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Serum levels of matrix metalloproteinase 2 in patients with breast cancer
Abstract
Matrix metalloproteinases (MMPs) have been reported to be associated with invasive and metastatic behaviors of human malignant tumors. However, there is still limited knowledge about the role of matrix metalloproteinases-2 (MMP-2) in breast cancer. This study was designed with the aim to elucidate the possible relationship between the preoperative circulating MMP-2 and breast cancer. Fifty-seven consecutive patients with invasive breast cancer undergoing surgery were prospectively included and evaluated. Venous blood samples were collected before the surgery. Sera were obtained by centrifugation, and stored at -70 degrees C until assayed. The control group consisted of 12 patients with benign breast tumor (six with fibrocystic disease and six with fibroadenoma). Serum concentrations of MMP-2 were measured by the quantitative sandwich enzyme immunoassay technique. The data on primary tumor stage, age, estrogen receptor, lymph node status, and TNM staging were reviewed and recorded. The mean value of serum MMP-2 in patients with invasive breast cancer was 694.3+/-140.5 ng/ml and those of control group were 593.3+/-134.0 ng/ml and the difference was significant (P=0.026). Furthermore, there were significantly higher serum levels of MMP-2 in the patients with more advanced primary tumor staging (P=0.005), in the patients with more advanced lymph node status(P=0.011) and in the patients with more advanced TNM staging (P<0.001). In multivariate analysis, TNM staging (P<0.001) appeared as independent factor regarding the significant higher serum levels of MMP-2. Patients with more advanced TNM staging were shown to have higher serum MMP-2 levels. Thus preoperative serum MMP-2 levels might reflect the severity of invasive breast cancer and deserve further evaluation.
... The over-expression of MMPs is closely related to tumor invasion and metastasis [3,4]. As one of the most important MMPs, MMP-2 has been identified as a biomarker of many tumor diseases, such as breast cancer and ovarian cancer [5][6][7][8]. Therefore, the determination of MMP-2 plays a key role in clinical diagnosis. ...
... The fresh normal human serum was purchased from Nanjing Liangwei Biotechnology Co. LTD. 6-mercapto-1-hexanol (MCH) was obtained from Sigma-Aldrich, tris(2-dimethylaminoethyl) amine (Me 6 TREN), CuBr 2 , LiClO 4 , KPF 6 , Tris(hydroxymethyl)methyl aminomethane, 2bromo-2-phenylacetic acid and ZrOCl 2 ⋅8H 2 O were provided by J&K Scientific Ltd (Shanghai, China). FMMA was purchased from Shanghai bide Pharmaceutical Technology Co., Ltd. ...
... FMMA was purchased from Shanghai bide Pharmaceutical Technology Co., Ltd. N,N-dimethylformamide (DMF), K 3 [Fe(CN) 6 ]、K 4 [Fe(CN) 6 ], KCl, KBr, Na 2 HPO 4 , NaH 2 PO 4 and Hydrochloric acid (HCl) were purchased from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China). ...
In this paper, a peptide-based biosensor was developed for sensitive and selective analysis of matrix metalloproteinase 2 (MMP-2) through a signal amplification strategy of electrochemically mediated atom transfer radical polymerization (eATRP). Briefly, α-bromophenylacetic acid (BPAA) initiators were connected to the end of the MMP-2 specific peptide, and then ATRP reaction was carried out with ferrocenylmethyl methacrylates (FMMA) monomers under the action of copper activators to obtain the polymers of Fc. The presence of target MMP-2 caused the part of peptides were specifically cleaved, and the electrochemical signal decreased of Fc. As a result, there was a linear relationship between Fc signal change value and MMP-2 concentration at 8 fM to 80 pM range. The MMP-2 detection limit was calculated as low as 0.53 fM, which was much lower than that in the reported literatures. The proposed peptide-based biosensor showed a good performance for the determination of MMP-2 in human serum samples. This highly efficient and simple method would have great potential in the clinical analysis of matrix metalloproteinase.
... Increased levels of MMP-2 protein in serum have been observed in breast cancer 18,19 . Sheen-Chen et al 18 found higher MMP-2 levels correlating with advanced tumour staging and advanced lymph node status. ...
... Increased levels of MMP-2 protein in serum have been observed in breast cancer 18,19 . Sheen-Chen et al 18 found higher MMP-2 levels correlating with advanced tumour staging and advanced lymph node status. However, Patel et al 19 observed that raised MMP-2 levels did not correlate with clinicopathological factors in breast cancer. ...
... Another study focussing on MMPs and their inhibitors in pancreatic cancer patients also reported elevated levels of MMP-2 23 . This was in agreement with the observations of Sheen-Chen et al 18 There are studies that have assessed not only concentration of total MMP-2 in the blood but also measured the activity of MMP-2 in serum or plasma with conflicting results. Decock et al 28 reported that the total MMP-2 concentration in plasma of patients with primary breast cancer was not significantly higher as compared to controls, but the MMP-2 activity was observed to be significantly increased in cancer patients than control. ...
Background & objectives:
Pancreatic cancer has a propensity for wide stromal invasion. Matrix metalloprotease-2 (MMP-2) is a protease that degrades the peri-tumoural tissue and helps in tumour dissemination. Thus, this study was aimed to assess any association of plasma MMP-2 levels with clinicopathological parameters and survival of patients with pancreatic cancer.
Methods:
Plasma samples from 127 pancreatic cancer patients were analyzed for MMP-2 levels by ELISA. Survival and other clinicopathological parameters of patients were analyzed for any correlation with plasma MMP-2 levels.
Results:
The mean MMP-2 levels in pancreatic cancer patients were 560.3±222.0 ng/ml which were significantly elevated compared to chronic pancreatitis patients (P<0.001) and healthy individuals (P<0.05). The plasma levels of MMP-2 significantly correlated with tissue expression of this protease (P=0.004). However, MMP-2 levels did not exhibit any association either with clinicopathological parameters or with survival.
Interpretation & conclusions:
Elevated MMP-2 levels were observed in blood of pancreatic cancer patients which correlated with its tissue expression. However, these levels did not associate with survival or any clinicopathological parameters of patients. Further studies need to be done to confirm the prognostic/ clinical significance of MMP-2 in cancer patients before and after surgery.
... FLI [6], PAI [21], MRI [22], PET [23], CHL imaging [24], and PL imaging [3] IETD [25] DDYVADC [26] MMPs GPLGVRGY [27] Expression levels of MMP-2 and MMP-9 are significantly elevated in most solid tumors, including breast cancer, bladder cancer, colon cancer, and prostate cancer [28]. For example, the MMP-2 concentration in normal human serum is 593.3 ± 134.0 ng/mL, whereas the MMP-2 concentration in serum of breast cancer patients is 694.3 ± 140.5 ng/mL [29]. The expression level of inactive platform MMP-2 (Mr: 72000) in normal cells and bladder cancer cells are 2.7 ± 0.6 units and 3.0 ± 0.5 units, respectively. ...
... The expression level of inactive platform MMP-2 (Mr: 72000) in normal cells and bladder cancer cells are 2.7 ± 0.6 units and 3.0 ± 0.5 units, respectively. Active species of MMP-2 (Mr: 59000 and Mr: 62000) are highly expressed in malignant tumor cells when compared with normal cells (1.9 ± 0.4 units versus 0.07 ± 0.05 units, respectively) [29]. In bladder tissue, the concentration of MMP-9 is 1.7 ± 0.8 units in normal cells and 136.0 ± 4.0 units in bladder cancer cells [30]. ...
Exploring and understanding the interaction of changes in the activities of various enzymes, such as proteases, phosphatases, and oxidoreductases with tumor invasion, proliferation, and metastasis is of great significance for early cancer diagnosis. To detect the activity of tumor-related enzymes, various molecular probes have been developed with different imaging methods, including optical imaging, photoacoustic imaging (PAI), magnetic resonance imaging, positron emission tomography, and so on. In this review, we first describe the biological functions of various enzymes and the selectively recognized chemical linkers or groups. Subsequently, we systematically summarize the design mechanism of imaging probes and different imaging methods. Finally, we explore the challenges and development prospects in the field of enzyme activity detection. This comprehensive review will provide more insight into the design and development of enzyme activated molecular probes.
... However, there are other less specific methods to measure circulating levels. Studies that have examined circulating levels of MMP-2 and breast cancer risk have measured either the latent, active or concentrations of both combined (latent plus active) (25)(26)(27)(28)(29)(30), but two studies did not specify the form (31,32). Eight research articles on circulating levels of MMP-2 are presented in Table I. ...
... Two studies found that serum mean levels of MMP-2 are higher in patients with breast cancer than in healthy patients (694.3 ng/ml vs. 593.3 ng/ml (31), 806.5 ng/ml vs. 771.2 ng/ml (32), p<0.05). ...
MMP-2 and MMP-9 genes have been suggested to play a role in breast cancer. Their functions have been associated with invasion and metastasis of breast cancer; however, their involvement in the development of the disease is not well-established. Herein, we reviewed the literature investigating the association between circulating levels and polymorphisms of MMP-2 and MMP-9 and breast cancer risk. Various studies report conflicting results regarding the relationship of polymorphisms in MMP-2 and MMP-9 and breast cancer risk. Nevertheless, it appears that the T allele in rs243865 and rs2285053 in MMP-2 are associated with reduced risk of breast cancer. In addition, high levels of latent form and low levels of active form of MMP-2 were observed in breast cancer patients compared to controls. For MMP-9, high latent levels and low total levels were found in breast cancer patients compared to controls. Additional studies are needed to comprehend the role of these genes in breast carcinogenesis.
... In several tumors, MMPs have been linked to tumor growth, metastasis and clinical outcome [7,10,[18][19][20]23,[81][82][83]. It has also been shown that proMMP-2 and proMMP-9 expression is increased in malignant cancers compared to benign cancers [84][85][86]. ...
