Urokinase-type plasminogen activator system and breast cancer (Review)

Malignant tumors have a capacity to degrade the extracellular matrix (ECM) by controlled proteolysis. One proteolytic system involved in these processes is the urokinase-type plasminogen activator (uPA) system, which consists of uPA, uPA receptor (uPAR) and uPA inhibitors 1 and 2 (PAI-1 and PAI-2). In the past two decades, study of the uPA system in human breast cancer has yielded valuable insights. Increased levels of uPA, PAI-1 and uPAR have been reported to be associated with poor prognosis in patients with breast cancer. Furthermore, uPA and PAI-1 may be new prognostic markers for axillary node-negative patients. To date, a growing body of evidence has suggested that uPA system promotes tumor metastasis by several different mechanisms, not just by breaking down the ECM. This article is focused on reviewing the current understanding of uPA system members as prognostic markers in breast cancer, and their cellular localization and tissue distribution. Correlations of the uPA system with other informative markers of breast cancer are also discussed.     

Urokinase-type plasminogen activator induces proliferation in breast cancer cells

Urokinase-type plasminogen activator (uPA) is implicated in various pathophysiological processes, including extracellular matrix turnover, cell migration and invasion. Our study aimed to determine the role of uPA in both proliferation and mitogen-activated protein kinase (MAPK) pathway. Hence, we analyzed the effects induced by exogeneous addition of domain-specific uPA antibodies and uPA-interacting molecules on proliferation of uPA-suppressed MDA-MB-231 breast cancer cells. uPA expression was reduced to 53% by stable transfection with an antisense/vector construct and to 65% by siRNA transfection. Immunocytochemical Ki67 staining and flow cytometry (S-phase) analysis indicated a strong decrease of cellular proliferation activity (35% and 38%, respectively). Exogenous addition of high molecular weight-uPA (HMW-uPA) or incubation with the amino terminal fragment (ATF), which lacks the enzymatic activity of uPA, lead to increased cell proliferation. A strong increase of proliferation was absent when the monoclonal anti-uPAR antibody IIIF10 (blocking uPA binding site), soluble uPAR (scavenger effect) and phosphatidyl-inositol-specific phospholipase C (PI-PLC, degrading uPAR) was added prior to the addition of HMW-uPA. In conclusion, HMW-uPA and ATF induce proliferation of breast cancer cells by binding to uPAR. Thereby, integrins situated adjacent to uPAR carry the signals into the cell, thus stimulating proliferation that is mediated via the MAPK pathway.     

Urokinase-type plasminogen activator and its inhibitor type 1 predict disease outcome and therapy response in primary breast cancer

Combined determination of urokinase-type plasminogen activator (uPA) and its inhibitor, activator inhibitor type 1 (PAI-1), supports risk-adapted individualized therapy concepts, particularly in node-negative breast cancer. The prognostic impact of both factors in primary breast cancer was substantiated by a pooled analysis of > 8000 patients with breast cancer and a multicenter prospective randomized therapy trial in node-negative breast cancer; findings achieved the highest level of evidence for tumor biomarkers. Patients with node-negative breast cancer with low antigen levels of uPA and PAI-1 in their primary tumor tissue have a very good prognosis and therefore may be spared the burden of adjuvant chemotherapy, whereas those with elevated uPA/PAI-1 antigen levels carry an increased risk of disease recurrence. Recent retrospective analysis of > 3000 patients indicated that patients with breast cancer with high uPA/PAI-1 values derive a significantly greater benefit from adjuvant chemotherapy than patients with low uPA/PAI-1 levels. Similarly, in the multicenter prospective Chemo N0 trial, administration of cyclophosphamide/methotrexate/5-fluorouracil-based chemotherapy led to a substantial reduction in risk of disease recurrence in patients with high uPA/PAI-1. However, benefit from adjuvant endocrine therapy appears to be independent of a patient's uPA/PAI-1 status. In metastatic breast cancer, retrospective studies showed that elevated uPA or PAI-1 present in the primary tumor tissue are associated with a poor response to later palliative endocrine therapy. These findings suggest that high levels of uPA and/or PAI-1 do reflect an aggressive phenotype that may be overcome or suppressed by early systemic therapy in the adjuvant setting but may be too advanced for response to palliative therapy at a later stage.     

