Carnitine palmitoyl transferase 1A is a novel diagnostic and predictive biomarker for breast cancer.

BACKGROUND: Carnitine palmitoyl transferase 1A (CPT1A), the key regulator of fatty acid oxidation, contributes to tumor metastasis and therapeutic resistance. We aimed to identify its clinical significance as a biomarker for the diagnosis and prediction of breast cancer. METHODS: Western blot, ELISA and in silico analysis were used to confirm CPT1A levels in breast cancer cell lines, cell culture medium and breast cancer tissues. Four hundred thirty breast cancer patients, 200 patients with benign breast disease, and 400 healthy controls were enrolled and randomly divided into a training set and a test set with a 7:3 ratio. Training set was used to build diagnostic models and 10-fold cross validation was used to demonstrate the performance of the models. Then test set was aimed to validate the effectiveness of the diagnostic models. ELISA was conducted to detect individual serum CPT1A levels. Receiver operating characteristic (ROC) curves were generated, and binary logistic regression analyses were performed to evaluate the effectiveness of CPT1A as a biomarker in breast cancer diagnosis. CPT1A levels between post-operative and pre-operative samples were also compared. RESULTS: CPT1A was overexpressed in breast cancer tissues, cell lines and cell culture medium. Serum CPT1A levels were higher in breast cancer patients than in controls and were significantly associated with metastasis, TNM stage, histological grading and molecular subtype. CPT1A levels were decreased in post-operative samples compared with paired pre-operative samples. Moreover, CPT1A exhibited a higher efficacy in differentiating breast cancer patients from healthy controls (training set: area under the curve, AUC, 0.892, 95% CI, 0.872-0.920; test set, AUC, 0.904, 95% CI, 0.869-0.939) than did CA15-3, CEA, or CA125. CONCLUSION: CPT1A is overexpressed in breast cancer and can be secreted out of breast cancer cell. Serum CPT1A is positively associated with breast cancer progression and could serve as an indicator for disease monitoring. Serum CPT1A displayed a remarkably high diagnostic efficiency for breast cancer and could be a novel biomarker for the diagnosis of breast cancer.

ALK-positive Histiocytosis of the Breast: A Clinicopathologic Study Highlighting Spindle Cell Histology.

Originally described as a systemic self-limiting disease in infancy, the spectrum of ALK-positive histiocytosis has recently been broadened to include localized diseases in older children and young adults. Despite different manifestations, these tumors share histologic characteristics and a highly recurrent KIF5B-ALK fusion. ALK-positive histiocytosis is poorly characterized in the breast. In this study, we report 3 cases of ALK-positive histiocytosis of the breast. The patients were Asian women, aged 16 to 45 years. Two patients presented with an isolated breast mass, while 1 exhibited multiorgan involvement. The latter patient received ALK inhibitor after surgery, which led to complete remission. Histologically, well-circumscribed tumors displayed fascicular and storiform growth of uniform, nonatypical spindle cells admixed with lymphocytic infiltrates. Fewer conventional epithelioid histiocytes with lobulated or clefted nuclei were observed within the same breast tumors in 2 cases or within a concomitant brain tumor in the third case. Touton-type giant cells were focally present in 2 cases. Immunohistochemically, tumor spindle, and epithelioid cells were diffusely positive for CD163 and ALK in all cases and focally positive for S100 protein in 1 of the cases. CD1a and langerin were negative. Actin-positive myofibroblasts were admixed within the tumor in 2 cases, and their reactive nature was highlighted using double immunostaining. Break-apart fluorescence in situ hybridization assay demonstrated gene rearrangements involving KIF5B and ALK in all the 3 cases. ALK-positive histiocytosis rarely occurs as a spindle cell breast tumor, and should be distinguished from other diseases such as inflammatory myofibroblastic tumors and spindled histiocytic reaction.

Expression and correlation of matrix metalloproteinase-9 and heparanase in patients with breast cancer.

