Relationship between morphological features and kinetic patterns of enhancement of the dynamic breast magnetic resonance imaging and tumor expression of metalloproteases and their inhibitors in invasive breast cancer.

AIM: Matrix metalloproteases (MMPs) expression and their inhibitors (TIMPs) play an important role in tumor physiopathology, so we investigated the relationship between the magnetic resonance (MR) and MMPs/TIMPs expression by breast carcinomas. MATERIALS AND METHODS: MRI parameters of 64 breast carcinomas were investigated. An immunohistochemical study was also performed in these cases using tissue microarrays and specific antibodies against MMP-1, MMP-2, MMP-7, MMP-9, MMP-11, MMP-13, MMP-14, TIMP-1, TIMP-2 and TIMP-3. RESULTS: Tumors with spiculated margins had a high global (score) values of MMP-1 or MMP-7, and high expression of TIMP-3 by tumor cells. Heterogeneous tumors had a higher score values of MMP-1, MMP-13, TIMP-2 or TIMP-3, and frequent expression of TIMP-3 by tumor cells. Tumors showing fast enhancement, had higher score values of MMP-1 or MMP-11. Associations between washout curve (type III) and MMP-1, MMP-11, MMP-13 and TIMP-1 expression by tumor cells, were found. CONCLUSIONS: MRI features may predict in some grade the expressions of MMPs/TIMPs in breast tumors, which might to contribute to a better biological characterization of breast cancer.

Impact of CD68/(CD3+CD20) ratio at the invasive front of primary tumors on distant metastasis development in breast cancer.

Tumors are infiltrated by macrophages, T and B-lymphocytes, which may favor tumor development by promoting angiogenesis, growth and invasion. The aim of this study was to investigate the clinical relevance of the relative amount of macrophages (CD68⁺), T-cells (CD3⁺ and B-cells (CD20⁺) at the invasive front of breast carcinomas, and the expression of matrix metalloproteases (MMPs) and their inhibitors (TIMPs) either at the invasive front or at the tumor center. We performed an immunohistochemical study counting CD3, CD20 and CD68 positive cells at the invasive front, in 102 breast carcinomas. Also, tissue sections were stained with MMP-2, -9, -11, -14 and TIMP-2 antibodies, and immunoreactivity location, percentage of reactive area and intensity were determined at the invasive front and at the tumor center. The results showed that an increased CD68 count and CD68/(CD3+CD20) ratio were directly associated with both MMP-11 and TIMP-2 expression by mononuclear inflammatory cells at the tumor center (p = 0.041 and p = 0.025 for CD68 count and p = 0.001 and p = 0.045 for ratio, respectively for MMP-11 and TIMP-2). In addition, a high CD68/(CD3+CD20) ratio (>0.05) was directly associated with a higher probability of shortened relapse-free survival. Multivariate analysis revealed that CD68/(CD3+CD20) ratio was an independent factor associated with distant relapse-free survival (RR: 2.54, CI: (1.23-5.24), p<0.01). Therefore, CD68/(CD3+CD20) ratio at the invasive front could be used as an important prognostic marker.

Comparative analysis of the expression of metalloproteases and their inhibitors in resected crohn's disease and complicated diverticular disease.