... A number of studies have linked elevated MMP-2 and -9 expression to increased metastasis and tumor stage, e.g. malignant versus benign breast tumors and advanced ovarian tumors versus benign [84][85][86]. In several invasive cell lines, MMP-9 expression was increased compared to non-invasive counterparts [154,155]. ...
In the past decades, a lot of effort has been put in identifying the role of matrix metalloproteinases (MMPs) in cancer. The main role of MMPs in angiogenesis, tumor growth and metastasis is degradation of extracellular matrix (ECM) and release and/or activation of growth factors through their degradative activity. The degradative activity finally results in cancer progression. MMP-inhibitors (MMPIs) have already been designed and tested, based on the degradative role of MMPs in cancer progression. First clinical trials with MMPIs have been performed with disappointing results, showing that in order to use MMP-inhibition the mechanisms underlying MMP-expression in cancer have to be further elucidated. This paper reviews the mechanisms of MMPs on molecular and cellular level and discusses the role for MMPs and MMP-inhibition in cancer with special focus on acute leukemia.
... [19][20][21][22][23], whereas few others have suggested inconsistent results (24,25). Although there were several studies on the association between the levels of serum MMP-2 and clinicopathologic factors, the results were inconsistent across the studies (26)(27)(28)(29)(30)(31)(32)(33)(34)(35). So far, only a few studies have investigated the association between MMP-2 levels and breast cancer prognosis (36,37). ...
... Other studies on the association between MMP-2 levels and clinicopathologic factors were not inconsistent. While several studies reported that higher serum levels of MMP-2 were shown in the patients with more advanced tumor stage (26,28,33,34), the others showed lower MMP-2 levels in patients with many poor prognostic factors including advanced tumor stage or grade (27,29,35). ...
Matrix metalloproteinase-2 (MMP-2) has been thought of as a predictor of recurrence or metastasis risk or prognostic markers in cancer. We evaluated whether preoperative serum levels of MMP-2 work as a prognostic biomarker in breast cancer prognosis.
Preoperative serum levels of MMP-2 were measured with ELISA in 303 patients with histologically confirmed breast cancer. The median follow-up time for all patients was 4.24 years. The relationship of MMP-2 to survival was investigated using Cox proportional hazard regression model adjusted for the tumor-node-metastasis (TNM) stage and estrogen receptor (ER) status.
In the multivariate analysis, disease-free survival (DFS) was worse among patients with the third tertile of MMP-2 level than with the first tertile of MMP-2 level [hazard ratio, 1.80; 95% confidence interval (CI), 1.04-3.11; P = 0.04]. However, when the patients were stratified by age, ER status, histologic grade, and nuclear grade, inverse correlation was shown between serum MMP-2 levels and prognostic factors, and the associations between MMP-2 and DFS were only significant among patients with poor prognostic factors (HR, 2.75; 95% CI, 1.32-5.73 in ER-negative; HR, 2.90; 95% CI, 1.42-5.92 in histologic grade III; and HR, 2.61; 95% CI, 1.26-5.39 in nuclear grade III).
Our results suggest that the preoperative serum levels of MMP-2 were associated with the survival in patients with breast cancer in ER-negative, higher histologic grade, or higher nuclear grade breast cancers.
Our results indicate that serum levels of MMP-2 may play a role as prognostic biomarker in breast cancer survival.
... Moreover, the components of the stroma promote the growth of epithelial cells by secreting growth factors that promote cell proliferation [16]. Some studies suggest a positive association of circulating levels of MMP-2 and MMP-9 with breast cancer risk [17][18][19][20][21][22]. Concerning polymorphisms located in these genes, the rs243865 in MMP-2 could be associated with breast cancer risk, but it is not a consistent finding. ...
Background
Matrix metalloproteinases (MMP)-2 and -9 may play an important role in adipogenesis and carcinogenesis. We investigated whether some polymorphisms located in these genes are associated with body adiposity and mammographic breast density, which are risk factors for breast cancer.Methods
Our study population included 731 premenopausal women. Multivariate generalized linear models were used to evaluate the association of polymorphisms rs243865 in MMP-2 and rs3918242, rs17576, rs2250889 and rs2274756 in MMP-9 with anthropometric factors that refer to adiposity and mammographic features (percent density, dense area and non-dense area) measured by computer-assisted method.ResultsThe number of copies of rs243865 T allele in MMP-2 was associated with increased means of anthropometric factors (ptrend < 0.05 for all except waist-to-hip ratio). The same allele of rs243865 was associated with decreased mean percent density (ptrend = 0.036) and increased mean non-dense area (ptrend = 0.031) when adjusted for potential confounders, but these associations were attenuated when further adjusted for adiposity.Conclusion
These findings suggest that the relation between rs243865 in MMP-2 and mammographic features could be mediated by adiposity.
... MMPs play a vital role in tumor growth, proliferation, angiogenesis and invasion. The secretion and activation of MMP2 and MMP9 are linked with the degradation of ECM and promotion of tumor metastasis [74,75]. The cell migration assay and colony formation assays revealed the efficiency of the algal bioactives to inhibit cell invasion and prolonged proliferation and correlated with the expression of MMPS. ...
Background:
Lung adenocarcinoma is the most common subtype of Non small cell lung cancer in which the PI3K/Akt cascade is frequently deregulated. The ubiquitous expression of the PI3K and the frequent inactivation of PTEN accounts for the prolonged survival, evasion of apoptosis and metastasis in cancer. This has led to the development of PI3K inhibitors in the treatment of cancer. Synthetic PI3K inhibitors undergoing clinical and preclinical studies are toxic in animals. Hence, there is a critical need to identify PI3K inhibitor(s) of natural origin. The current study aims to explore the efficacy of the red algae Gelidiella acerosaon inhibition of cell proliferation, migration and the expression of cell survival genes in lung adenocarcinoma cell line A549.
Methods:
The phytoconstituents of Gelidiella acerosa were extracted sequentially with solvents of different polarity, screened qualitatively and quantitatively for secondary metabolites and characterized by GC-MS. The in-vitro studies were performed to check the efficacy of the extract on cell proliferation (MTT assay), cell invasion (scratch assay and colony formation assay), apoptosis (fluorescent, confocal microscopy and flow cytometry) and expression of apoptosis and cell survival proteins including PI3K, Akt and GSK3β and matrix metalloproteinase MMP2 and MMP9 by Western blot method. The antitumor activity of GAE was analyzed in a tumor model of Zebrafish.
Results:
The outcomes of the in vitro analysis showed an inhibition of cell proliferation, induction of apoptosis, inhibition of cell migration and colonization by the crude extract. The analysis of protein expression showed the activation of caspases 3 and Pro apoptotic protein Bax accompanied by decreased expression of Bcl-2 and Bcl-XL. On the other hand, subsequent activation of GSK3β and down regulation of PI3K, Akt were observed. The decreased expression of MMP2 correlated with the antimetastatic activity of the extract. The in vivo studies showed an inhibition of tumor growth by GAE in Zebrafish.
Conclusion:
The phytoconstituents of algal extract contributed to the anticancer properties as evidenced by in vitro and in vivo studies. These phytoconstituents can be considered as a natural source of PI3K/Akt inhibitor for treatment of cancers involving the PI3K cascade.
... Increased expression of these markers correlates with increased severity of disease. Mmp2 and mmp9 can both be measured in serum and plasma, making them potentially useful blood-based markers for diagnosis, prognosis, and disease monitoring in breast cancer patients (171). We are currently measuring the levels of total and active mmp2 and mmp9 in plasma and serum of patients at high risk for breast cancer, comparing them with those of patients with benign breast disease and patients with breast cancer, in order to determine the prognostic significance of altered MMP levels in these patients. ...
Breast cancer is a complex disease that still imposes a significant healthcare burden on women worldwide. The etiology of breast cancer is not known but significant advances have been made in the area of early detection and treatment. The advent of advanced molecular biology techniques, mapping of the human genome and availability of high throughput genomic and proteomic strategies opens up new opportunities and will potentially lead to the discovery of novel biomarkers for early detection and prognostication of breast cancer. Currently, many biomarkers, particularly the hormonal and epidermal growth factor receptors, are being utilized for breast cancer prognosis. Unfortunately, none of the biomarkers in use have sufficient diagnostic, prognostic and/or predictive power across all categories and stages of breast cancer. It is recognized that more useful information can be generated if tumors are interrogated with multiple markers. But choosing the right combination of biomarkers is challenging, because 1) multiple pathways are involved, 2) up to 62 genes and their protein products are potentially involved in breast cancer-related mechanisms and 3) the more markers evaluated, the more the time and cost involved. This review summarizes the current literature on selected biomarkers for breast cancer, discusses the functional relationships, and groups the selected genes based on a Gene Ontology™ classification.
... Many different studies have investigated the serum levels of MMP-2, MMP-9 and VEGF in cancer patients with different and sometimes contradictory results 8,[24][25][26][27][28][29][30][31][32] . We wanted to incorporate these arguable markers into specific clinical events of two narrowly defined groups of cancer patients to prove or disprove their clinical utility. ...
Background and aims:
Metastases are a severe complication in cancer patients and biomarkers predicting their progression are still lacking for specific groups of patients. HER2 positive breast cancer (HER2 BC) patients on trastuzumab therapy are at risk of the development of unpredictable and often fatal central nervous system (CNS) metastases and castration resistant prostate cancer (CRPC) patients urgently need a marker of disease progression during therapy. Proposed metastatic markers: circulating tumor cells (CTC), serum levels of matrix metalloproteinase 2 (MMP-2), 9 (MMP-9) and vascular endothelial growth factor (VEGF) were prospectively studied to confirm their utility in these two narrowly defined groups of cancer patients.
Patients and methods:
The groups comprised 44 advanced HER2 BC, 24 CRPC patients and 42 healthy controls. An immunomagnetic separation method followed by PCR and electrophoretic detection (AdnaGen, Germany) were used for CTC determination. Serum marker levels were determined by the ELISAs (R&D System, USA).
