Prognostic and predictive value of the urokinase-type plasminogen activator (uPA) and its inhibitors PAI-1 and PAI-2 in operable breast cancer

The present study on the prognostic and predictive value of serine proteases was conducted in 460 early breast cancer patients mostly treated with some kind of adjuvant systemic therapy: 156 received chemotherapy, 141 hormone therapy and 111 a combination of both. Already in univariate analysis PAI-1 was the only proteolytic factor with a significant impact on DFS, which was retained in multivariate analysis (p = 0.020); PAI-2 showed borderline significance in univariate analysis (p = 0.0503) and uPA did not present as a significant prognostic factor for DFS in our patient series. In a separate univariate analysis of DFS on patient subgroups defined by adjuvant systemic therapy, a higher risk of relapse associated with higher uPA and PAI-1 levels was found in the subgroup of patients who did not receive any treatment; this difference did not reach the level of significance, probably due to the small number (n = 52) of patients in this group (HR 1.37; p = 0.71 and HR 2.14; p = 0.321, respectively). A higher risk of relapse was also found in the subgroup of patients treated with adjuvant chemotherapy (HR 1.44; p = 0.381 and HR 2.48; p = 0.003, respectively). In contrast, the bad prognostic impact of high uPA and PAI-1 levels was lost in the subgroup of patients treated with adjuvant hormone therapy (HR 0.79; p = 0.693 and HR 0.26; p = 0.204, respectively). The same observations were made for the uPA/PAI-1 combination. Our study confirmed the prognostic value of serine proteases in early breast cancer. In addition, it pointed to a possible predictive value of these tumor markers for response to adjuvant hormone therapy with tamoxifen, which should be confirmed in further studies.     

Prognostic factors for recurrence and survival in axillary node-negative breast cancer

The question of which node-negative breast cancer patients should be treated with adjuvant systemic therapy is a debatable topic. Our approach in San Antonio is to examine the risk profile for an individual patient and attempt to classify the patient into a good risk group or a high risk group in terms of disease recurrence. Features such as small tumor size (less than 2 cm), diploid tumors with low proliferative rate, and nuclear grade I, all indicate a good prognosis with a disease-free survival of approx. 90% at 5 yr. Examination of the cost vs benefits in this category of patients suggest that routine treatment with systemic adjuvant therapy is not appropriate.     

Limited prognostic value of c-erbB-2 compared to uPA and PAI-1 in primary breast carcinoma

In a retrospective study of 488 women with primary breast cancer, after a median follow-up of 10 years, we sought interactions between disease-free survival (DFS) and overall survival (OS) and tumor antigen levels of two components of the plasminogen system, urokinase-type plasminogen activator (uPA) and its inhibitor PAI-1, and the transmembrane growth factor receptor c-erbB-2. We used ELISAs (American Diagnostica, Greenwich, CT, USA) to quantify uPA and PAI-1 antigen levels in cytosols, and a double monoclonal antibody-based assay (EIA) (Ciba Corning Diagnostics, Alameda, CA, USA) to quantify c-erbB-2 in membrane extracts of the same tissues. Weak positive correlations were found between uPA and c-erbB-2 (r(s) = 0.146; p = 0.001) and between PAI-1 and c-erbB-2 (r(s) = 0.154; p < 0.001). In the overall population, using univariate analyses, c-erbB-2 overexpression and high uPA and PAI-1 antigen levels (> 300 IU/mg, > 1.40 ng/mg and > 5.53 ng/mg, respectively) were significantly associated with shorter DFS (p = 0.003, p < 0.001 and p < 0.001, respectively) and OS (p < 0.001 in all cases). Using multivariate analyses, PAI-1, node status and tumor size were independent predictors of DFS and c-erbB-2 was retained in the model only for OS. In the node-negative subgroup, PAI-1 was the strongest significant survival predictor both for OS (p = 0.003; HR 2.52) and DFS (p < 0.001; HR 2.39). This study shows that in primary breast cancer c-erbB-2 offers no additional prognostic information when uPA and/or PAI-1 are candidates in the multivariate analyses.     