BACKGROUND: Matrix metalloproteinases (MMPs) and their inhibitors (TIMPs), are expressed in the gastrointestinal tract by different cellular types. Nevertheless, the imbalance between MMPs and TIMPs plays an important role in the physiopathology of diverse intestinal inflammatory processes. METHODS: An immunohistochemical study was performed using tissue arrays and specific antibodies against MMPs -1, -2, -7, -9, -11, -13, -14, and TIMPs -1, -2 and -3. Immunohistochemical staining of intestinal samples from surgical interventions from 30 patients with complicated Crohn's disease (CD) and 25 patients with diverticulitis were performed at the inflamed mucosa and in adjacent noninflamed mucosa. A reverse-transcription polymerase chain reaction (RT-PCR) analysis was performed to confirm the results obtained by immunohistochemistry. In addition, western blot experiments were carried out. RESULTS: CD inflamed mucosa showed higher global expression of MMP-2, MMP-9, and MMP-13 than diverticulitis inflamed mucosa. However, inflamed and noninflamed diverticulitis mucosal samples showed higher global expression of MMP-1, TIMP-1, and 3 than the CD samples. Epithelial cells of inflamed mucosa showed higher expression of MMP-2, 9, and 13 in CD than diverticulitis. However, the latter showed higher expression of TIMP-1. Similar differences for fibroblast-like cells and mononuclear inflammatory cells were found. CD samples presented an increased expression of MMPs and a decreased expression of TIMPs compared to diverticulitis. CONCLUSIONS: These results indicate a differential pattern of expression of MMPs and TIMPs in CD and diverticulitis and the necessity to study the potential role of MMP inhibitors as new protective agents in both diseases.

Comparative study of the expression of metalloproteases and their inhibitors in different localizations within primary tumours and in metastatic lymph nodes of breast cancer.

Studies on metastasic lesions from human carcinomas are scarce. Therefore there is a need for such studies to identify the expression of the biological factors that will help in the assessment of the natural history of breast cancer. Here an immunohistochemical study was performed using tissue arrays and specific antibodies against matrix metalloproteinases (MMPs)-1, 2, 7, 9, 11, 13, 14 and tissue inhibitors of metalloproteases (TIMPs)-1, 2 and 3 in 39 patients with breast cancer. Specimens from 39 patients with node-positive carcinomas were examined and the analysis was performed at the central core of the tumour, at the invasive front, and in the metastasic axillary lymph nodes (MALNs). Global expression of MMP-1, 7 and 14, TIMP-1, and 3, were significantly higher at the centre of the tumour compared with the invasive front or the MALNs. Significantly higher expression of MMP-7 and 14, and TIMP-3, by fibroblast-like cells and mononuclear inflammatory cells (MICs) was seen in MALNs. In addition, in the tumour centre, the expression of MMP-11 and TIMP-1 and 2 by MICs, as well as TIMP-2 expression by fibroblast-like cells, were associated significantly with the occurrence of distant metastasis. In contrast, TIMP-3 expression by tumour cells or by fibroblast-like cells in this same tumour locations, as well as TIMP-1 expression by fibroblast-like cells at the invasive front, were associated significantly with poor prognosis. However, the expression of all of these biological factors in MALNs was not associated with the development of distant metastasis. Our data suggest that there is prognostic relevance to the expression of MMPs and TIMPs in the stromal cells of primary tumours, rather than to the expression of these enzymes in MALNs.

Immunohistochemical study of matrix metalloproteinases and their inhibitors in pure and mixed invasive and in situ ductal carcinomas of the breast.

We assessed differences in the patterns of expression of matrix metalloproteases and their inhibitors (tissue inhibitors of metalloproteases) in ductal carcinoma in situ alone and admixed with invasive ductal carcinomas (n = 40), as well as in pure invasive ductal carcinomas (n = 40), immunohistochemically and using tissue arrays. The invasive ductal carcinoma components showed higher expression of matrix metalloprotease-9 and -13 than did the admixed ductal carcinoma in situ, whereas stromal fibroblasts of the invasive components showed higher expression of matrix metalloprotease-2, -7, -9, -13, and -14 and tissue inhibitor of metalloprotease-1 and -3 than did fibroblasts around the neoplastic ducts of the admixed ductal carcinoma in situ. Expression of matrix metalloprotease-14 and tissue inhibitor of metalloprotease-3 was significantly higher in the mononuclear inflammatory cells of the invasive components. By contrast, matrix metalloprotease-1 expression was significantly higher in stromal cells of the ductal carcinoma in situ admixed with invasive ductal carcinoma. The pure invasive ductal carcinomas had significantly higher expression of matrix metalloprotease-1, -9, -11, and -14 and tissue inhibitor of metalloprotease-1 and -3 than the invasive ductal carcinomas admixed with ductal carcinoma in situ. Our findings indicate a significant association of matrix metalloprotease expression by the periductal stromal cells of the ductal carcinoma in situ component of mixed tumors and the occurrence of distant metastasis. Our data suggest that the molecular matrix metalloprotease/tissue inhibitor of metalloprotease profile can contribute to better characterization of early breast carcinomas.