Results:
MMP-2 serum level was significantly higher in HER2 BC patients who developed CNS metastases, especially if there were also bone metastases. CTCs were a negative predictive marker for overall survival in HER2 BC patients. MMP-9 serum level was significantly higher in CRPC patients in whom disease progression occurred. CTC vanished from the blood of most of the CRPC patients (from 88% to 37%) during chemotherapy.
Conclusion:
MMP-2 serum level and CTCs show the potential to predict CNS metastases and overall survival in BC patients. CTCs and MMP-9 serum level could be a promising therapy response marker in CRPC patients.
... Liczne doniesienia naukowe mówią o wzroście aktywności żelatynazy A i żelatynazy B w surowicy krwi w następstwie rozwoju raka piersi [36]. Poziom MMP koreluje z agresywnością guza, i co za tym idzie, z ryzykiem zgonu [37,38]. Enzymy te stanowią istotny czynnik pozwalający określić stan zaawansowania choroby, co może w znaczącym stopniu ułatwić prognozowanie jej przebiegu i dobór odpowiedniej terapii [3,38]. ...
... MMP-2 and -9 activity in serum or plasma, measured via quantitative gelatin zymography, has shown potential for discrimination among breast cancer subclassifications of varying risk (54,55), for prediction of lymph node metastasis (56), and for assessment of response to therapy in breast cancer patients (57). Some studies have shown measurement of MMP-9 protein in serum by ELISA (56,(58)(59)(60) or by Luminex multiplexed protein assays (61,62) to provide an effective alternative measure of similar prognostic value, and high serum MMP-2 measured by ELISA has also been associated with poor prognosis (63,64). By contrast, one recent large study of 465 breast cancer patients specifically examining the © 1996-2015 concentration of MMP-9/TIMP-1 complexes in plasma by ELISA and by in-solution proximity ligation assay found no correlation of this complex with disease-free survival (65). ...
Members of the matrix metalloproteinase (MMP) family have been identified as poor prognosis markers for breast cancer patients and as drivers of many facets of the tumor phenotype in experimental models. Early enthusiasm for MMPs as therapeutic targets was tempered following disappointing clinical trials that utilized broad spectrum, small molecule catalytic site inhibitors. However, subsequent research has continued to define key roles for MMPs as breast cancer promoters, to elucidate the complex roles that that these proteins play in breast cancer development and progression, and to identify how these roles are linked to specific and unique biochemical features of individual members of the MMP family. Here, we provide an overview of the structural features of the MMPs, then discuss clinical studies identifying which MMP family members are linked with breast cancer development and new experimental studies that reveal how these specific MMPs may play unique roles in the breast cancer microenvironment. We conclude with a discussion of the most promising avenues for development of therapeutic agents capable of targeting the tumor-promoting properties of MMPs.
... For example, a correlation between levels of uPA in tumour tissue and serum was found, and high levels indicated poor prognosis in patients with soft-tissue sarcomas [165]. The proteases in serum and plasma specimens can be detected either by immunological methods [126,140] or by using activity-based assays with zymographic- [190], optical [117,135] or MS monitoring [34,36,47,52] of the proteolytic substrate turnover. The diagnostic power of protease profiling with immunological methods has been recently demonstrated and the combined measurement of cathepsin B, cathepsin L, and uPA in serum specimens has been proved to be a more sensitive indicator for colorectal cancer than These are not the final page numbers commonly used tumour markers [166]. ...
Targeted MS is becoming increasingly important for sensitive and specific quantitative detection of proteins and respective PTMs. In this article, Ceglarek et al. [Proteomics Clin. Appl. 2013, 7, 794-801] present an LC-MS-based method for simultaneous quantitation of seven apolipoproteins in serum specimens. The assay fulfills many necessities of routine diagnostic applications, namely, low cost, high throughput, and good reproducibility. We anticipate that validation of new biomarkers will speed up with this technology and the palette of laboratory-based diagnostic tools will hopefully be augmented significantly in the near future.
... These events require diverse proteolytic enzymes, among which MMP-2 and MMP-9 play a significant role in degradation of type IV collagen, the major component of the basement membrane (Shah et al., 2009). Elevated levels of both MMP-2 and MMP-9 have been observed in blood from breast cancer patients and were repeatedly found to be associated with advanced stage, lymph node metastasis, and poor prognosis (Coskun et al., 2007; Farias et al., 2000; La Rocca et al., 2004; Leppa et al., 2004; Liu et al., 2006; Quaranta et al., 2007; Ranuncolo et al., 2003; Sheen-Chen et al., 2001; Talvensaari-Mattila and Turpeenniemi-Hujanen, 2005; Zucker et al., 1993 Zucker et al., , 1999). MMP-9 is abundantly expressed in various malignant tumors and is postulated to play a critical role in tumor invasion and angiogenesis (Chandler et al., 1997; Liabakk et al., 1996). ...
Ethnopharmacological relevance
Iridoid glycosides have been associated with decreased risks of cancer, such as hepatocarcinoma. Although Picrorrhiza kurroa has shown activity against hepatocarcinogenesis, its mechanism of action is poorly understood, further the anticancer activity of iridoid glycosides present in this plant has not been tested so far.
Aim of the study
Here, MCF-7 cell lines (Human breast cancer) were used to test whether P. kurroa extract (PE) and its isolated iridoid glycosides Picroside I (PS), Kutkoside (KS), and Kutkin (KT) exerts the anti-invasion activity via down-regulation of the expression of matrix metalloproteinases (MMPs). MMPs play an important role in solid tumor invasion and migration.
Materials and methods
The activity and expression of gelatinases (MMP-2 and MMP-9) and collagenases (MMP-1 and MMP-13), protein, and mRNA were detected by gelatin zymography, and RT-PCR. The migratory and invasive capacities of MCF-7 cell lines were measured by the wound scratch migration assay. The preliminary cytotoxicity testing was done by MTT assay and propidium iodide staining. Further the inhibition of inflammatory mediators was also done by quantification of nitrite inflammatory mediators.
Results
The study showed that PE and its isolated iridoids glycosides PS, KS, and KT exhibited considerable cytotoxic potential in a dose-dependent manner. Further PE, PS, KS, and KT inhibited MCF-7 cell invasion and migration, and decreased MMP-2, 9 and MMP-1, 13 activities. Furthermore, PS, KS, and KT reduced MMPs expression at protein and mRNA levels, and suppression of the inflammatory mediators was also exhibited.
Conclusions
Our results suggest that PS, KS, and KT may be the valuable anti-invasive drug candidates for cancer therapy by suppressing Collagenases and Gelatinases. PS, KS, and KT showed good results in comparison with PE. PS and KS exhibit almost comparable down regulation while KT exhibited maximum suppression of invasion, migration, and expression of MMPs.
... In addition to the degradation of the extracellular matrix (ECM), MMPs are suspected of having other roles in cancer development, such as angiogenesis, inhibition of apoptosis [6], and regulation of cell proliferation [3]. High serum concentration and over-expression of MMP2, MMP3 and MMP9 in breast cancer patients is related to poor prognosis, decreased survival, increased tumor size, increased invasiveness and metastatic behavior [9,11,12,16,18,14,39,93,94,95]. Data from experimental animal models also correlates MMP expression with invasive behavior and increased metastasis [39]. ...
In order to study the expression of MMP2, MMP 3 and MMP9 in breast cancer brain and lung metastasis, we used a syngeneic rat model of distant metastasis of ENU1564, a carcinogen-induced mammary adenocarcinoma cell line. At six weeks post inoculation we observed development of micro-metastasis in the brain and lung. Immunohistochemistry and Western blotting analyses showed that MMP 2, -3 and -9 protein expression is consistently significantly higher in neoplastic brain tissue compared to normal brain tissue. Lung metastases express abundant MMP2, -3 and -9 in neoplastic cell cytoplasm. In situ zymography revealed gelatinase activity within the brain metastasis. Gel zymography showed an increase in MMP2 and MMP3 activity in brain metastasis. Furthermore, we were able to significantly decrease the development of breast cancer brain and lung metastasis in animals by treatment with PD 166793, a selective synthetic MMP inhibitor. In addition, PD 166793 decreased the in vitro invasive cell behavior of ENU1546. TIMP2 overexpression also decreased the development of breast cancer lung metastasis in our model. Our results suggest that MMP2, -3 and -9 may be involved in the process of metastasis of breast cancer to the brain and lung. Because astrocytes have been associated with breast cancer brain metastasis we evaluated the role of astrocytes and ERK2 pathway in MMP2 up-regulation in BC brain metastasis. A significant decrease in brain metastases development, and orthotopic tumor size and weight were observed in animals inoculated with ENU1564-TIMP2 cells. These were associated with decreased MMP2 activity, as demonstrated by gel zymography. Rat astrocyte-conditioned media increased expression of MMP2 in ENU15645 cells and increased in vitro cell invasion of ENU1564 and ENU1564-TIMP2 cells. Blockage of ERK1/2 phosphorylation by treatment with PD98059 decreased the expression of MMP2 in cancer cells grown in rat astrocyte-conditioned media. We determine that MMP2 plays a role in in vivo development of breast cancer brain metastases. Additionally, we conclude that astrocytes are associated with expression of MMP2 in cancer cells via ERK1/2 signaling pathway.
... As with prostate cancer, increased MMP production in breast cancer is generally associated with poor prognostic factors, with some exceptions. Numerous studies done in breast cancer reported increased total MMP2 or MMP9 protein expression to be associated with poor prognostic factors (60)(61)(62)(63)(64)(65)(66). However, two studies have reported evidence for an inverse association with poor prognosis: One study found lower levels of active MMP2 protein to confer a decreased recurrence-free survival in breast cancer patients and an inverse relationship between increasing total pro-MMP2 protein in breast cancer serum with nodal status, grade, and stage (67); the second reported a negative association between tumor grade and pro-MMP2 and pro-MMP9 protein expression in breast cancer patient serum (68). ...
