenomena were also observed in the DAG-treated culture. These findings suggested that the MMP-9 expression in the breast cancer tissue may be more sensitive for the PKC activation.

Expression of cyclooxygenase-2 in malignant and benign breast tumors.

Cyclooxygenase-2 (COX-2) is reported to play an important role in carcinogenesis. We examined COX-2 expression in patients with breast cancer and benign breast tumors. Immunohistochemical staining revealed a high level of COX-2 expression in malignant lesions of invasive ductal carcinoma (IDC) at a rate of 40% and a low level of expression in 15% of adjacent normal-appearing breast epithelia. Similarly, in ductal carcinoma in situ (DCIS), a high level of COX-2 expression was found in 80% of malignant lesions and a low level of expression in 50% of normal epithelia. Reverse transcriptional polymerase chain reaction (RT-PCR) performed in 7 of these cases disclosed that COX-2 expression was restricted to the malignant lesion. Further, all 10 cases of fibroadenoma and 10 cases of intraductal papilloma, both of which are benign tumors, had a high level of COX-2 expression. When overexpression of COX-2 was analyzed in relation to the clinicopathological features of the patients, no characteristic correlation was noted. Our results demonstrated that COX-2 is expressed in mammary tissue during tumorigenesis of the breast gland, suggesting that the cyclooxygenase isoenzyme may be a target for the prevention and treatment of breast cancer.

Antioxidant profile in the circulation of patients with fibroadenoma and adenocarcinoma of the breast.

OBJECTIVES: To correlate the extent of lipid peroxidation with the antioxidant status in the circulation of patients with fibroadenoma and adenocarcinoma of the breast. DESIGN AND METHODS: Ten fibroadenoma and thirty breast cancer patients and an equal number of age- and sex- matched normal subjects were chosen for the study. Lipid peroxidation as evidenced by thiobarbituric acid reactive substances (TBARS) and the status of the antioxidants superoxide dismutase (SOD), catalase (CAT), reduced glutathione (GSH), glutathione peroxidase (GPx), glutathione S-transferase (GST) ascorbic acid and vitamin E were estimated. RESULTS: Enhanced lipid peroxidation with concomitant depletion of antioxidants was observed in breast cancer patients as compared to normal subjects and fibroadenoma patients (p < 0.05). A similar pattern of changes was seen in fibroadenoma patients as compared to corresponding normal subjects (p < 0.05). CONCLUSION: This study has revealed an imbalance in the redox status in patients with fibroadenoma and adenocarcinoma of the breast.

[Significance of telomerase expression in diagnosis and treatment of breast tumor].

BACKGROUND AND OBJECTIVE: It was reported that the activation of telomerase can turn normal cell into tumor cell. This study was designed to investigate the expression of telomerase in breast tumor and its value for early diagnosis and treatment. METHODS: Telomerase expression in 178 breast tumor specimens was detected by using PCR-ELISA immunohistochemical method. RESULTS: Positive expression rate of telomerase in primary breast carcinoma, precancerous lesion of breast, breast fiber tumor were 86.72% (111/128), 46.67% (7/15), and 26.66% (4/15), respectively. Negative expression of telomerase was shown in normal breast tissue and cystic hyperplasia tissue. Positive expression rate of telomerase in different clinical stage were 63.16% in stage I, 85.96% in stage II, 95.65% in stage III, 100% in stage IV, respectively. Positive expression rate of telomerase in the patients with and without positive axillary lymph node were 95.83% (69/72) and 75(42/56), respectively. CONCLUSION: Expression of telomerase was closely associated with clinical stages of breast cancer (P < 0.005) and metastases of axillary lymph node (P < 0.01). Special importance is that its expression was found in breast precancerous lesion, indicating its is significance in early diagnosis of breast cancer.

Quantitative determination of telomerase activity in breast cancer and benign breast diseases.

Telomerase plays an important role in maintaining the stability of chromosomes. This ribonucleoprotein prevents chromosome ends (telomeres) from gradual loss with each cell division. It enables tumor cells to maintain telomere length, allowing indefinite replicative capacity. Telomerase activity has been detected in the majority of tumor and germ cells and in immortalized cell lines. Quantitative telomerase PCR-ELISA (TeloTAGGG Telomerase PCR ELISA(PLUS)) was evaluated for distinguishing benign and malignant breast tissue. Activity of telomerase was determined in 27 samples of fibrocystic and dysplastic tissues, 28 fibroadenomas and phylloid tumors, and 154 breast cancer tissues; 59 specimens were analyzed retrospectively. Analytical precision and linearity of the assay was tested using breast carcinoma cell line ZR-75-1 and breast tumor tissue extracts. About 4% of tumor samples were excluded from analysis due to interferences in the PCR reaction. Relative telomerase activity differed significantly in the groups of dysplastic tissues, fibroadenomas and carcinomas. The highest activity was found in breast cancer tissue. This method can identify breast cancer tissue with 73% clinical sensitivity and 93% specificity as compared to benign breast tumors. We did not find a correlation between telomerase activity and the tissue levels of estrogen and progesterone receptors, HER-2/neu oncoprotein concentration, tumor size, and lymph node positivity. Probability of disease-free survival was significantly lower for patients with telomerase activity higher than median value. As the assay for telomerase activity has very high analytical sensitivity and high specificity for cancer cells, this routinely used method may prove useful for distinguishing malignant phenotype of breast tissues.

Increased gelatinase-A and gelatinase-B activities in malignant vs. benign breast tumors.

Matrix metalloproteinases (MMP) types 2 and 9 (also known as gelatinase A and B) are thought to be causally involved in cancer invasion and metastasis. In normal as well as in malignant tissue, both these MMPs occur in multiple forms such as inactive precursors, active enzymes and enzyme-inhibitor complexes. Using newly developed quantitative activity assays, the levels of active MMP-2, total (active and activatable) MMP-2 and total MMP-9 were found to be significantly higher in breast carcinomas than in fibroadenomas. In addition, active MMP-2 and MMP-9 were detected more frequently in malignant than in benign breast carcinoma. These new quantitative activity assays are likely to be of use in studying the mechanism of action of both MMP-2 and -9, assessing their potential prognostic value in different cancers and in the design of MMP inhibitors for preventing cancer metastasis.

Suppression of tumor growth and metastasis in Mgat5-deficient mice.

Golgi beta1,6N-acetylglucosaminyltransferase V (MGAT5) is required in the biosynthesis of beta1,6GlcNAc-branched N-linked glycans attached to cell surface and secreted glycoproteins. Amounts of MGAT5 glycan products are commonly increased in malignancies, and correlate with disease progression. To study the functions of these N-glycans in development and disease, we generated mice deficient in Mgat5 by targeted gene mutation. These Mgat5-/- mice lacked Mgat5 products and appeared normal, but differed in their responses to certain extrinsic conditions. Mammary tumor growth and metastases induced by the polyomavirus middle T oncogene was considerably less in Mgat5-/- mice than in transgenic littermates expressing Mgat5. Furthermore, Mgat5 glycan products stimulated membrane ruffling and phosphatidylinositol 3 kinase-protein kinase B activation, fueling a positive feedback loop that amplified oncogene signaling and tumor growth in vivo. Our results indicate that inhibitors of MGAT5 might be useful in the treatment of malignancies by targeting their dependency on focal adhesion signaling for growth and metastasis.