Angiogenesis as an independent prognostic indicator in node-negative breast cancer

OBJECTIVE: To quantitate tumor angiogenesis in carcinoma of the breast (stage T2N0M0) by computerized image analysis of CD-31-stained histologic sections and, keeping in view the heterogeneity of tumors, to assess which areas of neovascularization provide the best prognostic indicator. STUDY DESIGN: Thirty-six cases of infiltrating duct carcinoma of the breast, stage T2N0M0, with follow-up of five years, were analyzed. All cases had received uniform initial treatment in the form of mastectomy with axillary clearance and radiotherapy. Intratumoral microvessel density (IMD) counts were done on "hot spots" and "non-hot spots" on CD-31-stained sections using computerized image analysis. Angiogenesis was correlated with other variables, such as age, menopausal status, histologic grade and proliferative activity by univariate and multivariate analyses. RESULTS: Hot-spot IMD counts were highly significant independent prognostic markers in univariate and multivariate analyses. Background vascularity of a tumor was of no value in prognosticating. CONCLUSION: In patients with node negative breast carcinoma, IMD counts in hot spots are an independent prognostic factor in disease-free and overall survival and can be used to separate out patients with T2N0M0 stage in need of systemic adjuvant therapy.     

A structural basis for differential cell signalling by PAI-1 and PAI-2 in breast cancer cells

PAI-1 and PAI-2 (plasminogen-activator inibitor types 1 and 2) are inhibitors of cell surface uPA (urokinase plasminogen activator). However, tumour expression of PAI-1 and PAI-2 correlates with poor compared with good patient prognosis in breast cancer respectively. This biological divergence may be related to additional functional roles of PAI-1. For example, the inhibition of uPA by PAI-1 reveals a cryptic high-affinity site within the PAI-1 moiety for the VLDLr (very-low-density-lipoprotein receptor), which sustains cell signalling events initiated by binding of uPA to its receptor. These interactions and subsequent signalling events promote proliferation of breast cancer cells. Biochemical and structural analyses show that, unlike PAI-1, the PAI-2 moiety of uPA-PAI-2 does not contain a high-affinity-binding site for VLDLr, although uPA-PAI-2 is still efficiently endocytosed via this receptor in breast cancer cells. Furthermore, global protein tyrosine phosphorylation events were not sustained by uPA-PAI-2 and cell proliferation was not affected. We thus propose a structurally based mechanism for these differences between PAI-1 and PAI-2 and suggest that PAI-2 is able to inhibit and clear uPA activity without initiating mitogenic signalling events through VLDLr.     

tPA, uPA, PAI-1和 PAI-2在人和恒河猴胎盘中的原位杂交和荧光免疫定位(英文)

用原位杂交和荧光免疫定位方法研究了组织型（ｔ）和尿激酶型（ｕ）纤溶酶原激活因子ｔＰＡ、ｕＰＡ和相应的抑制因子ＰＡＩ－１、ＰＡＩ－２在人和恒河猴胎盘中的定位和分布。结果表明：（１）激活因子ｔＰＡ、ｕＰＡ（Ｆｉｇ．１＆４）和抑制因子ＰＡＩ－１（Ｆｉｇ．２）、ＰＡＩ－２（Ｆｉｇ．３）一般都在不同程度上定位于两者胎盘的相同部位;（２）它们主要分布在绒毛干和蜕膜的血管壁、 ＲＯｈｒ’ｓ和Ｎｉｔａｂｕｃｈ’ｓ纹间的基盘外绒毛滋养层细胞、滋养壳、蜕膜细胞和腺体细胞。并且发现;ｔＰＡ和它的抑制因子ＰＡＩ－１更明显地定位于邻近母体组织离层界面的区域,而ｕＰＡ和抑制因子ＰＡＩ－１更集中在绒毛滋养层和外绒毛滋养细胞中;（３）激活因子和抑制因子的ｍＲＮＡ和蛋白的定位和分布基本上一致,但是、在绒毛核体滋养层细胞上未发现其ｍＲＮＡ表达,却有很强的免疫荧光的分布;（４）激活因子和抑制因子合成和分布部位与其作用底物,即纤蛋白类分子的产生部位一致;（５）未发现上述分子在人和恒河猴胎盘分布上的不同。上述实验结果说明,在妊娠的各个阶段 ＰＡ和它的抑制因子协同表达,局限在作用范围很小的特定产生底物的区域。它们的相互作用可能是维持正常妊娠所必需。在妊     