Genome-wide association studies have accelerated the discovery of single nucleotide polymorphisms (SNP) associated with susceptibility to complex diseases, including many malignancies. The matrix metalloproteinase (MMP) family of proteases are involved in many cell processes, most notably the degradation of the extracellular matrix, and differences in gene and protein expression have been reported to be associated with many cancers. Surprisingly, none of the SNPs located within these genes have been identified to be associated with cancer in the genome-wide association studies published to date. This may be in part due to the proportion and the tagging efficiency of MMP SNPs covered by high-throughput genotyping chips. This review will provide an overview of current evidence for MMPs and associated SNPs in endometrial and other hormone-related cancers, to provide justification for the further detailed studies of MMP SNPs as cancer markers.
... MMPs are endopeptidases that degrade extracellular matrix (ECM) components (14). MMPs are known to be responsible for the detachment of tumor cells from the tumor cell mass and the invasion of the surrounding extracellular matrix, and MMP-2 expression is highly increased in a variety of cancer cell types (15)(16)(17). We assessed MMP-2 expression in several selected cancer cell lines. ...
Controlled gene expression in specific cells is a valuable tool for gene therapy. We attempted to determine whether the lentivirus-mediated Tet-On inducible system could be applied to cancer gene therapy. In order to select the genes that induce cancer cell death, we compared the ability of the known pro-apoptotreic genes, Bax and tBid, and a cell cycle inhibitor, p21cip1/waf1, and determined that Bax was the most effective. For the cancer cell-specific expression of rtTA2(S)-M2, we tested the matrix metalloproteinase-2 (MMP-2) promoter and determined that it is highly expressed in cancer cell lines, including SNU475 cells. The co-transduction of two lentiviruses that contain sequences for TRE-Bax and rtTA2(S)-M2, the expression of which is controlled by the MMP-2 promoter, resulted in the specific cell death of SNU475, whereas other cells with low MMP-2 expression did not evidence significant cell death. Our data indicate that the lentivirus-mediated Tet-On system using the cancer-specific promoter is applicable for cancer gene therapy.
... Depending upon the specificity of their target, metalloproteinases are cathegorised into four functional subgroups, including collagenases -MMP-1, -8, -13 and -18 (degrade fibrillary collagen), gelatinases -MMP-2 and -9 (degrade gelatins and elastins), stromyelysins -MMP-3, -10 and -11 (degrade proteoglycans, fibronectins and laminins) and membraneous type metalloproteinases -MMP-14, -15, -16 and -17 (12). High interest, particularly in oncology, is focused on the group of gelatinases, MMP-2 and MMP-9 in particular, the overexpression of which and correlation with prognostically unfavourable factors were demonstrated in female mammary carcinoma (3,(17)(18)(19)(20). The proteinases have been shown to be involved with the degradation of collagen type IV, which forms a groundwork of BM in vascular endothelium. ...
The purpose of the present study was to determine the expression of the proteins related to tumour metastatic potential, including matrix metalloproteinase (MMP)-9 and E-cadherin, in correlation with the expression of proliferation-associated antigen (Ki-67) in canine mammary adenocarcinomas.
Material for the studies was obtained during surgery from 35 dogs of various breeds, aged 7 to 16 years. Neoplastic tumours were verified by a pathologist. The studied proteins were detected by immunohistochemical reactions. The microphotographs of the studied tumours were subjected to computer-assisted image analysis using MultiScaneBase V 14.02 software.
Expression of MMP-9 was noted in almost 83% of the tumours, expression of E-cadherin in 77% of tumours, while expression of Ki-67 antigen was detected in fewer than 26% of studied tumours.
The positive correlation (r=0.375) between expressions of MMP-9 and Ki-67 and negative correlations between E-cadherin and Ki-67 (r=-0.383) as well as between MMP-9 and E-cadherin (r=-0.45) could suggest that expression and biological significance of the studied markers in mammary adenocarcinomas in dogs resembles the pattern noted in ductal carcinoma, i.e. in the most frequent histological type of malignant tumour in humans. This may point to suitability of the animal model in studies on mechanism of neoplasia and metastases in humans.
... Although MMP expression has been associated with invasive behavior of many human tumor types, conflicting results have been reported concerning the role of MMP-2 in breast cancer. While some studies found a highly significant correlation of MMP-2 expression with tumor grading and lymph node involvement (28,29), other studies found no or even an inverse correlation between MMP-2 expression and the clinical outcome of the disease (30). Thus, it seems that neither IMD nor MMP-2 expression alone can reliably predict the invasive and metastatic behavior of breast tumors. ...
The metastatic potential of tumors is dependent on the ability of tumor cells to degrade extracellular matrix components by the expression of matrix metalloproteinases (MMPs) and to induce vascularisation of the tumor tissue. Thus, expression of MMPs and the number of blood vessel in tumor tissue may serve as prognostic markers of aggressive and metastasizing tumor growth. We have determined the vascularisation and the expression of MMP-2 by immuno-histochemical staining of 19 benign and 75 malignant breast tissue specimens with CD31- and MMP-2 specific antisera. The degree of vascularisation was expressed by intratumoral microvascular density (IMD), which takes into account all vessels present in a hot spot irrespective of their size. In addition, we have introduced a novel parameter, vascular grading (VG), which describes the percentage of small microvessels of <20 microm in diameter in the total number of blood vessels. IMD tended to indicate an elevated risk for metastasis formation and disease recurrence, while VG did not correlate with metastasis formation. Similarly, MMP-2 expression neither correlated with the clinical outcome of the disease nor with the classical histo-pathological parameters such as stage, grade, lymph node involvement and estrogen receptor status. Tumor cell-specific MMP-2 expression, however, showed a highly significant correlation with VG but not with IMD. These results indicate that MMP-2 expression is rather involved in the formation of small capillaries than in vessel maturation and tumor cell invasion. Thus, MMP-2 expression by tumor cells may serve as indicator of strong angiogenic induction potential of breast tumor cells.
Ovarian cancer is one of the most common gynecologic malignancies. It is characterized by a high mortality rate, which is mainly due to the asymptomatic course of the disease. In light of the high mortality rate and increasing morbidity, new diagnostic methods are being explored to enable earlier detection, better monitoring, and improved prognosis. Such diagnostic methods include the assessment of tumor markers in various biological samples. Among the markers currently being investigated, extracellular matrix metalloproteinases (MMPs) are of particular interest. The objective of this article was to compile the existing knowledge of MMPs in ovarian cancer patients and to describe their potential diagnostic utility. Additionally, this article provides an overview of the symptoms, complications, and risk factors associated with ovarian cancer and the role of MMPs in physiology and pathology. Preliminary results indicate that tissue expression and blood and body fluid levels of MMPs may be different in ovarian cancer patients than in healthy women. The expression and concentration of individual MMPs have been shown to be correlated with cancer stage and disease severity. In addition, the preliminary value of some of these enzymes in predicting prognosis is discussed. However, as the amount of data is limited, more studies are needed to fully evaluate the potential function of individual MMPs in ovarian cancer patients. Based on the knowledge gathered for this article, it seems that MMP-2, MMP-3, MMP-7, MMP-8, MMP-9, MMP-13, are tentatively the most useful. A thorough evaluation of their utility as modern biomarkers in ovarian cancer requires further investigation.
Syndecan-1 (SDC-1) is an integral membrane heparin sulfate proteoglycan that is involved in inflammatory response, cell-signaling, cell proliferation, and numerous other cell-matrix interactions. Like the other members of the syndecan family, very little is known about structural conformations and dynamics of SDC-1. A majority of interactions occur through the extracellular ectodomain, therefore we have dedicated our research efforts to the study this specific portion of SDC-1. The ectodomain is often shed from the cell surface due to various stimuli. The released fragment has already been used as a useful biomarker for prognosis of some diseases and cancers. SDC-1 can be cleaved in different locations depending on the sheddase, generating soluble shed ectodomains that can be carried away in blood sera. In this study, we focus specifically on two main cleavage fragments that can be generated. We show the first successful expression and purification of recombinant SDC-1 ectodomains. Production of SDC-1 in E. coli allows the production of the core protein without risking heterogeneous post-translational modifications such as glycosylation, allowing a certain level of control over protein homogeneity that is not possible in mammalian expression. An expression vector was used to generate two different fusion proteins consisting of a His-tag and a TEV cleavage site for the removal of the fusion partner. SDS-PAGE was used to track the expression as well as the purification. Masses of the isolated proteins were determined using mass spectrometry and the purity and homogeneity were evaluated by solution NMR.
Owing to anticancer properties of selected natural substances, it is assumed that they have potential to be used in oncological therapy. Here, the recently proven effects of the selected natural polyphenols, resveratrol and curcumin, are described. Secondly, the potential of probiotics and prebiotics in modulation of immunological response and/or enhancing the chemotherapeutic treatments is reported based on the recent clinical trials. Further, the chapter presents current knowledge regarding the targeted supplementation of the patient with probiotic bacteria and known efficacy of probiotics to support immunotherapy. The major clinical trials are listed, aiming to verify whether, and to which extent the manipulation of patient’s microbiome can improve the outcome of chemotherapies. In the end, a potential of natural substances and feed ingredients to pose epigenetic changes is highlighted. The chapter provides an insight into the scientific proofs about natural bioactive substances in relation to cancer treatment, leaded by the question – do they really work?