Matrix metalloproteinase-9 (MMP-9) and heparanase (HPSE) are thought to be involved in tumor progression and metastasis. However, up to now, there are no studies that simultaneously investigated the expression levels of MMP-9 and HPSE in tumor tissue and serum of breast cancer patients. Their correlation in breast cancer pathological processes is unknown. The purpose of this study was to investigate the expression profile of MMP-9 and HPSE in breast cancer and to assess their clinicopathological significance. We measured serum MMP-9 and HPSE by enzyme-linked immunosorbent assay in healthy women, and in patients with benign and malignant breast disease. We also evaluated the expression of MMP-9 and HPSE protein in paraffin-embedded tumor tissues by immunohistochemistry. We then correlated serum and tissue levels of MMP-9 and HPSE in breast cancer samples and their expression with patients' clinicopathologic characteristics. We found that serum levels of MMP-9 and HPSE were significantly higher in breast cancer patients than in benign breast disease and in healthy controls (P = 0.001). There was positive correlation between MMP-9 and HPSE in breast cancer patients. The tissue and serum levels of MMP-9 were associated with histology grade, lymph node status, pathological stage, and lymphovascular invasion (all P < 0.05). The tissue levels of MMP-9 were also associated with ER (P = 0.038) and Ki-67 (P = 0.032). The tissue and serum levels of HPSE expression were associated with tumor size, histology grade, lymph node status, and pathological stage (all P < 0.05). Our findings suggested that MMP-9 and HPSE might further be evaluated as biomarkers for predicting progression and prognosis of breast cancer.

An inflammatory myofibroblastic tumour of the breast with ALK overexpression.

Inflammatory myofibroblastic tumours (IMTs), also known as inflammatory pseudotumours, include a diverse group of lesions characterised by inflammatory cell infiltration and variable fibrotic responses. Their occurrence in the breast is unusual. We present a case of an IMT of the breast in a 46-year-old woman who complained of a breast mass with palpable axillary lymph node. The initial clinical diagnosis was breast cancer, and the patient underwent a conservative excision with apparently negative margins and an axillary lymph node excisional biopsy. A histopathological examination showed the presence of myofibroblastic spindle cells with mixed inflammatory infiltrates, and the pathological diagnosis was IMT. Significantly, the case we present here is unique in showing anaplastic lymphoma kinase 1 (ALK1) overexpression and ALK1 gene amplification in IMT of the breast. Therefore, our case suggests that ALK1 gene amplification in IMT of the breast has important diagnostic and therapeutic implications.

Phosphatidylinositol-3-kinase pathway mutations are common in breast columnar cell lesions.

The phosphatidylinositol-3-kinase pathway is one of the most commonly mutated pathways in invasive breast carcinoma, with PIK3CA mutations in ∼25% of invasive carcinomas, and AKT1 mutations in up to 5%. Ductal carcinoma in situ, and benign papillomas harbor similar mutations. However, activating point mutations in breast columnar cell lesions have been infrequently studied. Twenty-three breast resection specimens containing columnar cell lesions were identified; 14 with associated invasive carcinoma or carcinoma in situ. DNA extracts were prepared from formalin-fixed paraffin-embedded tissue and screened for a panel of point mutations (321 mutations in 30 genes) using a multiplex PCR panel with mass-spectroscopy readout. PIK3CA mutations were identified in 13/24 columnar cell lesions (54%) and 3/8 invasive carcinomas (37%). The mutation status of columnar cell lesions and associated carcinoma was frequently discordant. Of the 14 cases, only 5 demonstrated the same genotype in matched samples of columnar cell lesions and carcinoma (4 wild type, 1 PIK3CA H1047R). Interestingly, five patients had mutations in columnar cell lesions with wild-type carcinoma; two patients had different point mutations in columnar cell lesions and carcinoma. Only three cases had wild-type columnar cell lesion and mutated carcinoma. The 50% PIK3CA mutation prevalence in columnar cell lesions is greater than reported in most studies of invasive breast cancer. Further, columnar cell lesions and carcinoma were frequently discordant for PIK3CA/AKT1 mutation status. These findings raise interesting questions about the role of PIK3CA/AKT pathway in breast carcinogenesis, and the biologic/precursor potential of columnar cell lesions.

Immunolocalization of estrogen-producing and metabolizing enzymes in benign breast disease: comparison with normal breast and breast carcinoma.