E-钙黏附素与尿激酶型纤溶酶激活物在浸润性乳腺癌中的表达及其临床意义

目的通过检测E-钙黏附素(ECD)和尿激酶型纤溶酶原激活物(UPA)基因产物在乳腺癌组织中的表达情况,探讨它们与乳腺癌浸润和转移及其与乳腺癌生物学行为的关系.方法采用免疫组化SABC方法检测ECD与 UPA在86例乳腺浸润癌(其中淋巴结转移62例,无淋巴结转移24例)中的表达,分析其与临床病理的关系及内在联系.结果淋巴结转移组UPA表达比无淋巴结转移组明显高(P<0.01),而ECD表达则减低(P<0.01).UPA的高表达与乳腺癌的高侵袭力、淋巴转移、组织低分化和病理分期升高有关,且ECD(-)/UPA( )组较ECD( )/UPA(-)组有明显的恶性生物学行为.结论 ECD表达减低和UPA高表达均提示其恶性生物学行为,有助于确定术后治疗方案.     

Urokinase-type plasminogen activator expression correlates with tumor angiogenesis and poor outcome in gastric cancer

Urokinase plasminogen activating system (PA system) and vascular endothelial growth factor (VEGF) were recently suggested to contribute synergistically to tumor progression. To evaluate the roles of the PA system and VEGF in gastric cancer, the effects of the PA system and VEGF on tumor angiogenesis and the survival of patients with gastric cancer were investigated. Cancer tissues from 101 gastric cancer patients were assayed immunohistochemically for expression of urokinase-type plasminogen activator (uPA), uPA receptor (uPAR), PA inhibitor-1 (PAI-1) and VEGF protein. The positive rates of uPA, uPAR, PAI-1, VEGF expression were 22.8%, 32.7%, 36.6% and 26.7%, respectively. Positive staining was observed in tumor cells (uPA, uPAR, VEGF), or in both tumor cells and stromal cells (PAI-1). The expressions of uPA, uPAR, PAI-1 and VEGF were significantly correlated with the clinicopathological factors: uPA, depth of tumor invasion, differentiation, lymphatic and vascular invasion; uPAR, tumor size, depth, lymph node involvement, differentiation, vascular invasion; PAI-1, tumor size, depth, lymph node involvement, differentiation, vascular invasion; VEGF, differentiation, vascular invasion. The microvessel density (MVD) assessed immunohistochemically was significantly higher in the patients with expression of uPA, uPAR or VEGF, and stepwise analysis identified uPA as an independent correlated factor with MVD. Furthermore, multivariate analysis demonstrated that depth of tumor invasion, lymph node involvement and uPA expression were independent prognostic factors. uPA is a key factor in the PA system, being associated with a poor outcome of gastric cancer, and contributing not only to invasive activity, but also to angiogenesis. (Cancer Sci 2003; 94: 43–49)     