Matrix metalloproteinase 2 (MMP-2) is a crucial biomarker of tumor growth, invasion and metastasis. In the present study, a core-satellite magnetic-fluorescent-plasmonic nanosensor ([email protected]) was constructed through biological self-assembly to generate localized SERS “hot spots” and an efficient FRET system for the sensitive determination of MMP-2 activity in a SERS-fluorescence dual-mode assay. In this hybrid nanosensor, a biotin-labeled peptide containing a specific MMP-2 substrate (PLGVR) was employed as a bridge for the assembly of gold nanoparticles (AuNPs) and avidin functionalized fluorescent-magnetic nanospheres (FMNS). The modified RB on FMNS served as a Raman reporter and a donor of FRET, while the AuNPs assembled on FMNS acted as SERS substrates and acceptors of FRET. In the presence of MMP-2, the SERS “hot spot” effect was weakened and the FRET system was disrupted through enzymatic cleavage of PLGVR, resulting in a reduction of SERS signal and the recovery of fluorescence emission. Importantly, this combination of SERS and fluorescence assay methods in the dual-mode nanosensor broadened the detection range for MMP-2 to 1–200 ng mL⁻¹, with a limit of detection of 0.35 ng mL⁻¹ and a limit of quantitation of 1.17 ng mL⁻¹. In addition, our novel nanosensor affords semi-quantitative sensing of MMP-2 by naked-eye observation and accurate detection of MMP-2 through dual-mode analysis. The practicality of [email protected] was validated by determination of MMP-2 activity in cell secretions and human serum samples. The designed [email protected] nanosensor holds great potential for clinical diagnosis of protease-related diseases.
TGFβ is a pleiotropic cytokine that plays critical roles to define cancer cell phenotypes, construct the tumor microenvironment, and suppress antitumor immune responses. As such, TGFβ is a lynchpin for integrating cancer cell intrinsic pathways and communication among host cells in the tumor and beyond that together affect responses to genotoxic, targeted, and immune therapy. Despite decades of preclinical and clinical studies, evidence of clinical benefit from targeting TGFβ in cancer remains elusive. Here, we review the mechanisms by which TGFβ acts to oppose successful cancer therapy, the reported prognostic and predictive value of TGFβ biomarkers, and the potential impact of inhibiting TGFβ in precision oncology. Paradoxically, the diverse mechanisms by which TGFβ impedes therapeutic response are a principal barrier to implementing TGFβ inhibitors because it is unclear which TGFβ mechanism is functional in which patient. Companion diagnostic tools and specific biomarkers of TGFβ targeted biology will be the key to exploiting TGFβ biology for patient benefit.
A simple, selective and sensitive “signal on” electrogenerated chemiluminescence (ECL) biosensing method was developed for matrix metalloproteinase 2 (MMP‐2). Ru(bpy)32+, gold nanoparticles (AuNPs) and Nafion were modified onto glassy carbon electrode (GCE) to form Ru(bpy)32+/AuNPs/Nafion/GCE as sensitive ECL platform and then ferrocene (Fc) labeled peptide was assembled onto the modified electrode to form ECL sensing platform. The ECL intensity increased when the ECL biosensing electrode reacted with MMP‐2 because of MMP‐2‐induced cleavage of Fc labeled peptide. The ECL method was applied to determine MMP‐2 with detection limit of 0.3 ng/mL and one‐step recognition, which is promising for point‐of‐care test of protease.
Enzyme-activatable ratiometric near-infrared (NIR) fluorescent probes enabling noninvasive imaging of enzyme activity in vivo are promising for biomedical research; however, such probes with ratiometric fluorescence emissions both in NIR window under a single NIR light excitation are largely unexplored. Here, a quenched NIR fluorophore of Cy5.5 is integrated with NIR fluorescent poly[2,6-(4,4-bis-(2-ethylhexyl)-4H-cyclopenta[2,1-b;3,4-b′]dithiophene)-alt-4,7(2,1,3-benzothiadiazole)] (PCPDTBT)-based semiconducting polymer nanoparticles (SPNs), and an αvβ3 integrin-targeting and matrix metalloproteinase-2 (MMP-2)-activatable ratiometric fluorescent probe (SPN-MMP-RGD) is developed. Under excitation at 660 nm, SPN-MMP-RGD shows “always-on” fluorescence of PCPDTBT (830 nm) and activatable fluorescence of Cy5.5 (690 nm) toward MMP-2, affording a remarkable ≈176-fold enhancement in fluorescence intensity ratio between 690 and 830 nm (I690/I830) for sensitive detection of MMP-2 activity in vitro and in tumor cells. By virtue of ratiometric fluorescence imaging independently of probe's concentration, SPN-MMP-RGD can not only accurately report on MMP-2 levels regarding different tumor sizes, but also noninvasively delineate MMP-2-positive tiny gastric tumors metastasis in vivo. The authors’ study reveals the potential of SPN-MMP-RGD for ratiometric fluorescence imaging of MMP-2 activity via combining two independent NIR fluorophores, which can be amenable for the design of other enzyme-activatable ratiometric NIR fluorescent probes for reliable in vivo imaging.
Herein, a label-free and homogeneous electrochemical strategy for monitoring of matrix metalloproteinase 2 (MMP-2) activity was proposed based on electrodes modified with orderly distributed mesoporous silica films (MSFs). In the absence of target MMP-2, an artificially substrate peptide with positive charge was absorbed on the surface of MSFs by electrostatic interaction, which could prevent electrochemical molecules [Ru(NH3)6]Cl3 from approaching the electrode surface. When the substrate peptide was hydrolyzed by target MMP-2, [Ru(NH3)6]Cl3 could arrive to the electrode surface and lead to the increase of electrochemical signal. This assay showed considerable sensitivity to target MMP-2, which could measure it down to 0.98 ng. mL⁻¹. Meanwhile, a satisfied response to the inhibitor of MMP-2 was also achieved (IC-50 value = 1.68 μM). Significantly, it displayed satisfactory performances in the complicated biological samples including cell lysates and human serum. Taking advantages of the anti-fouling ability in biological complex samples of MSFs and the high efficiency of homogeneous sensing, this assay realized the electrochemical detection of MMP-2 with accuracy and sensitivity, which exhibited significant potential in clinical biomedicine and biological analysis of cancer-related protease.
The efficient signal amplification capacity of several class 2 CRISPR-Cas systems with trans-cleavage activity has exhibited great value in molecular diagnostics, but its potential application for non-nucleic-acid targets is yet underdeveloped. Here, we deploy CRISPR-Cas system for the ultrasensitive detection of protease biomarkers by the coupling of proteolysis-triggered transcription. In this strategy, a protease-activatable RNA polymerase is adopted for the conversion of each protease-catalyzed proteolysis event into the output of multiple programable RNA sequences by in vitro transcription, and the transcribed RNA subsequently serves as the guide RNA of Cas12a proteins with trans-cleavage activity. The rational design of the transcribed RNA efficiently couples the signal conversion and amplification of proteolysis-triggered transcription and the self-signal amplification of CRISPR-Cas12a, resulting in a two-stage amplified detection of target protease. The versatility of this strategy has been demonstrated in the detection of protease biomarkers including MMP-2 and thrombin with femtomolar sensitivity, which is 5–6 orders of magnitude lower than that of the standard peptide-based methods. Moreover, the proposed method has been further applied in the analysis of MMP-2 secreted by different cancer cell lines as well the assessment of MMP-2 activity in clinical serum samples, providing a generic method for the ultrasensitive detection of protease biomarkers in biochemical research and clinical diagnosis.
The clinical medicine and biomaterials are two fields that are conducive to achieve the goal of precise medicine on diagnostics and therapeutics of various diseases. The interdiscipline of materials and medicine, termed/abbreviated as “materdicine,” seeks to address the dominant medical shortcomings and challenges faced by conventional medicine, including inferior bioavailability, systemic toxicity, poor targeting specificity, and unsatisfied diagnostic/therapeutic efficacy. In this review, we present the perspective and discussion on the use of diverse biomaterials for disease diagnosis and treatment from a broad perspective, especially on nanoscale biomaterials. We initially highlight the recent advances on the engineering of abundant contrast agents for diagnostic bioimaging, such as optical fluorescence imaging, magnetic resonance imaging, and photoacoustic imaging. In addition, discussions on the diagnostic biosensing for point‐of‐care diagnosis, such as fluorescent and plasmonic sensors, are supplemented. Furthermore, we outline several materdicine‐enabled therapeutic modalities for disease treatments, including the following exemplified scaffold biomaterials for regenerative medicine. Especially, rational modulation of toxicity issues of micro‐/nanoscale biomaterials is also accounted for addressing the possible concerns of biocompatibility and biosafety on further clinical translation of materdicine. Finally, we summarize facing challenges and outlook future developments relating to the clinical translation of these distinctive biomaterials in materdicine. This review proposes and highlights the perspective of materdicine, providing a deep discussion on the rational design, construction, and application of versatile biomaterials/medmaterials with unique physiochemical property and specific biological effects for disease diagnosis and treatment from a broad perspective, especially on nanoscale biomaterials.
The 72-kDa type IV collagenase or gelatinase A is the second member of the matrix metalloproteinase family, MMP-2. Since the discovery of its first two substrates within components of the extracellular matrix, denatured interstitial type I collagen and native type IV collagen, the roles and various levels of regulation of MMP-2 have been intensively studied, mainly in vitro. Its (over)expression in most if not all tumors was considered a hallmark of cancer aggressiveness and boosted investigations aiming at its inhibition. Unfortunately, the enthusiasm subsided like a soufflé after clinical trial failures, mostly because of insufficient knowledge of in vivo MMP-2 activities and detrimental side effects of broad-spectrum MMP inhibition.
Nowadays, MMP-2 remains a major topic of interest in research, the second in the MMP family after MMP-9. This review presents a broad overview of the major features of this protease. This knowledge is crucial to identify diagnostic or therapeutic strategies focusing on MMP-2. In this sense, recent publications and clinical trials underline the potential value of measuring circulating or tissular MMP-2 levels as diagnostic or prognostic tools, or as a useful secondary outcome for therapies against other primary targets. Direct MMP-2 inhibition has benefited from substantial progress in the design of more specific inhibitors but their in vivo application remains challenging but certainly worth the efforts it receives.