It is well known that estrogens play important roles in the cell proliferation of breast carcinoma. Benign breast disease (BBD) contains a wide spectrum of diseases, and some are considered an important risk factor for subsequent breast carcinoma development. However, the significance of estrogens in BBD has remained largely unknown. Therefore, in this study, we examined tissue concentrations of estrogens and immunolocalization of estrogen-producing/metabolizing enzymes in BBD, and compared these findings with those in the normal breast and ductal carcinoma in situ (DCIS). Tissue concentration of estradiol in BBD (n = 9) was significantly (3.4-fold) higher than normal breast (n = 9) and nearly the same (0.7-fold) as in DCIS (n = 9). Immunoreactivity of estrogen sulfotransferase in BBD was significantly lower (n = 82) than normal breast (n = 28) but was not significantly different from DCIS (n = 28). Aromatase and steroid sulfatase immunoreactivities tended to be higher (P = 0.07) in BBD than in normal breast, and 17ß-hydroxysteroid dehydrogenase type 1 immunoreactivity was significantly higher in BBD than normal breast in the postmenopausal tissues. Immunoreactivity of estrogen and progesterone receptors was also significantly higher in BBD than normal breast. These results suggest that tissue concentration of estradiol is increased in BBD at a level similar to DCIS, which is considered mainly due to loss of estrogen sulfotransferase expression. Increased local estradiol concentration in BBD due to aberrant expression of estrogen-producing/metabolizing enzymes may play important roles in the accumulation of estradiol-mediated growth and/or subsequent development of breast carcinoma.

The role of lactate dehydrogenase, alkaline phosphatase and aspartate aminotransferase in the diagnosis of subclinical intramammary infections in dairy sheep and goats.

The objective was to investigate the changes occurring in the activities of the enzymes lactate dehydrogenase (LDH), alkaline phosphatase (ALP) and aspartate aminotransferase (AST) in sheep and goat milk as a result of subclinical intramammary infections (IMI) and to evaluate the use of these enzymes for the diagnosis of subclinical IMI in dairy sheep and goats. A total of 206 samples of sheep milk and 162 samples of goat milk, obtained from equal udder halves, were used in the study. For each species they were divided into two groups: a no-infection group and a subclinical infection group. Activities of LDH, ALP and AST were significantly higher in the subclinical infection group than in the no-infection group (P<0.05) in both sheep (LDH: 350.42+/-11.25 v. 120.91+/-4.41; ALP: 2773.43+/-105.18 v. 2189+/-94.24; AST: 29.57+/-0.74 v. 17.32+/-0.46) and goats (LDH: 354.07+/-13.33 v. 103.79+/-3.75; ALP: 311.13+/-25.74 v. 137.24+/-19.62; AST: 27.59+/-6.42 v. 15.87+/-0.45). The activity of LDH was identified as indicator for subclinical IMI in both sheep and goats. The optimum cut-off values for LDH activity, offering the highest diagnostic sensitivity (DSn) and diagnostic specificity (DSp), determined by receiver operating characteristic (ROC) analysis, were at 197 U/l, 185 U/l and 197 U/l for sheep, goats and both species, respectively. DSn for sheep, goats and both species at these cut-off values was 92.8%, 98.2% and 94.0%, whereas DSp was 95.4%, 96.3% and 96.3%, respectively. It was concluded that the determination of LDH activity in milk serum is a sensitive and reliable method for the detection of subclinical IMI in dairy sheep and goats.

Cathepsin-D expression in breast lesion: an immunohistochemical study.