Evolution of adjuvant chemotherapy of breast cancer from Bonadonna to the taxanes

The author summarizes the progress of adjuvant chemotherapy of breast cancer from the classical Bonadonna-type CMF over the anthracyclines to the taxanes. The CMF regimen represented the prototype of combination chemotherapy which significantly improved early and long term results. After 20 years the patients given adjuvant combination chemotherapy with CMF had significantly better rates of relapse-free survival (p<0.001) and overall survival (p=0.03) compared with no chemotherapy. 6 cycles of CMF was the gold standard of adjuvant chemotherapy in breast cancer for decades. The Milan research group decided in the early 1980s to challange this popular CMF combination by introducing doxorubicin within the adjuvant program. Compared with standard CMF, anthracyclin-containing regimens reduced the annual risk of recurrence by 12% and the annual risk of death by 11%, equating to a 3.2% absolute reduction in recurrence and a 2.7% absolute reduction in mortality at 5 years. This small but real difference seen with regimens containing three or more agents (e.g. CEF and CAF, FAC, FEC, etc.), whereas 4 cycles of 2-drug regimens (e.g. AC or EC) appears to be equivalent to 6 cycles of CMF. Among the novel chemotherapeutic drugs introduced in the 1990s the taxanes have emerged as the most powerful compounds in breast cancer. Several large, adjuvant clinical trials are currently ongoing or have recently completed accrual. The available results from innumerable clinical studies are still inconclusive and do not support the routine use of taxanes in the adjuvant setting - with the exception of the BCIRG 001 docetaxel trial, in which significant improvement was documented in disease free survival with 6 x TAC compared with 6 x FAC (82% vs 74%). Studies on the effect of the new trastuzumab (an antibody against the extracellular domain of the HER2) in adjuvant setting was initiated in early 2000. The Herceptin adjuvant trial programme is extensive, involving more than 12,000 patients worldwide. This trials will potentially offer many women with HER2-positive disease the chance of improved survival.     

Histopathological diagnosis of thyroid cancer in a multicenter trial

INTRODUCTION: In the front of the problems related to the differentiation between benign and malignant thyroid tumors we decided to perform a multicentre study in order to validate diagnoses of malignant thyroid tumors and assess the inter-observer variability. MATERIAL AND METHODS: Material included 690 cases of malignant and benign thyroid lesions with primary histopathology established in 1985-1999. These cases were selected to multicentre study. The studies were sent from centres which agreed to participate in the project and than coded in the independent centre--Department of Nuclear Medicine and Endocrine Oncology. 40 pathologists from 25 centres provided their diagnoses which were compared with the reference ones. RESULTS: 10 547 diagnoses were evaluated, both on their accuracy of the distinction between malignant and benign lesions and on their accuracy of cancer histotype definition. The reference diagnosis was made by an agreement between four expert pathologists (D.L., S.S., J.S. and A.K.). The participants diagnosed 21% of cases differently than experts. Concerning the diagnosis of cancer histotype, the difference between participants diagnosis and the reference one was even higher. The best concordance was achieved in the diagnosis of papillary thyroid cancer, however, on the cost of cancer overdiagnosis by some participants. Follicular cancer was diagnosed accurately only in 75.4% of cases. CONCLUSION: The study documents a high inter-observer variability of thyroid cancer diagnosis and confirms the lesser accuracy of diagnosis of follicular cancer.     