Matrix metalloproteinases-2 (MMP-2) plays an important role in tumorigenesis and metastasis of gastric cancer. Therefore, specific and sensitive detection of MMP-2 is of great significance for clinical diagnosis and therapeutic evaluation of gastric cancer. Herein, we developed a new approach for accurate detection of MMP-2 activity both in gastric cancer cells and clinical tissues using a fluorescent probe Dab-PLGVRGY-FITC. The capability of probe for detecting MMP-2 was systematically evaluated in gastric cancer MGC-803 cells and clinical gastric cancer tissues. Around 40-fold fluorescence enhancement at the wavelength of 520 nm was determined for the assays of probe with recombinant MMP-2. The detection limit of MMP-2 was as low as 42 ng/mL. Strong fluorescent signals were detected in MGC-803 cells and mice gastric cancer tissues. Fluorescent signal enhancements of 11 clinical gastric cancer tissue lysates were 5.8～9 times (7.53 ± 0.77 on average) higher than those from gastritis patients. This probe could be potentially used for early diagnosis of gastric cancer in clinics.
The abnormal expression of tumor-related proteases plays a critical role in cancer invasion, progression, and metastasis. Therefore, it is considerably meaningful to noninvasively assess the proteases’ activity in vivo for both tumor diagnosis and therapeutic evaluation. Herein, we report an activatable probe constructed with a near-infrared dye (Cy5.5) and a quencher (QSY21) covalently linked through a peptide substrate of matrix metalloproteinases-2 (MMP-2) that was chosen a model for tumor-associated proteases. Upon cleavage with activated MMP-2, the above probe emitted an MMP-2 concentration dependent fluorescence. Quite unexpectedly, owing to the variation in the aggregation state of both dye and its quencher in consequence of the cleavage, the responsive probe presented a dramatic MMP-2 concentration dependent absorption at around 680 nm, while that at around 730 nm was MMP-2 concentration independent. These features allowed detection of MMP-2 activity via both fluorescence and photoacoustic (PA) imaging in vitro, respectively. Moreover, taking the PA signal at 730 nm as internal reference, the PA signal at 680 nm allowed quantitative detections of MMP-2 expression in breast cancer in vivo. We thus envision that our current approach would offer a useful tool for studying the malignant impacts of versatile tumor-associated proteases in vivo.
Matrix Metalloproteinase-2 is a family of extra cellular matrix degrading proteinases. Study results on the role of MMP-2 in breastcancer progression and metastasis are still controversial. The aim of this study was to know the MMP-2 level by analyzing in metastaticand non metastatic breast cancer patients by using a cross sectional design, it was carried out in the Wahidin Sudirohusodo, Ibnu Sina,Labuang Baji Hospitals and at the Research Unit of UNHAS Makassar from April 2012 until June 2012. The 56 breast cancer patientswere divided into two groups: metastatic and non metastatic based on their thorax photo, ultrasonography and/or bone scan results. Theywere also divided into early and advanced stage, based on their TNM staging. The matrix metalloproteinase-2 level was determined byELISA method. The result of this study showed that the mean levels of MMP-2 in metastatic and non metastatic breast cancer were 20.18ng/mL and 17.14 ng/mL, respectively. The independent sample T test showed there was a significant difference (p=0.018) in MMP-2levels between metastatic and non metastatic breast cancer. The mean level of MMP-2 in early and advanced stages was 17.10 ng/mLand 18.31 ng/mL, respectively, the independent sample T test showed no significant difference of MMP-2 level between both stages. Oneway ANOVA test showed no significant difference of MMP-2 level based on tumour size and regional lymph node infiltration. The MMP-2contributed to the metastasis in breast cancer patients. The matrix metalloproteinase-2 level in metastatic was higher than in the nonmetastatic breast cancer, so this condition could be used as a biomarker to predict the metastasis of breast cancer.
Here, we propose a highly sensitive and rapid bio-sensor for the detection of bio-markers for stroke and cancer-related diseases, based on utilizing the adsorption properties of ruthenium carbonyl (Ru-CO) clusters onto monolayer graphene (MG). A fast rate of decarbonylation of Ru-CO to form ruthenium oxide nanoparticles (RuO2 NPs) on MG was observed. Quantitative detection of matrix metalloproteinase-2 (MMP-2) enzyme (bio-marker for stroke and vascular diseases) was demonstrated by tracking the spectral shift of the characteristic G band of graphene caused by the adsorption of RuO2 NPs. A concentration of as low as 17 ng/ml of MMP-2 was detected in a simulated clinical serum sample. This effective bio-sensor has the potential to revolutionize the biomedical field in the early detection and possible prevention of stroke disease and cancer diagnosis.
Introduction:
The overexpression of matrix metalloproteinase (MMP) plays an important role in the context of tumor invasion and metastasis, and MMP-2 has been characterized as the most validated target for cancer. Therefore, it is necessary to design matrix metalloproteinase inhibitors (MMPIs) that would be active and selective against MMP-2 but non-selective toward other MMPs. Areas covered: This article clearly describes the structural character of MMP-2 followed by a review of the recent development of selective MMP-2 inhibitors based on their basic structures. Expert opinion: Over the past 30 years, MMPs have been considered to be attractive cancer targets, and several different types of synthetic inhibitors have been identified as anticancer agents, but only a small number of small MMPIs have been examined in clinical trials, and none of these molecules has been established as anticancer drugs due to their adverse effects. One major possibility is that the MMPIs used in clinical trials were broad-spectrum drugs that also inhibited the anti-tumor effects and influenced the mediation of the normal physiological processes of MMPs. MMP-2 has recently been characterized as the most validated target for cancer. Therefore, the design and synthesis of selective MMP-2 inhibitors would be helpful for the treatment of cancer.
Highly selective bioassays are extremely important in biological and chemical sensing. Herein, a selective biosensor incorporating sensitive electrogenerated chemiluminescence (ECL) is reported for simultaneous detection of two matrix metalloproteinases (MMPs) that are related to pathological development and progression of cancer. An Ir complex ((dfppy)2Ir(dcbpy)PF6, Ir1) and a Ru complex ((Ru(bpy)2(mcbpy-O-Su-ester)(PF6)2, Ru1) were employed as potential resolved ECL labels. It was found that the Ru1 and Ir1 in the presence of tripropylamine could produce two well-separated strong ECL emissions at +0.9 V and +1.4 V vs Ag/AgCl, respectively, with a large difference of ECL peak potential (∼0.5 V) at the gold electrode, which provided an access for potential-resolved detection of dual targets. A multiplexed biosensor was simply fabricated by co-immobilizing the Ir1 and Ru1 labeled peptides on a gold electrode via self-assembling technique. It was facile to determine MMP-2 and MMP-7 in a single run with detection limit as low as 5 ng/mL and 10 pg/mL, respectively. The advantages of the multiplexed ECL biosensor were further exemplified by analyzing two MMPs released from living HeLa, K562, HCT116 and macrophage cells. The multiplexity of the biosensor will open a new door for facile and rapid detection of multiple biomarkers.
A simple and sensitive electrogenerated chemiluminescence biosensor was developed to monitor matrix metalloproteinase 2 (MMP-2) by employing a specific peptide (CGPLGVRGK) as a molecular recognition substrate. Bis(2,2′-bipyridine)-4′-methyl-4-carboxybipyridine-ruthenium N-succinimidyl ester-bis(hexafluorophosphate) (Ru(bpy)2(mcbpy-O-Su-ester)(PF6)2 (Ru1) was used as ECL-emitting species and covalently labeled onto the peptide through NH2-containing lysine on the peptide via acylation reaction to form Ru1-peptide as an ECL probe. An ECL peptide-based biosensor was fabricated by self-assembling the ECL probe onto the surface of gold electrode. MMP-2 can specifically cleave the Ru1-peptide on the electrode surface, which led the partly Ru1-peptide to leave the electrode surface and resulted in the decrease of the ECL intensity obtained from the resulted electrode in 0.1 M phosphate-buffered saline (pH 7.4) containing tri-n-propylamine. The decreased ECL intensity was piecewise linear to the concentration of MMP-2 in the range from 1 to 500 ng/mL. Moreover, the ECL biosensor is successfully applied to detection of MMP-2 secreted by living cell, such as HeLa cells. Additionally, the biosensor was also applied to the evaluation of matrix metalloproteinase inhibitors. The strategy presented here is promising for other disease-related matrix metalloproteinase assay and matrix metalloproteinase inhibitor profiling with sensitivity and simplicity.
Graphical Abstract
Detection of MMP-2 released from living cells by ECL peptide-based biosensor
Matrix metalloproteinase 2 (MMP2) is an enzyme with important functions in breast cancer invasion and metastasis. However, it is unclear whether circulating MMP2 levels may predict breast cancer risk. We conducted a prospective nested case-control analysis in the Nurses' Health Study among 1136 cases who were diagnosed with invasive breast cancer between 1992 and 2004 and 1136 matched controls. All participants provided blood samples in 1989-1990, and a subset (170 cases, 170 controls) contributed an additional sample in 2000-2002. Pre-diagnostic plasma MMP2 levels were measured via immunoassay, and conditional logistic regression was performed to calculate odds ratios (ORs) and 95% confidence intervals (95% CIs), adjusted for breast cancer risk factors. No association was observed between plasma MMP2 levels and risk of total invasive breast cancer (top vs. bottom quartile, OR=1.0; 95% CI: 0.7, 1.2; p-trend=0.89). Findings did not vary significantly by time since blood draw, body mass index, postmenopausal hormone use, or menopausal status at either blood draw or breast cancer diagnosis. MMP2 was associated with a greater risk of nodal metastases at diagnosis (top vs. bottom quartile, OR=1.5; 95% CI: 1.0, 2.2; p-heterogeneity, any vs. no lymph nodes=0.002), but no significant associations were observed with other tumor characteristics or with recurrent or fatal cancers. Plasma MMP2 levels do not appear to be predictive of total invasive breast cancer risk, although associations with aggressive disease warrant further study.