Cathepsin-D (CathD) is an aspartyl lysosomal protease expressed in all tissues that might play a role in antigen processing, cell proliferation and tissue renewal, and activation of different pro hormones. The aim of our study was to compare the expression of CathD in most common breast tumors and tumor-like breast lesions. The study includes 21 patients with histologically verified breast lesions (adenosis, ductal hyperplasia, fibroadenomas, and different types of invasive carcinoma). We investigated the cathepsin-D expression in these breast lesions using immunohistochemistry (IH; paraffin-embedded tissues). Cathepsin-D staining within each lesion was assessed by estimating the area of the objects and the medium pixel intensity per object, as the integrated optical density (IOD). The immunostaining was more obvious in breast invasive carcinomas and macrophages. The reaction in tumor tissue was heterogeneous with little variation of staining intensity in positive tumor cells. Adenosis had the maximum area/signal intensity from all studied breast benign lesions (p<0.001, Student t-test). The general tendency (all benign lesions, lobular carcinomas and G3 ductal invasive carcinoma) was a more prominent representation of the cellular compartment. In the G3 ductal invasive carcinoma-type, the group of patients with metastases had a stronger expression in the cellular compartment. These results suggest that CathD expression was strongest in malignant than in benign breast disease, the positivity being present in both epithelial neoplastic and stromal cells. We also conclude that our procedure in IOD measurement is prone to less subjective-related biases, and thus more accurate and constant than other methods employed by other authors.

Prognostic significance of MMP-9 and TIMP-1 serum and tissue expression in breast cancer.

Tumor progression and metastasis contribute to the great majority of breast cancer deaths. Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) are thought to be involved in tumor progression and metastasis. Thus, we determined whether the expression of MMP-9 and TIMP-1 is associated with prognosis in breast cancer patients. We measured serum MMP-9 and TIMP-1 by enzyme-linked immunosorbent assay in 60 breast cancer patients, 18 benign breast disease patients and 15 healthy controls. We also evaluated the expression of MMP-9 and TIMP-1 protein and mRNA in paraffin-embedded tumor tissues from the 60 breast cancer patients by immunohistochemistry and in situ hybridization. We then correlated serum and tissue levels of MMP-9 and TIMP-1 in breast cancer samples and their expression with patients' clinicopathologic characteristics. We found that serum levels of MMP-9 and TIMP-1 were significantly higher in breast cancer patients than in benign breast disease and in healthy controls. High serum levels of MMP-9 and TIMP-1 were associated with lymph node metastasis, higher tumor stage and lower relapse-free and overall survival (OS) rates. Compared to low expression, high tissue expression of MMP-9 protein was associated with lymph node metastasis and higher tumor stage; and high tissue expression of TIMP-1 was associated with a lower OS rate. Our findings suggest that MMP-9 and TIMP-1 may further be evaluated as biomarkers for predicting progression and prognosis of breast cancer.

Expression of cytochrome P450 1B1 and catechol-O-methyltransferase in breast tissue and their associations with breast cancer risk.

Cytochrome P450 1B1 (CYP1B1) and catechol-O-methyltransferase (COMT) are important estrogen-metabolizing enzymes that may affect breast cancer risk. Few studies have directly measured the expression of CYP1B1 and COMT genes in breast tissue samples. The subjects in this study were a subgroup of participants of the Shanghai Breast Cancer Study including 64 patients diagnosed with breast cancer and 68 patients diagnosed with benign breast diseases (BBD) who provided samples of tumor tissue and adjacent nontumor tissue to the study. We compared CYP1B1 and COMT mRNA expression in tumor tissue and adjacent nontumor tissue in both breast cancer patients and BBD patients. High levels of CYP1B1 expression and low levels of COMT expression in adjacent nontumor tissue were associated with a significantly increased breast cancer risk in a nonlinear manner. Odds ratios and 95% confidence intervals (in parentheses) for the midpoints of the first, second, fourth, and fifth quintiles of gene expression levels compared with the overall median levels in BBD subjects were 0.21 (0.07-0.67), 0.81 (0.69-0.95), 1.20 (1.05-1.38), and 1.55 (1.12-2.15) for CYP1B1 and 1.72 (1.17-2.55), 1.19 (1.05-1.35), 0.83 (0.73-0.95), and 0.78 (0.65-0.93) for COMT, respectively. These results support the hypothesis that the formation and accumulation of catechol estrogens in breast tissue through increased CYP1B1 expression and reduced COMT expression may play a significant role in breast cancer risk.

Circulating MMP2 and MMP9 in breast cancer -- potential role in classification of patients into low risk, high risk, benign disease and breast cancer categories.