Readministration of tamoxifen after adjuvant therapy for recurrent breast cancer

BACKGROUND: Previous series concerning tamoxifen (TAM) rechallenge did not obtain satisfactory results. Using stricter criteria, we now assess the usefulness of readministration of TAM as an initial therapy for patients with recurrent breast cancer. METHOD: The eligibility criteria were postmenopausal, estrogen receptor (ER) positive or unknown, at least 12 months of adjuvant TAM, a 6-month or longer drug-free period and no previous therapy after recurrence. A total of 10 patients were enrolled. TAM was administered in daily doses of 20 or 30 mg. RESULTS: The mean age of the patients at the time of recurrence was 64.8 years. The receptor status was positive in 8 patients and unknown in 2. The median disease-free interval (DFI) after mastectomy was 71.7 months. A complete response was observed in one patient, a partial response in 6, stable disease in 2, and progression in one. The response rate was thus 70%, with an additional two patients showing no progression over 6 months. Although only one patient with a DFI of less than 48 months showed a positive response, all patients with a DFI longer than 48 months showed a clinical response. The duration of response was less than 12 months in 3 patients and longer in 4. CONCLUSION: The post-adjuvant readministration of tamoxifen is a useful first choice therapy for postmenopausal recurrent breast cancer patients with positive ER and longer DFI.     

胃癌组织中E—CD和UPA的表达及临床病理学评价

目的：探讨胃癌浸润、转移中细胞粘附分子ECD和基质分解酶UPA所发生的功能性改变及其与胃癌生物学行为的关系。方法：采用免疫组化SABC法对42例胃癌标本的ECD、UPA表达情况进行研究，分析了其与临床病理因素的关系及内在联系。结果：ECD在42例胃癌中阴性表达为66．7％（28例），UPA阳性率为64．3％（27例），ECD减低的表达，UPA的高表达与胃癌的高侵袭力、淋巴结转移、组织低分化和病理分期升高有关，且ECD（－）／UPA（＋）组较之ECD（＋）／UPA（－）组有明显的恶性生物学行为。结论：ECD表达减低和UPA的高表达均提示胃癌具恶性生物学行为，ECD／UPA共表达对胃癌生物学行为评估有重要意义。     

Urokinase-type plasminogen activator receptor: a beacon of malignancy?

Discriminating between benign and malignant disease is a pivotal diagnostic issue in the care of women with pelvic masses. Identification of serum biomakers that can rellably make the distinction would aid in the proper referral for patient care and may provide leads in the development of early detection strategies.     

尿激酶型纤溶酶原激活因子及抑制剂表达与人大肠癌细胞转移能力的关系

目的　探讨尿激酶型纤溶酶原激活因子 ( u PA)、尿激酶型纤溶酶原激活因子受体 ( u PAR)和纤溶酶原抑制剂 1( PAI- 1)的表达与人大肠癌细胞系转移能力的关系。方法　用 EL ISA方法测定 3个人大肠癌细胞系培养上清液中 u PA、u PAR和 PAI- 1含量 ;用免疫组化 ABC方法检测 u PA、u PAR和 PAI- 1在细胞中的表达 ;分析其表达与大肠癌细胞转移能力的关系。结果　在培养上清液中具有高转移能力的 HT- 2 9d细胞的 u PA、u PAR及 PAI- 1含量明显高于低转移的 HT- 2 9和不转移的 Wi Dr细胞 ,而 HT- 2 9细胞的 u PA、u PAR及 PAI- 1含量则高于 Wi Dr细胞。免疫组化显示 u PA和 PAI- 1在 HT- 2 9d细胞中的表达高于 HT- 2 9和 Wi Dr细胞。结论　 u PA、u PAR和 PAI-1的表达与大肠癌细胞的转移能力密切相关。     

Relationship of PS2 with response to tamoxifen therapy in patients with recurrent breast cancer.