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A novel fluorescence nanoprobe for the detection of matrix metalloproteinase 2 (MMP2) has been developed by engineering the fluorescein isothiocyanate-labeled peptide onto the surface of poly(m-phenylenediamine) (PMPD) nanoparticles through covalent linkage. The nanoprobe itself displays a low background signal due to the effective fluorescence quenching by electron-rich PMPD, but its reaction with MMP2 causes 11-fold fluorescence enhancement. Compared with similar fluorescence nanosystems for MMP2 assembled through physical adsorption, the as-prepared nanoprobe is significantly more stable and displays a strikingly higher signal-to-background ratio, which leads to a high sensitivity for MMP2 assay, with a detection limit of 32 pM. Most notably, the nanoprobe has been successfully applied to determine MMP2 in human serum samples, demonstrating that the MMP2 level in serum from colorectal cancer (CRC) patients is 2 times higher than that from healthy people. Moreover, the nanoprobe has also been used to monitor MMP2 secreted by CRC cells that were grown under normoxic and hypoxic conditions, respectively, and the results show that the cells under hypoxic conditions produce higher level of MMP2 than those under normoxic conditions. Our method is simple, and can offer a highly sensitive detection of MMP2 in relevant clinical samples.
Breast cancer is the most common malignant cancer among women, both in Poland and worldwide. Due to the constantly increasing number of breast cancer cases, it is vital to develop effective activities in primary and secondary prevention. One of the promising methods of best value, connecting both types of cancer prevention, appears to be chemoprevention. Chemoprevention uses natural or synthetic compounds to inhibit, delay or reverse the process of carcinogenesis. Among ingredients of natural origin, great attention is paid to curcumin - a broad-spectrum anti-cancer polyphenol derivative, extracted from the rhizome of Curcuma longa L. Curcumin has a number of chemopreventive properties such as anti-inflammatory activity, induction of apoptosis, inhibition of angiogenesis as well as tumor metastasis. Numerous in vitro and in vivo studies have demonstrated the mentioned anti-cancer effect in the epithelial breast cell line MCF-10A and in the epithelial breast cell lines MCF-7, BT-474, SK-BR-3-hr and MDA-MB-231. The main problem associated with the use of curcumin as a chemopreventive agent in humans is its low absorption from the gastrointestinal tract, poor solubility in body fluids and low bioavailability. Current studies are underway to increase the bioavailability and effectiveness of curcumin in vivo. Good results in the prevention and the treatment of breast cancer could be ensured by curcumin nanoparticles coated with albumin, known as nanocurcumin. The studies using nanocurcumin, however, are still in the preclinical stage, which is why there is a need to conduct extensive long-term randomized clinical trials to determine its effectiveness.
This paper analyses data of 150 female patients undergoing surgical treatment for invasive ductal breast cancer at the University Hospital for Tumors from January 2006 to January 2007. The control group consisted of 50 healthy women. The patients were classified into three groups, depending on their tumor differentiation, i.e. grade I, II and III tumor groups. Each group consisted of 50 patients. Traditional prognostic factors including: age, tumor size and differentiation grade, axillary lymph node status, presence of distant metastases, steroid receptor findings, vascular invasion of the primary tumor, presence of an extensive intraductal component (EIC) in the primary tumor, HER-2 protein expression were evaluated. Both the patients' and controls' serum levels of proMMP-2 (pro-matrix metalloproteinase-2) were assessed using the ELISA method. The aim of the study was to assess pathohistological prognostic factors and the level of serum proMMP-2 in the three patient groups and the controls, compare the relationship between the prognostic factors and the level of serum proMMP-2 in the patient groups, and upon the results, determine possible features of proMMP-2 as a prognostic factor in breast cancer patients. The study results showed no difference in proMMP-2 concentrations between the three patient groups and the controls. No statistically significant difference in the serum proMMP-2 concentration was found between the patient groups, although the grade III group values were the highest showing a trend toward statistical significance. Comparison of proMMP-2 and prognostic factors revealed a statistically significant correlation between proMMP-2 and age in patients with histologic grade I tumors. There was no statistically significant correlation between circulating proMMP-2 and other pathohistological prognostic factors.
Elevated levels of matrix metalloproteinases (MMPs) have been found to associate with poor prognosis in various carcinomas. This study aimed at evaluating plasma levels of the collagenases MMP1, MMP8 and MMP13 as diagnostic and prognostic markers of breast cancer. Using ELISA, plasma levels of MMP1, MMP8 and MMP13 were measured in 42 control individuals and in 208 patients – of which 21 were inflammatory breast cancer patients – and were correlated with standard clinicopathological data. Plasma MMP1 levels were higher in breast cancer patients than in control individuals, while the opposite was true for MMP8. Plasma MMP13 levels could not be detected. We found a negative correlation of plasma MMP1 with tumour size (P = 0.07); and a positive association of MMP8 with the premenopausal status (P = 0.06), Nottingham Prognostic Index (P = 0.06) and Her2 expression (P = 0.07). Further, a twofold decrease in MMP1 (P = 0.025) and MMP8 (P = 0.007) levels was observed in inflammatory breast cancer patients, a very rare and not well understood aggressive disease. Most interestingly, we observed a peculiar relation between plasma MMP8 levels and lymph node metastasis. We found that both control individuals and patients without lymph node involvement (pN0) have lower plasma MMP8 levels than patients with moderate lymph node involvement (pN1, pN2) (P = 0.001); and that they show a trend for higher MMP8 levels as compared with patients with extensive lymph node metastasis (pN3) and a strong predisposition to distant metastasis. In summary, we observed differences in MMP1 and MMP8 plasma levels between distinct breast cancer patient groups. As it is not clear to date whether MMPs in blood and body fluids have a physiological function per se, we hypothesize that altered levels in blood reflect local changes in the extracellular microenvironment. As such, a positive association of blood MMP levels with clinical characteristics and tumour features reflects a negative association with tissue MMP levels and vice versa. Therefore, our results suggest that both MMP1 and MMP8 in the tumour may contribute to the aggressive phenotype of inflammatory breast carcinomas. Interestingly, our results suggest that tumour MMP8 expression may affect the metastatic behaviour of breast cancer cells with a greater protective effect against lymph node metastasis than against distant metastasis.
At Windber Research Institute we have started research programs that use artificial neural networks (ANNs) in the study of
breast cancer in order to identify heterogeneous data predictors of patient disease stages. As an initial effort, we have
chosen matrix metalloproteinases (MMPs) as potential biomarker predictors. MMPs have been implicated in the early and late
stage development of breast cancer. However, it is unclear whether these proteinshold predictive powerfor breast disease
diagnosis, and we are not aware of any exploratory modeling effortsthat address the question. Here we report the development
of ANN models employing plasma levels of these proteins for breast disease predictions.
Clinical proteomic profiling by mass spectrometry (MS) aims at uncovering specific alterations within mass profiles of clinical specimens that are of diagnostic value for the detection and classification of various diseases including cancer. However, despite substantial progress in the field, the clinical proteomic profiling approaches have not matured into routine diagnostic applications so far. Their limitations are mainly related to high-abundance proteins and their complex processing by a multitude of endogenous proteases thus making rigorous standardization difficult. MS is biased towards the detection of low-molecular-weight peptides. Specifically, in serum specimens, the particular fragments of proteolytically degraded proteins are amenable to MS analysis. Proteases are known to be involved in tumour progression and tumour-specific proteases are released into the blood stream presumably as a result of invasive progression and metastasis. Thus, the determination of protease activity in clinical specimens from patients with malignant disease can offer diagnostic and also therapeutic options. The identification of specific substrates for tumour proteases in complex biological samples is challenging, but proteomic screens for proteases/substrate interactions are currently experiencing impressive progress. Such proteomic screens include peptide-based libraries, differential isotope labelling in combination with MS, quantitative degradomic analysis of proteolytically generated neo-N-termini, monitoring the degradation of exogenous reporter peptides with MS, and activity-based protein profiling. In the present article, we summarize and discuss the current status of proteomic techniques to identify tumour-specific protease-substrate interactions for functional protease profiling. Thereby, we focus on the potential diagnostic use of the respective approaches.
Matrix metalloproteinases (MMPs), particularly MMP-2 and MMP-9, have been reported as putative tumor markers because of their involvement in cancer invasion and metastasis. The aim of our study was to elucidate the possible role of MMP-2 and -9 as serum prognostic biomarker for breast cancer classification and correlate it with the clinicopathological variables.
Our study consisted of 60 females with primary breast cancer, 40 cases of benign breast disease and 60 healthy female volunteers as controls. The serum MMP-2 and -9 levels were quantitatively measured by ELISA technique.
A significantly raised MMP-2 and MMP-9 levels were observed in breast cancer patients. Significant rise in serum MMP-9 concentration was found in patients presenting with metastasis as well as in those cases who presented with a duration of less than 1year. ROC analyses depicted a serum cutoff value of 315ng/mL for MMP-9 to discriminate the breast cancer patients from the control group.
Our results suggest that serum MMP-9 level is a better marker than serum MMP-2 in predicting the breast cancer development and progression.
Borstkanker is nog steeds de meest voorkomende dodelijke vorm van kanker bij Europese vrouwen. Dankzij intensieve screening de ontwikkeling van efficiëntere systemische therapieën is de mortaliteit in Europa gedaald. De meerderheid van de sterfgevallen is te wijten aan de verspreiding van tumorcellen in het lichaam, ook wel metastasering genoemd. Tot op heden wordt de prognose van een patiënt bepaald op basis van de tumor grootte, differentiatiegraad, oestrogeen en progesteron receptor expressie, Her2 status en aanwezigheid van lymfeklier aantasting. Hoewel deze factoren allemaal een sterke prognostische waarde hebben, worden op basis van deze parameters niet alle patiënten met een verhoogd risico op metastasering geïdentificeerd. Bijgevolg is er een grote nood aan nieuwe prognostische merkers of biomerkers voor borstkanker. Hierbij wordt meer en meer aandacht besteedt aan de identificatie en evaluatie van circulerende biomerkers aangezien deze het mogelijk maken om op een niet-invasieve manier het ziekteverloop en de respons op therapie op te volgen. Proteasen, waaronder de matrix metalloproteinasen (MMP) en cathepsines, worden als goede kandidaat biomerkers beschouwd door de diverse functies die ze uitoefenen tijdens tumor groei en metastasering. In deze thesis werd hun klinische relevantie in borstkanker nagegaan.