Analysis of the expression of hyaluronan in intraductal and invasive carcinomas of the breast.

AIMS: To evaluate hyaluronan expression at different stages of tumoral progression in primary breast cancer. METHODS: Hyaluronan expression was evaluated by histochemical techniques in 42 cases of pure DCIS, in 15 cases of DCIS with a microinvasive component, and in 32 cases of invasive ductal carcinoma of the breast. Staining results were evaluated by calculating the percentage of stained areas by means of a specific software program. RESULTS: Our results show higher values of hyaluronan expression in invasive breast carcinomas [median of percentage of stained areas 41.1 (range 8-69.2)] and in DCIS with a microinvasive component [48.6 (16.8-62.8)] than in pure DCIS [14.5 (1-44.4)] (p < 0.001, for both). CONCLUSIONS: Our study indicates a proportionally higher area of hyaluronan expression in DCIS with a microinvasive component than in pure DCIS, suggesting a key role of this glycosaminoglycan in the early invasive phase of breast carcinomas. Thus, hyaluronan could play an important function in determining the migratory phenotype of cancer cells. Larger size tumors appear to demonstrate an intricate balance between hyaluronan synthesis and degradation, thus conditioning intratumoral heterogeneity in the hyaluronan metabolism.

Expression of metalloproteases and their inhibitors in primary tumors and in local recurrences after mastectomy for breast cancer.

AIMS: To investigate the expression of matrix metalloproteases (MMPs) and their inhibitors (TIMPs) in patients who develop local recurrence (LR) after mastectomy. METHODS: We analyzed the expressions of MMP-1, -2, -7, -9, -11, -13, -14, TIMP-1, -2, and -3, using immunohistochemical techniques, in primary tumors from patients without tumoral recurrence (n = 50), patients who developed distant metastasis (n = 50), and from patients who develop LRs (n = 25). LRs of the latter group were also analyzed for MMPs expression. All the patients underwent mastectomy. RESULTS: Score values for all MMPs and TIMPs were significantly higher in primary tumors of patients with distant metastasis. Primary tumors from patients with LR have lower expressions of MMPs and TIMPs compared with those from patients who developed distant metastasis, and with patients without recurrence for some MMPs. Remarkably, however, primary tumors from patients with LR showed significantly higher percentage of TIMP-1 and 2 expression in stromal cells compared to primary tumors from patients with distant metastasis or primary tumors from patients without tumoral progression. Furthermore, LRs had significantly higher MMP-9 expression than their corresponding primary tumors. CONCLUSIONS: Our data indicate differences in MMPs/TIMPs expression between primary tumors of patients with LRs and of those with distant metastasis, both after mastectomy for breast cancer.

Expression and prognostic significance of metalloproteases and their inhibitors in luminal A and basal-like phenotypes of breast carcinoma.

To analyze the expression and prognostic value of matrix metalloproteases and their tissue inhibitors in luminal A and basal-like breast carcinomas, an immunohistochemical study was performed on cancer specimens from 93 randomly selected patients with invasive primary ductal tumors of the breast (46 with and 47 without distant metastasis) and with luminal A (n = 48) (ER+, HER2-) or basal-like (HER2-, ER-, PgR-) (n = 45) lesions. Luminal B cases were too few to analyze. Specimens were also studied using tissue microarrays and specific antibodies against matrix metalloproteases 1, 2, 7, 9, 11, 13, and 14 and tissue inhibitors 1, 2, and 3. There were no significant differences in matrix metalloprotease or tissue inhibitor expression in the 2 phenotypes of tumors. In basal-like carcinomas, high scores for matrix metalloproteases 9 and 11 were significantly associated with a high distant metastasis rate. Likewise, data showed associations between matrix metalloprotease/tissue inhibitor expression by either stromal fibroblasts or mononuclear inflammatory cells and distant relapse-free survival in both tumor phenotypes. In addition, in infiltrating luminal A and basal-like tumors, we identified a prometastatic phenotype of mononuclear inflammatory cells, showing a high matrix metalloprotease/tissue inhibitor molecular profile. Expression of matrix metalloproteases and tissue inhibitors is related to the characteristics of breast tumor cells. As prognostic factors in breast carcinomas of both luminal A and basal-like phenotypes, our results point to the importance of the expression of matrix metalloproteases and tissue inhibitors by the stromal cells.