Aromatase inhibitors as adjuvant therapies in patients with breast cancer

There is increasing evidence that endocrine therapy has an important role in patients with oestrogen receptor positive breast cancer. Several large meta-analyses have reinforced the value of both ovarian ablation and tamoxifen in improving survival. Over the past decade, aromatase inhibitors have become the treatment of choice for second-line therapy of metastatic breast cancer, and the third generation inhibitors have now an established reputation for good patient tolerability. Early studies indicated that aminoglutethimide/hydrocortisone could benefit postmenopausal patients with primary breast cancer, and in 2001, the ATAC study showed that the third generation aromatase inhibitor, anastrozole, seemed superior to tamoxifen in that anastrozole-treated patients had a longer disease-free survival. Other studies will report on the relative merits of the steroidal inhibitor exemestane as well as non-steroidal letrozole. The exact duration and sequencing of treatment, together with the long-term effects on bone are at present, unknown.     

Prognostic significance of GRP78 expression patterns in breast cancer patients receiving adjuvant chemotherapy

This study examined the associations between GRP78 expression and breast cancer recurrence and survival in patients treated with anthracyclines in the adjuvant setting. GRP78 expression was assessed in 106 stage II/III breast cancer patients. Tissue microarray was used to perform immunohistochemistry and to determine the GRP78 expression in endoplasmic reticulum and cell membrane of breast tumors. Four distinct scenarios (low and high thresholds) were developed. For high thresholds, 16% and 40% of our cases were GRP78-positive for endoplasmic reticulum and cell membrane, respectively. For low thresholds, 74% and 87% of our cases were GRP78-positive for endoplasmic reticulum and cell membrane, respectively. In the endoplasmic reticulum high-threshold scenario, GRP78 positive was found to be significantly frequent in T3 tumors (p=0.02), and inversely related to ERBB2 overexpression (p=0.03). There was a lower proportion of GRP78-positive cases among women between 50 and 65 years of age (p=0.02). In the endoplasmic reticulum low-threshold scenario, the proportion of GRP78-positive cases was significantly higher in women younger than 50 years and in those who were premenopausal (p=0.04). No statistically significant difference was found in survival probabilities among the scenarios examined. In our cohort, GRP78 overexpression was not a predictor of overall or disease-free survival of patients receiving anthracycline-based adjuvant chemotherapy.     

Surgical perspectives from a prospective, nonrandomized, multicenter study of breast conserving surgery and adjuvant electronic brachytherapy for the treatment of breast cancer

Background

The influence of viral hepatitis status on prognosis in patients undergoing hepatic resection for hepatocellular carcinoma (HCC) remains a matter of debate. This study is a meta-analysis of the available evidence.

Methods

A literature search was performed to identify comparative studies reporting postoperative survival of HCC in different types of viral hepatitis. Pooled odds ratios (OR) and weighted mean differences (WMD with 95% confidence intervals (95% CI) were calculated using either the fixed effects model or random effects model.

Results

Twenty studies matched the selection criteria and reported on 4744 subjects, of whom 2008 in the HBV-positive (B-HCC) group, 2222 in the HCV-positive (C-HCC) group, and 514 in the hepatitis B- and C-negative (NBNC-HCC). Meta-analysis showed that patients with HBV or HCV infection had a worse 5-year disease-free survival when compared to patients with NBNC-HCC (respectively: OR: 0.39, 95% CI: 0.28 to 0.53, P < 0.001; WMD: 0.37, 95% CI: 0.22 to 0.64, P < 0.001). There was a tendency toward higher 5-year overall survival rates in the NBNC-HCC group compared to those in the other two groups, although these differences were not statistically significant. Both the 5-year overall survival and disease-free survival were not different among the B-HCC and C-HCC groups.

Conclusions

Patients with positive serology for hepatitis B or C undergoing resection for HCC had a poor prognosis compared to patients with negative serology.     