Matrix metalloproteinase (MMP) 2 and 9 are involved in cancer invasion and metastasis, and increased levels occur in serum and plasma of breast cancer (BC) patients. It is, however, unclear whether changes in serum levels can be exploited for early detection or classification of patients into different risk/disease categories. In our study, we measured concentration and activity of MMP2/9 in sera of 345 donors classified as low risk (Gail score <1.7), high risk (HR) (Gail score > or =1.7), benign disease or BC. Kruskal-Wallis and Mann-Whitney nonparametric tests showed that total-MMP2 concentration is higher in HR compared to control (p = 0.012), benign (p = 0.001) and cancer (p = 0.007). Active MMP2 (aMMP2) concentration is higher in control than benign and cancer (p < 0.001, respectively). Total and aMMP9 concentrations are higher in cancer than benign (p < 0.001, p = 0.002, respectively). Total-MMP2 and total-MMP9 activities are lower in control than benign (p < 0.001, p = 0.002, respectively) and cancer (p < 0.001, respectively). Total-MMP2 and MMP9 activities are also higher in cancer than benign (p = 0.004, p < 0.001) and HR (p = 0.008, p = 0.007, respectively). These results were not affected by age or inclusion/exclusion of donors with noninvasive cancer or atypical hyperplasia. Linear discriminant analysis revealed that HR donors are characterized by lower total-MMP2 and higher aMMP2. Overall group classification accuracy was 64.5%. Independent validation based on the leave-one-out cross validation approach gave an overall classification of 63%. Our study provides evidence supporting the potential role of serum MMP2/9 as biomarkers for breast disease classification.

Plasma concentration and activity of matrix metalloproteinase 2 and 9 in patients with breast disease, breast cancer and at risk of developing breast cancer.

Matrix metalloproteinases (MMPs) are involved in extracellular matrix modification and associated with invasive and metastatic behavior of human malignant tumors. Specifically, MMP2 and MMP9 are implicated in both early and late processes of tumor development. It is reported that MMPs occur as inactive precursors, active enzymes or enzyme inhibitor complexes in biological samples. However, there is limited knowledge on the role of each form in disease and/or the significance of changes in the plasma concentration and/or activity in breast cancer patients. The aim of this study was to determine if patients with breast cancer, benign disease and at risk for developing breast cancer display characteristic levels of active and/or total MMP2 and MMP9 in plasma. Concentration and activity of MMP2 and MMP9 were determined quantitatively in the plasma of 124 female volunteers diagnosed with breast cancer (n=31), benign disease (n=38), or determined by the Gail Model to be at high risk (n=31) or low risk (controls, n=24) of developing breast cancer. Data obtained was statistically analyzed to search for differences/patterns characteristic of each category. Concentration of total MMP2 was significantly lower in control individuals than benign, high risk (P<0.001 respectively) and breast cancer patients (P=0.002). Activity of total MMP2 was significantly lower in controls compared to cancer, benign and high risk patients (P<0.001 respectively). Attempts to build a predictive/descriptive model using canonical discriminant analysis (utilizing all eight features; concentrations and activity levels of active/total MMP2 and MMP9) enabled the distinction of the controls from the high risk, benign and cancer groups. Our results suggest that preoperative plasma concentration and activity of MMP2 and MMP9 may permit sub-classification of female patients with breast disorders.

UDP-N-acetyl-D-galactosamine: polypeptide N-acetylgalactosaminyltransferase-6 as a new immunohistochemical breast cancer marker.

Mucin O-glycosylation is characterized in cancer by aberrant expression of immature carbohydrate structures (Tn, T, and sialyl-Tn antigens). The UDP-N-acetyl-D-galactosamine: polypeptide N-acetylgalactosaminyltransferases (ppGalNAc-T) family enzymes regulate the initial steps of mucin O-glycosylation and could be responsible for the altered glycosylation observed in cancer. Considering that we recently found the ppGalNAc-T6 mRNA expressed in breast carcinomas, we produced a highly specific monoclonal antibody (MAb T6.3) to assess the expression profile of ppGalNAc-T6 protein product in breast tissues. The expression of ppGalNAc-T6 by breast carcinoma cells was confirmed on MCF-7 and T47D cell lines. In formalin-fixed tissues, ppGalNAc-T6 expression was observed in 60/74 (81%) breast cancers, 21/23 (91.3%) adjacent ductal carcinoma in situ (DCIS), 4/20 benign breast lesions (2/2 sclerosing adenosis and 2/13 fibroadenoma), and in 0/5 normal breast samples. We observed a statistically significant association of ppGalNAc-T6 expression with T1 tumor stage. This fact, as well as the observation that ppGalNAc-T6 was strongly expressed in sclerosing adenosis and in most DCIS, suggests that ppGalNAc-T6 expression could be an early event during human breast carcinogenesis. Considering that an abnormal O-glycosylation greatly contributes to the phenotype and biology of breast cancer cells, ppGalNAc-T6 expression could provide new insights about breast cancer glycobiology.