Androgen receptor expresion in breast cancer: relationship with clinicopathological characteristics of the tumors, prognosis, and expression of metalloproteases and their inhibitors.

BACKGROUND: In the present study we analyze, in patients with breast cancer, the tumor expression of androgen receptors (AR), its relationship with clinicopathological characteristics and with the expression of several matrix metalloproteases (MMPs) and their inhibitors (TIMPs), as well as with prognosis. METHODS: An immunohistochemical study was performed using tissue microarrays and specific antibodies against AR, MMPs -1, -2, -7, -9, -11, -13, -14, and TIMPs -1, -2 and -3. More than 2,800 determinations on tumor specimens from 111 patients with primary invasive ductal carcinoma of the breast (52 with axillary lymph node metastases and 59 without them) and controls were performed. Staining results were categorized using a score based on the intensity of the staining and a specific software program calculated the percentage of immunostained cells automatically. RESULTS: A total of 83 cases (74.8%) showed a positive immunostaining for AR, but with a wide variation in the staining score values. There were no significant associations between the total immunostaining scores for AR and any clinicopathological parameters. However, score values for MMP-1, -7 and -13, were significantly higher in AR-positive tumors than in AR-negative tumors. Likewise, when we considered the cellular type expressing each factor, we found that AR-positive tumors had a higher percentage of cases positive for MMP-1, -7, -11, and TIMP-2 in their malignant cells, as well as for MMP-1 in intratumoral fibroblasts. On the other hand, multivariate analysis demonstrated that patients with AR-positive tumors have a significant longer overall survival than those with AR-negative breast carcinomas (p = 0.03). CONCLUSION: Our results confirm that AR are commonly expressed in breast cancer, and are correlated with the expression of some MMPs and TIMP-2. Although we found a specific value of AR expression to be a prognostic indicator in breast cancer, the functional role of AR in these neoplasms is still unclear and further data are needed in order to clarify their biological signification in breast cancer.

The mammographic appearance of breast carcinomas of invasive ductal type: relationship with clinicopathological parameters, biological features and prognosis.

OBJECTIVES: To evaluate the clinical significance of the mammographic appearance of tumors in 411 patients with infiltrating ductal carcinoma of the breast. STUDY DESIGN: Tumors were classified into five radiographic subgroups: spiculated mass (A-type), diffuse changes with or without suspicious microcalcifications (B-type), microcalcifications with a mass (C-type), circumscribed (D-type), and not visible (E-type). Intratumoral levels of estrogen (ER) and progesterone (PR) receptors, c-erbB-2, EGFR, pS2, cathepsin D and tPA, ploidy and S-phase fraction, were analysed in a significant number of cases. RESULTS: A-type A radiographic pattern was detected in 234 patients (57%), B-type in 46 (11%), C-type in 46 (11%), D-type in 68 (17%), and E-type in 17 patients (4%). On the other hand, a total of 155 tumors (37.8%) showed microcalcifications. The percentage of tumors showing A-type pattern was more frequent in postmenopausal women, in well-differentiated tumors, and in those showing higher levels of ER, pS2 of tPA. However, B-type pattern was detected in a high percentage of premenopausal women and in those showing larger tumors, positive nodes, poor differentiation or high S-phase fraction. Cox multivariate analysis showed that B-type pattern and the absence of microcalcifications were factors significantly associated to high risk for relapse. CONCLUSIONS: Our results suggest that the mammographic appearance of tumor may to provide useful clinical information in addition to classical prognostic factor in infiltrating ductal carcinoma of the breast.