CD44/CD24 as potential prognostic markers in node-positive invasive ductal breast cancer patients treated with adjuvant chemotherapy

The hypothesis on cancer stem cells assumes the existence of small subpopulation of cells that possess the ability to undergo self-renewal and can give rise to the diversity of differentiated cells that form the tumour. It has been accepted that CD44⁺/CD24^?/low phenotype is one of the features characterizing breast cancer stem cells. The aim of our study was to assess (1) prognostic significance of CD44/CD24 expression as well as (2) a relation between the above-mentioned phenotype and breast cancer subtypes [based on estrogen (ER), progesterone receptors, human epidermal growth factor receptor 2 and Ki67 status] and expression of selected markers such as fascin, laminin-5 gamma-2 chain, cytokeratin (CK) 5/6 and 8/18, epidermal growth factor receptor (EGFR), smooth muscle actin, P-cadherin and lymphocytic infiltration in invasive ductal breast cancer patients (T ≥ 1, N ≥ 1, M0), who underwent mastectomy followed by chemotherapy (with taxanes and/or anthracyclines) or/and hormonotherapy. We noted that most cancers with CD44?/CD24? and CD44?/CD24+ phenotype were ER positive. The majority of CD44?/CD24?, CD44?/CD24+ and CD44+/CD24? tumours were characterized by CK5/6 and EGFR negativity. In univariate analysis we demonstrated that patients with pN1/pN2 and with CD44 +/CD24- carcinomas had significantly lower risk of progression or cancer-related death than those with pN3 or tumours characterised by other CD44/CD24 expression patterns. We also found 100 % DFS in 12 patients with CD44+/CD24?/CK5/6+/ER? phenotype. Other analysed parameters were insignificant. We conclude that tumours with immunophenotypes: CD44+/CD24? and CD44+/CD24?/CK5/6+/ER? might be more sensitive for chemotherapy based on taxanes and/or anthracyclines.     

Effect on natural killer and antibody-dependent cellular cytotoxicity of adjuvant cytotoxic chemotherapy including melphalan in breast cancer

Natural killer (NK) cell activity and antibody-dependent (K) cell activity were studied sequentially in 30 patients with early node-positive breast cancer entered into an adjuvant chemotherapy trial. The drugs used were melphalan, and melphalan with methotrexate, given for 12 months. Estimations were made 3-monthly during chemotherapy, and then at 15 and 24 months to assess recovery. Mean values for NK-cell activity during chemotherapy were significantly lower than the mean pre-chemotherapy baseline value at all time-points from 3 to 15 months, but there was recovery by 24 months. Mean values for K-cell activity during chemotherapy did not appear to differ from the mean pre-chemotherapy value, but variability in individual values was high. Over a 4-year follow-up period, a comparison of 16 patients who did not develop recurrent breast cancer with 14 who did showed that NK-cell activity was significantly lower in the latter group 12 months after the start of chemotherapy.     

Combined evaluation of LC3B puncta and HMGB1 expression predicts residual risk of relapse after adjuvant chemotherapy in breast cancer

In spite of adjuvant chemotherapy, a significant fraction of patients with localized breast cancer (BC) relapse after optimal treatment. We determined the occurrence of cytoplasmic MAP1LC3B/LC3B (microtubule-associated protein 1 light chain 3B)-positive puncta, as well as the presence of nuclear HMGB1 (high mobility group box 1) in cancer cells within surgical BC specimens by immunohistochemistry, first in a test cohort (152 patients) and then in a validation cohort of localized BC patients who all received adjuvant anthracycline-based chemotherapy (1646 patients). Cytoplasmic LC3B⁺ puncta inversely correlated with the intensity of SQSTM1 staining, suggesting that a high percentage cells of LC3B⁺ puncta reflects increased autophagic flux. After setting optimal thresholds in the test cohort, cytoplasmic LC3B⁺ puncta and nuclear HMGB1 were scored as positive in 27.2% and 28.6% of the tumors, respectively, in the validation cohort, while 8.7% were considered as double positive. LC3B⁺ puncta or HMGB1 expression alone did not constitute independent prognostic factors for metastasis-free survival (MFS) in multivariate analyses. However, the combined positivity for LC3B⁺ puncta and nuclear HMGB1 constituted an independent prognostic factor significantly associated with prolonged MFS (hazard ratio: 0.49 95% confidence interval [0.26–0.89]; P = 0.02), and improved breast cancer specific survival (hazard ratio: 0.21 95% confidence interval [0.05–0.85]; P = 0.029). Subgroup analyses revealed that within patients with poor-prognosis BC, HMGB1⁺ LC3B⁺ double-positive tumors had a better prognosis than BC that lacked one or both of these markers. Altogether, these results suggest that the combined positivity for LC3B⁺ puncta and nuclear HMGB1 is a positive predictor for longer BC survival.     