Upregulation of focal adhesion kinase (FAK) expression in ductal carcinoma in situ (DCIS) is an early event in breast tumorigenesis.

Focal adhesion kinase (FAK) is a protein tyrosine kinase that is overexpressed in a subset of invasive breast cancers. FAK transmits signals that mediate several functions including tumor cell proliferation, migration, adhesion and survival. We used immunohistochemical techniques to assess FAK expression in patients with fibrocystic disease (FCD), atypical ductal hyperplasia (ADH), ductal carcinoma in situ (DCIS) and infiltrating ductal carcinoma (IDC). Formalin-fixed, paraffin-embedded (FFPE) tissue sections were obtained from 119 patients (12 FCD, 38 ADH, 51 DCIS and 18 IDC). The anti-FAK 4.47 monoclonal antibody was used to detect FAK expression. FAK expression was scored as high (3 or 4 intensity and > or =90% positive cells) or low. The DCIS tissue sections demonstrated high FAK expression in 34/51 (66%) of the sections. High FAK expression was demonstrated in 6/18 (33%) of the IDC tissue sections and 8/38 (21%) of the ADH tissue sections. None (0/12) of the FCD tissues sections stained high for FAK. The pattern of FAK expression in DCIS was significantly higher than ADH (p < 0.0001) and IDC (p = 0.02). We conclude that FAK overexpression in preinvasive, DCIS tumors precedes tumor cell invasion or metastasis, suggesting that FAK may function as a survival signal and be an early event in breast tumorigenesis.

[Quantitative detection of telomerase activity and its association with clinicopathological characteristics in breast cancer].

BACKGROUND & OBJECTIVE: Telomerase is a ribonucleoprotein enzyme, which plays an important role in cell immortalization and carcinogenesis. Recent studies on the association of telomerase activity with prognostic factors of breast cancer were controversial due to different methods. This study was to establish a feasible assay of quantitative detection of telomerase activity based on silver staining, and investigate possible association between telomerase activity and clinicopathological prognostic factors in primary breast cancer. METHODS: Highly sensitive silver-staining telomeric repeat amplification protocol assay (SS-TRAP) was used to quantify telomerase activity in 52 frozen human breast cancer samples and their adjacent breast tissues, 32 benign lesions, and 14 normal mammary gland lesions. The association between telomerase activity and clinicopathological data was analyzed. RESULTS: Telomerase activity was detected in 47 of the 52 breast cancer samples (90.38%), and in 10 of the 32 benign lesions, whereas no activity was detected in 37 of 52 adjacent nonmalignant breast tissues, and all 14 normal mammary gland tissues. The telomerase activity levels were 36.91+/-15.35, 8.27+/-4.37, 14.10+/-5.28, 0 (unit: TPG) in breast cancer, adjacent tissue of cancer, benign lesion, and normal tissue, respectively. The difference of telomerase activity was significant between breast cancer and the other 3 groups by using ANVOA (all P< 0.01). A strong correlation was found between telomerase activity and pathological category, and differentiation degree by logistic regression analysis, i.e. with ongoing tumor progression, telomerase activity appeared to increase in primary breast cancer (P=0.003, and P=0.004). No correlation was seen between telomerase activity and disease course, age, and menopause status of patients (all P >0.05). Telomerase activity level of invasive non-special cancer was higher than that of invasive special cancer(P< 0.05). Telomerase activity level of moderately/poorly differentiated carcer was higher than that of highly differentiated cancer (P< 0.05), while no obvious difference was found between moderately and poorly differentiated cancer (P >0.05). CONCLUSIONS: The activation of telomerase might occur early in breast cancer,and plays a critical role in carcinogenesis and tumor development. Telomerase may serve as a specific marker of early diagnosis and prognosis in mammary gland neoplasm.