Prognostic value of pre-operative serum CA 15.3 levels in breast cancer.

BACKGROUND: CA15.3 (also known as MUCI) is the most widely used marker in breast cancer. The aim of the present work was the evaluation of the prognostic value of preoperative serum CA15.3 levels in patients with primary breast cancer. PATIENTS AND METHODS: This study included 818 women with a histologically verified diagnosis of invasive breast cancer. The serum values of CA15.3 were investigated at the time of primary diagnosis by means of an immunoradiometric assay based on the "sandwich" principle. The median follow-up period of patients free of recurrence was 38 months. RESULTS: Pre-operative CA15.3 serum levels ranged from 6 to 452 U/ml. Elevated CA15.3 levels (>30 U/ml) were found in 15.2% of patients. Statistical analysis showed that pre-operative CA15.3 serum levels were significantly higher in patients with large size tumors (T3 or T4) (p = 0.0001), as well as in those with node-positive tumors (p = 0.0001). In the univariate analysis, high CA15.3 levels were significantly associated with a lower probability of both relapse-free and overall survival in the overall group of patients (p = 0.0001 and p = 0.004, respectively) and in the subgroup with node-positive breast cancer (p = 0.001 and p = 0.03, respectively). In addition, multivariate analysis demonstrated that pre-operative levels of the antigen were significantly and independently associated with relapse-free survival in the overall group of patients, as well as in the subgroup of patients with node-positive breast cancer (p = 0.02 and p = 0.01, respectively). CONCLUSION: These results show that high pre-operative CA15.3 levels correlate with large size tumors and the presence of lymph node metastases and suggest that this antigen could be used as an additional prognostic marker.

Expression and clinical signification of cytosolic hyaluronan levels in invasive breast cancer.

BACKGROUND: Hyaluronic acid (HA), a high-molecular weight glycosaminoglycan, has been considered to be involved in the growth and progression of malignant tumors in several experimental studies. The objective of this work was to evaluate the cytosolic HA content in breast cancer, its possible relationship with clinicopathological tumor parameters and steroid receptor status, as well as its potential prognostic significance. METHODS: Cytosolic HA levels were examined by means of immunoradiometric techniques in 850 patients with invasive breast cancer. The mean follow-up period for these patients was 55.1 months. RESULTS: Cytosolic HA levels ranged widely in tumors (4-59767 ng/mg protein; median: 4960). Statistical analysis showed that HA levels were significantly higher in younger patients (p=0.0001), as well as in premenopausal than in postmenopausal patients (p=0.001). HA levels were also significantly higher in ductal or lobular histological type than in other histological types (coloid, medullar or papillar types) (p=0.0001). Likewise, HA correlated significantly and positively with tumoral levels of PgR (r sub S: 0.11; p=0.001) in the all group of patients. In the subgroup of patients with ductal invasive type, HA levels were also significantly higher in well differentiated tumors and in diploid tumors. In addition, in this latter group of patients, HA levels in tumors correlated also positively and significantly with the either estrogen-inducible proteins: PgR (r sub S: 0.11; p=0.001), pS2 (r sub S: 0.117; p=0.008) and tPA (r sub S: 0.314; p=0.0001). On the other hand, significant association between HA intratumoral levels and relapse-free survival and overall survival in the overall group of patients was not found. However, high HA intratumoral levels were significantly associated with longer relapse-free survival in the subgroup of patients with ductal histological type tumors (p=0.01), as well as in those patients without any type of systemic adjuvant treatment (p=0.01). CONCLUSIONS: Our results suggest that high intratumoral levels of HA may be associated with tumors of favorable evolution in certain subgroups of patients with breast cancer. Thus, HA may provide additional prognostic information to that given by other biochemical markers currently used in breast cancer.