Feasibility of breast conservation after neoadjuvant taxene based chemotherapy in locally advanced breast cancer: a Prospective Phase I trial

Background

Acute cholecystitis can be the result of retention of bile in the gallbladder with possible secondary infection and ischaemia. The aim of the present study was to investigate whether internal drainage of the gallbladder could protect against the development of acute cholecystitis in a pig model.

Materials and methods

Twenty pigs were randomized to either internal drainage (drained) or not (undrained). Day 0 acute cholecystitis was induced by ligation of the cystic artery and duct together with inoculation of bacteria. Four days later the pigs were killed and the gallbladders were removed and histologically scored for the presence of cholecystitis. Bile and blood samples were collected for bacterial culturing and biochemical analyses.

Results

The histological examination demonstrated statistical significant differences in acute cholecystitis development between groups, the degree of inflammation being highest in undrained pigs. There were no differences in bacterial cultures between the two groups.

Conclusion

Internal drainage of the gallbladder protected against the development of acute cholecystitis in the present pig model. These findings support the theory that gallstone impaction of the cystic duct plays a crucial role as a pathogenetic mechanism in the development of acute cholecystitis and suggest that internal drainage may be a way to prevent and treat acute cholecystitis.     

The optimal dose of CAF regimen in adjuvant chemotherapy for breast cancer patients at stage IIB

This paper deals with the idea of balancing drug effects on tumor and normal cell populations based on a variety of criteria, which is evaluated by the oncologist for breast cancer patients at stage IIB. In this paper, the optimal controller represents the optimal drug dosage of CAF (Cyclophosphamide, Adriamycin and Fluorouracil) regimen in adjuvant chemotherapy after surgery for these patients. We determined the doses of CAF regimen by minimizing a cost function with some constraints. The cost function includes the cancer cell and the normal cell growth dynamics with prescribed weighting coefficients for each patient. The physician determines these weighting coefficients based on some individual parameters. The optimal treatment schedules are computed based on a trade-off between the cancer cell reduction and the normal cell preserving. Numerical simulations are given to illustrate the accuracy of the optimal controller.     

Inhibition of PAI-1 expression in breast cancer carcinoma cells by siRNA at nanomolar range

Plasminogen activator inhibitor type I (PAI-1) plays a central role in metastatic behavior by increasing cells' migratory capacities as shown in several tumoral cell lines. Moreover, in vivo high expression of this factor helps tumoral growth, both by its role in extracellular matrix remodeling and by favoring angiogenesis. High levels of PAI-1 are correlated with bad prognosis in several cancers, particularly in breast cancer. The effect of PAI-1 upon angiogenesis is also involved in atherosclerosis, in which high levels of PAI-1 expression are observed. Breast carcinoma MDA MB 231 cells are known for both having important metastatic capacities and expressing high levels of PAI-1. We have demonstrated in these cells that the transfection of PAI-1 specific small interfering RNAs (siRNA) specifically inhibited the expression of this factor by 91%. We evaluated siRNA activity by determining PAI-1 mRNA level, as well as intracellular and extracellular PAI-1 protein by using RT Q-PCR, Western blot and ELISA analyses, respectively. Data confirmed inhibition at mRNA levels (primary aim of interference), intracellular protein, and secreted PAI-1, the latter being operative successfully in the cell microenvironment. The lipidic vector Delivery Liposomes System (DLS) used was adapted to siRNA delivery as observed by particle size distribution analysis, confocal microscopy and transfection into MDA MB 231, in the presence of serum. SiRNA activity was clearly detected at concentrations as low as 10 nM. Moreover, the low cytotoxicity of this vector makes it a good candidate for future in vivo siRNA delivery.