Assessment of apoptosis in human breast tissue using an antibody against the active form of caspase 3: relation to tumour histopathological characteristics.

Apoptosis is of important significance in the pathogenesis of cancer. Many methods are available for the measurement of apoptosis but the 'gold standard' is to identify apoptotic cells by their morphological features using microscopy. Caspase 3 is a cytosolic enzyme that is activated only in cells committed to undergo apoptosis. The activation of caspase 3 precedes the development of the classical morphological features of apoptosis. Using immunohistochemistry with an antibody against the active form of caspase 3, the apoptotic index (AI) was measured in 116 samples of human breast tissue (22 normal/benign and 94 invasive carcinomas). The AI obtained by measuring caspase activation has a strong correlation with the AI derived by morphological assessment (r = 0.736, P < 0.01). The AI is higher in the invasive group than in the benign group (P = 0.008), and in invasive cancer high AI is associated with high tumour grade (P = 0.013), positive node status (P < 0.001) and negative steroid receptor status (P = 0.001 for ER; P = 0.004 for PR). No significant association is observed between AI and tumour size. Measurement of apoptosis by immunohistochemistry using an antibody against the active form of caspase 3 is therefore reliable and correlates strongly with morphological assessment.

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.

[Metabolism of adenosine and thymidine in healthy females of different ages and females with mastopathies]. / Metabolizm adenozynu ta tymidynu v orhanizmi zdorovykh ta khvoryk na mastopatiiu zhinok riznoho viku.

The activity of thymidine kinase, thymidine phosphorylase, adenosine deaminase and 5'-nucleotidase of AMP was studied in tissues of 39 healthy females, as well as blood serum and lymphocytes of 60 healthy females, as well as in 50 patients with fibrocavernous mastopathy aged as 23-70. Comparative determination of adenosine metabolism enzymes activity in lymphocytes was carried out simultaneously with studying some immunological indexes in the organism of the same-aged healthy females and ones with mastopathy. It was revealed that age-related changes in the activity of thymidine kinase in blood serum reflected the analogous changes in enzyme activity in tissues of the healthy women. A direct correlation was established between thymidine kinase activity and age both in the healthy females and those with mastopathy. A significant decrease in activity of thymidine phosphorylase was demonstrated in blood serum of the patients with mastopathy in the age 46-60. Determined 4-fold increase in the activity of adenosine deaminase in serum was accompanied by decreased enzyme activity in lymphocytes and decreased Lymphocyte Blast Transformation Index in the same age range. Changes of immunological status are more expressed in T-system of immunity. The revealed metabolic changes in DNA-precursors metabolism in the patients with mastopathy aged as 46-60 might be one of the reasons of increased risk of oncological disease in this age group.

Copper, zinc and superoxide dismutase in precancerous, benign diseases and gastric, colorectal and breast cancer.

The aim of the present study was to assess serum levels of copper and zinc levels and erythrocytes Cu,Zn-SOD activity and to determine probable changes in gastric and colorectal precancerous diseases, benign breast diseases, gastric, colorectal and breast cancer. The study included 165 subjects with cancer, 348 subjects with precancerous (atrophic gastritis, gastric adenoma, colon adenoma, rectal adenoma) and/or benign diseases (weak dysplasia, severe dysplasia, fibroadenoma, cystic disease) and 161 randomly selected healthy controls. Our results suggest that while in gastric and colorectal cancer there were mostly increased copper levels, in breast cancer they were not changed. Zinc levels were weakly decreased in atrophic gastritis, gastric adenoma and breast cancer. There was a strong positive correlation between zinc levels and SOD activity in fibroadenoma and a weak positive correlation in colorectal adenoma and colorectal cancer without any correlation between SOD activity and copper in these groups. In gastric precancerous disease there was a positive correlation between SOD and copper. The results of this study suggest that serum trace element levels and activity of related enzymes might be different in various neoplastic processes. This variation in neoplastic processes might be influenced by other factors that have to be considered in complex relationships between the whole body and neoplastic cells.