PS2, an oestrogen-inducible protein, was measured in the cytosol of 230 primary tumours from patients who were subjected to first-line tamoxifen therapy for advanced disease without prior adjuvant therapy with tamoxifen. PS2 correlated positively with oestrogen receptor (ER, P < 0.01) and progesterone receptor content (PgR, P < 0.001), and with the length of progression-free survival (PFS, P = 0.05). Although not statistically significant, higher levels of PS2 (> or = 10 ng mg-1 protein) were also associated with increased probability of response to tamoxifen treatment and a longer total post-relapse survival (PRS). ER, PgR, menopausal status, site of disease and prior adjuvant chemotherapy were all associated with response to tamoxifen therapy and with PFS. In multivariate analysis for PFS, low levels of ER and PgR, visceral metastasis, a disease-free interval of less than 1 year and prior adjuvant chemotherapy were all significantly associated with an increased probability of a rapid disease progression after start of tamoxifen therapy. In the subset of 83 tumours with intermediate levels of ER and PgR (both > or = 10, but not both > or = 75 fmol mg-1 protein), PS2 was positively related with the length of PFS (P < 0.01) and PRS (P < 0.05). PS2 remained the strongest factor in multivariate analysis for PFS (P < 0.01) in this ER/PgR intermediate subgroup, but was not of predictive value in univariate or multivariate analysis for both PFS and PRS in tumours classified as ER/PgR low or high (> or = 75 fmol mg-1 protein). It is concluded that PS2 status may be used as a parameter, additional to ER and PgR, for better refinement of prediction of response to tamoxifen treatment in advanced breast cancer patients especially with intermediate ER/PgR levels in their primary tumour.     

Urokinase-type plasminogen activator and plasminogen activator inhibitor type 1 and type 2 in stage I malignant melanoma

The urokinase-type plasminogen activator (uPA) and its inhibitors type 1 (PAI-1) and type 2 (PAI-2) are considered to have a key role in the process of invasion and metastasis. We investigated the differences in uPA, PAI-1 and PAI-2 concentrations in primary cutaneous melanoma and normal skin and correlations with well-established melanoma prognostic factors. The study was performed on 43 patients (19 men, 24 women; mean age 57 years) with histologically confirmed primary melanomas <1.5 mm thick. The uPA concentrations were determined in 36 pairs of triton extracts, and the PAI-1 and PAI-2 concentrations in 43 pairs of cytosols prepared from the tumour and adjacent normal tissue samples (matched pairs). The uPA, PAI-1 and PAI-2 concentrations were measured by enzyme-linked immunosorbent assay (ELISA). Significantly higher concentrations of both uPA and PAI-1 were measured in melanomas than in normal surrounding skin (uPA: 1.08 vs 0.48 ng/mg total protein (mgp), p<0.001; PAI-1: 14.07 vs 2.07 ng/mgp, p<0.001). The melanoma uPA, PAI-1 and PAI-2 concentrations correlated significantly (p<0.05) with normal skin (r=0.73, 0.54, 0.38 respectively). The uPA concentrations positively correlated with those of PAI-1 measured in melanomas (r=0.45, p<0.01). PAI-1 values were significantly lower (p<0.001) in the melanomas of Breslow thickness < or =0.75 mm, Clark invasion 0.75 mm, Clark invasion of > or =II and < or =III, with microscopic ulceration and vascular invasion (22.25, 17.67, 27.67, 37.77, respectively). Determination of uPA and PAI-1 can provide significant additional prognostic information for melanoma patients.     

Clinical utility of urokinase-type plasminogen activator and plasminogen activator inhibitor-1 determination in primary breast cancer tissue for individualized therapy concepts

Invasion factors urokinase-type plasminogen activator (uPA) and its plasminogen activator inhibitor (PAI-1) are the only novel tumor biological prognostic factors validated at the highest level of evidence with regard to their clinical utility in breast cancer. Antigen levels of both factors present in extracts of primary tumor tissue are determined by standardized, quality-assured enzyme-linked immunosorbent assays. Numerous studies showed that patients with low levels of uPA and PAI-1 have a significantly better survival than patients with high levels of either factor. Recently, these data have been validated by a European Organization for Research and Treatment of Cancer pooled analysis comprising more than 8000 breast cancer patients. The particular combination of both factors, uPA/PAI-1 (both low vs. either or both factors high), outperforms the single factors as well as other traditional prognostic factors with regard to risk group assessment, particularly in node-negative breast cancer. Node-negative breast cancer patients with low levels of uPA and PAI-1 have a very good prognosis and, as such, may be candidates for being spared the burden of adjuvant chemotherapy. In contrast, node-negative patients with high uPA/PAI-1 are at a substantially increased risk of relapse, comparable to that of patients with > or = 3 involved axillary lymph nodes. First results from a multicenter prospective randomized therapy trial in node-negative breast cancer (Chemo N(0)) as well as recent retrospective analyses indicate that these high-risk patients benefit from adjuvant chemotherapy. Thus, combined determination of the invasion factors uPA and PAI-1 supports risk-adapted individualized therapeutic strategies in patients with primary breast cancer, particularly in those with node-negative breast cancer.     

Tissue-type plasminogen activator in breast cancer: relationship with estradiol and progesterone receptors

Plasminogen activator (PA) is an estradiol-inducible enzyme and therefore a potential marker for a functional estradiol receptor (ER) in human breast carcinomas. In this investigation tissue-type PA (t-PA) correlated significantly with both ER and progesterone receptors (PR) in human breast carcinomas. In contrast, neither total PA activity nor urokinase-like PA showed any significant correlation with either ER or PR. Other proteases such as a trypsin-like protease, a chymotrypsin-like protease, and cathepsin B also showed no correlation with ER and PR. It was concluded that the t-PA form of PA may be a marker for a functional ER in breast carcinoma and thus be of value in predicting hormone-dependent breast cancers.     

Predictive impact of urokinase-type plasminogen activator: plasminogen activator inhibitor type-1 complex on the efficacy of adjuvant systemic therapy in primary breast cancer

One of the most thoroughly studied systems in relation to its prognostic relevance in patients with breast cancer, is the plasminogen activation system. This system comprises of, among others, the urokinase-type plasminogen activator (uPA) and its main inhibitor (PAI-1). In this study we investigated whether the uPA:PAI-1 complex is associated with the responsiveness of patients with primary breast cancer to adjuvant systemic therapy. Quantitative enzyme-linked immunosorbent assays were used to assess the levels of uPA, PAI-1, and uPA:PAI-1 complex in 1119 tumors of patients with primary invasive breast cancer. These patients were followed for a median follow-up time of 59 months (range, 2-267 months) after the primary diagnosis. Correlations with well-known clinicopathological factors, and univariate and multivariate survival analyses were performed. High uPA:PAI-1 complex levels were correlated with an adverse histological grade, and inversely associated with negative estrogen and progesterone receptor status. High tumor levels of uPA:PAI-1 complex predicted an early relapse in the univariate relapse-free survival analysis (P < 0.001). The multivariate analysis showed that high uPA:PAI-1 complex levels were associated with a decreased relapse-free survival time (P = 0.033), independently of age, tumor size, number of lymph nodes affected, progesterone receptor status, uPA, adjuvant endocrine, and chemotherapy. More important, it was demonstrated that there is a larger benefit from adjuvant chemotherapy for patients with higher versus lower tumor levels of uPA:PAI-1 complex. The results of this study imply that the expression of uPA:PAI-1 complex independently predicts the efficacy of adjuvant chemotherapy in patients with primary breast cancer.     

Arena: Overprolonged adjuvant tamoxifen therapy in breast cancer

While adjuvant tamoxifen therapy given continuously for 2–3years can lead to a modest improvement in survival rates inearly breast cancer, there is no evidence that prolonging tamoxifenadministration beyond that time is likely to improve survivalrates any further in unselected cases. In the case of advanceddisease, an alternating tamoxifen/progestagen regimen has beenshown to increase the response rate and also its duration, beyondthat to be expected from either agent alone. The next generationof adjuvant trials in breast cancer needs to explore the potentialof an alternating tamoxifen/megestrol regimen. breast cancer, tamoxifen therapy, progestagen therapy, megestrol therapy, adjuvant therapy     

Prognostic significance of urokinase-type plasminogen activator and plasminogen activator inhibitor-1 in primary breast cancer.

The uPA-mediated pathway of plasminogen activation is central to cancer metastasis. Whether uPA and PAI-1 are related to local recurrence, metastatic spread or both is not clear. We present a retrospective study of 429 primary breast cancer patients with a median follow-up of 5.1 years, in which the levels of uPA and PAI-1 in tumour extracts were analysed by means of an enzyme-linked immunosorbent assay. The median values of uPA and PAI-1, which were used as cut-off points, were 4.5 and 11.1 ng mg(-1) protein respectively. The levels of uPA and PAI-1 were correlated with tumour size, degree of anaplasia, steroid receptor status and number of positive nodes. Patients with high content of either uPA or PAI-1 had increased risk of relapse and death. We demonstrated an independent ability of PAI-1 to predict distant metastasis (relative risk 1.7, confidence limits 1.22 and 2.46) and that neither uPA nor PAI-1 provided any information regarding local recurrence.     

The urokinase plasminogen activator system as a target for prognostic studies in breast cancer

The identification of patients at high risk of relapse is currently one of the most important issues in breast cancer research. However, the selection of high-risk patients continues to be difficult due to the unpredictable course of this disease. Axillary lymph node status is currently recognized as the best clinical discriminant between good and poor prognosis, yet almost 30% of node- negative patients and 65% of node-positive patients will experience a relapse. Additional prognostic markers are therefore urgently needed.Since metastatic disease is the main cause of cancer patient morbidity and mortality, the measurement of molecules functionally involved in the regulation of tumor invasion and metastasis is attractive as a means to predict prognosis.Cancer invasion is a complex process in which degradation of the extracellular matrix plays a crucial role. This degradation is accomplished by the concerted action of several proteolytic enzyme systems, including generation of plasmin by the urokinase pathway of plasminogen activation, matrix metallo-proteases, and other extracellular proteases. Increased expression and secretion of urokinase plasminogen activator (uPA) strongly correlates with the malignant phenotype of many types of cells, and the central role of uPA in tumor invasion is now well established.This review will focus on the prognostic impact of components of the urokinase plasminogen activation system in breast cancer with emphasize on methodological issues.     

Dissemination risk index based on plasminogen activator system components in primary breast cancer.

PURPOSE: To study interactions between disease-free survival (DFS) and four components of the plasminogen activator system: urokinase-type plasminogen activator (uPA), its two inhibitors (PAI-1 and PAI-2), and its membrane receptor uPAR. PATIENTS AND METHODS: We conducted a retrospective study of 499 primary breast cancer patients (median follow-up, 6 years). uPA, PAI-1, and PAI-2 were determined on cytosols and uPAR on solubilized pellets, using enzyme-linked immunoadsorbent assay kits (American Diagnostica, Greenwich, CT). Classical univariate and multivariate statistical methods were used together with multiple correspondence analysis to graphically examine interactions between the variables and outcome. RESULTS: By univariate analysis, higher uPA and PAI-1 values were significantly related to shorter DFS (P =.002; P <.00002). PAI-2 was not significantly related to DFS, although patients with high and very low PAI-2 values had a longer DFS. Multiple correspondence analysis showed the parallel impact of uPA and PAI-1 on outcome, and the clearly different behavior of PAI-2 compared with PAI-1. The prognostic contribution of uPAR seemed weak by both methods. A dissemination risk index [uPA x PAI-1/(PAI-2 + 1)], taking into account the modulation of uPA proteolytic activity by the ratio of its two inhibitors, was then tested. Dissemination risk index was selected as an independent variable in the Cox model in the overall population (P <.000001) and in node-positive patients (P <.00001). It was the only variable selected in node-negative patients (P =. 003). CONCLUSION: A dissemination risk index determined on primary tumor and taking into account the different effects of PAI-1 and PAI-2 on uPA can be of major help in clinical management of breast cancer, particularly in node-negative patients.