In een eerste luik werd het expressieprofiel van 6 MMPs in luminaal A type invasief ductale borsttumoren bestudeerd en in relatie gebracht met de standaard prognostische factoren voor borstkanker. Postmenopausale patiënten vertoonden een sterke tumorale expressie van MMP2 en MMP11 mRNA, wat in overeenstemming is met hun sex steroïd hormoon-afhankelijke regulatie. De verhoging in MMP expressie in luminaal A type borsttumoren van postmenopausale vrouwen is op het eerste zicht paradoxaal. Luminaal A type tumoren komen namelijk vooral voor bij postmenopausale vrouwen waar ze geassocieerd zijn met een goede prognose, terwijl een verhoogde MMP expressie geassocieerd is met een slechte prognose in diverse tumortypes. Deze schijnbare paradoxale bevinding zou er echter op kunnen wijzen dat luminaal A borsttumoren van premenopausale vrouwen een ander, meer agressief, biologisch gedrag vertonen dan deze van postmenopausale vrouwen.
In een tweede luik werd de prognostische waarde van circulerend MMP2 en MMP9 in het plasma van borstkanker patiënten geëvalueerd, waarbij een onderscheid gemaakt werd tussen circulerend actief MMP en totaal MMP. Patiënten met een Her2-positieve tumor vertoonden een verhoogde activiteit van MMP2 in hun bloed, wat voorgaande bevindingen van in vitro werk en dierstudies bevestigt. Gebruik makend van humane stalen, hebben wij kunnen bevestigen dat er een sterke interactie bestaat tussen de signaaltransductieweg van Her2 en deze van de MMPs.
In een derde luik werd onderzocht of bepaling van MMP1, MMP8 en MMP13 expressie in plasma kan bijdragen tot de diagnostiek en prognosestelling van borstkanker. Bepaling van plasma MMP1 en MMP8 expressie maakte het mogelijk om borstkanker patiënten, onder wie inflammatoire borstkanker patiënten, te onderscheiden van gezonde vrijwilligers. Bovendien konden er verschillen in MMP1 en MMP8 expressie aangetoond worden tussen patiënten met verschillende klinische en pathologische kenmerken. Verder vonden we dat de MMP8 concentratie hoger is in patiënten met een beperkte lymfeklier aantasting (pN1, pN2) dan in gezonde controlepersonen, terwijl het omgekeerde geldt voor patiënten met een uitgebreide lymfeklier aantasting (pN3) en een verhoogde kans op het ontwikkelen van metastases. Op basis van de hypothese dat er een omgekeerde relatie bestaat tussen proteïne gehaltes in het bloed en deze in het tumorweefsel, met een hogere expressie in het bloed wanneer het betreffende proteïne geconcentreerd wordt in de tumor of het stroma, bevestigen onze resultaten de bevindingen van vorige studies dat MMP8 een beduidend beschermend effect heeft op lymfeklier metastasering en in mindere mate op metastasering naar andere organen zoals de longen.
In een vierde luik werd de expressie van cathepsine B, H en X alsook van hun endogene inhibitor cystatin C in serum van vrouwen met vroegstadium of inflammatoire borstkanker bepaald. Doordat chemotherapie verschillende ongunstige bijwerkingen veroorzaakt is er een groot debat gaande of patiënten met gunstige prognostische kenmerken zoals een tumor grootte kleiner dan 5 cm en de afwezigheid van lymfeklier aantasting, adjuvant behandeld moeten worden met chemotherapie. Inflammatoire borstkanker is een zeldzame, zeer agressieve vorm van borstkanker waarvan tot op heden de onderliggende pathologie ongekend is. Aangezien cathepsines een duidelijke rol spelen in kanker, zou de bepaling van hun serum concentraties extra prognostische informatie kunnen verschaffen. Lagere concentraties serum cathepsine B en hogere hoeveelheden cystatine C werden teruggevonden in respectievelijk patiënten met een slecht gedifferentieerde tumor en postmenopausale patiënten. Op basis van serum cystatine C expressie konden vroegstadium borsttumoren ingedeeld worden in 2 groepen zijnde tumoren kleiner dan 2 cm enerzijds en groter dan 2 cm anderzijds, waarbij de laatste groep gekenmerkt werd door een verhoogde cystatine C expressie. Deze verhoging zou het gevolg kunnen zijn van een compensatie mechanisme, hoewel blijkbaar inefficiënt, voor een verhoogde cathepsine activiteit in de tumor. Verder hadden patiënten met inflammatoire borstkanker een opmerkelijk verschillende hoeveelheid serum cathepsine H en X en in minder mate cathepsine B en cystatine C expressie in vergelijking met niet-inflammatoire borstkanker patiënten. Deze observatie suggereert dat cathepsine H en X mogelijks betrokken zijn bij de onderliggende processen die deze zeer agressieve vorm van borstkanker veroorzaken.
In een vijfde luik werd de invloed van genetische variatie, meer specifiek ‘single’ nucleotide polymorfismen (SNP), in het MMP8 gen op de expressie en de paradoxaal beschermende rol van MMP8 op metastasering nagegaan. Dragers van het minst voorkomend allel (minor allele) van de rs11225395 SNP, gelegen in de promoter regio van MMP8, vertoonden een verlaagd risico op lymfeklier aantasting. In vitro functie-analyses toonden aan dat de MMP8 promoter met het ‘minor allele’ een hogere activiteit heeft dan deze met het meest voorkomend allel (major allele). Verder toonden we aan dat de promotersequentie met het ‘minor allele’ gebonden kan worden door diverse nucleaire proteïnen wat de verhoogde activiteit van de promoter zou kunnen verklaren. Bovendien vonden we dat dragers van het ‘minor allele’ minder frequent metastases ontwikkelden en een opmerkelijk betere algemene overleving en ziektevrije-overleving hadden dan dragers van het ‘major allele’, en dit specifiek in de groep van patiënten met vroegstadium borstkanker. Ten eerste bevestigen deze resultaten dat MMP8 genvariatie een invloed kan hebben op de expressie en functie van MMP8. Ten tweede wijzen deze data erop dat, in analogie met wat reeds aangehaald werd voor hoofdstuk 5, MMP8 een beschermende rol tegen lymfeklier aantasting heeft.
Breast cancer remains the most common deadly form of cancer in European women. Thanks to early diagnosis and administration of adjuvant systemic therapy, its mortality rate is dropping in the Western world. The majority of treatment failures and deaths are attributed to metastasis or spread of tumour cells to distant organs. Currently, the established prognostic factors for operable breast cancer are tumour size, histologic grade, steroid hormone receptors, Her2 expression and lymph node involvement. In spite of their strong association with recurrence-free and overall survival, these parameters do not allow us to identify all patients at high risk of metastasis. Therefore, a great deal of research is devoted to the identification of new prognostic markers or biomarkers for breast cancer. In particular, circulating biomarkers have been the focus of interest since they are readily accessible and permit serial assessment for monitoring of therapy response and disease progression. Proteases such as matrix metalloproteinases (MMPs) and cathepsins have been suggested as candidate biomarkers of cancer due to their numerous functions in tumour progression and metastasis. Hence, we evaluated their clinical relevance in breast cancer.
In a first study, we explored the mRNA expression profile of a panel of 6 MMPs in luminal A invasive ductal carcinomas and correlated it with clinical and pathological prognostic markers. We observed a strong positive correlation between tumour MMP2 and MMP11 expression and the postmenopausal status which is in line with a MMP sex steroid dependent regulation. At first sight, an increase in MMP expression in luminal A breast tumours of postmenopausal women seems paradoxical. First, luminal A tumours are primarily found in postmenopausal women being associated with a good prognosis. Second, increased MMP expression has been associated with a worse prognosis in various tumour types. However, this apparent paradoxical finding might suggest that luminal A breast tumours of premenopausal women have a more aggressive biological behaviour than those of postmenopausal women.
In a second study, we evaluated the prognostic value of plasma MMP2 and MMP9 in breast cancer, discriminating the expression of the active MMP fraction from the total expression. We demonstrated a significant increase in MMP2 activity in plasma from patients with a Her2 positive tumour. Hence, we confirmed the link between the Her2 and MMP pathway in human samples, a relation that was previously solely observed in in vitro systems and in animal models.
In a third study, we investigated whether measurement of circulating levels of the collagenases MMP1, MMP8 and MMP13 may be a useful tool for breast cancer diagnosis and prognosis. Plasma MMP1 and MMP8 levels enabled the diagnosis of breast cancer, including inflammatory breast cancer. We found differences in MMP1 and MMP8 levels between patients with different clinical and pathological characteristics. Furthermore, MMP8 levels were higher in patients with moderate lymph node involvement (pN1, pN2) than in healthy controls, while the opposite was true for patients with an extended lymph node metastasis (pN3) and a strong predisposition to distant metastasis. Based on the hypothesis of a reverse association of blood and tissue protein levels, with lower blood levels in case of concentration in the tumour and/or stroma, our observations support previous studies that MMP8 might influence the metastatic behaviour of breast cancer cells. Moreover, our results suggest a greater protective effect against lymph node metastasis than against distant metastasis.
In a fourth study, we examined the expression of cathepsins B, H, X and their endogenous inhibitor cystatin C in serum of early stage and inflammatory breast cancer patients. As chemotherapy is known to have adverse side effects, the debate is ongoing whether patients with favourable prognostic characteristics, i.e. tumours smaller than 5 cm and without lymph node involvement, should receive adjuvant chemotherapy. On the other hand, inflammatory breast cancer is as yet a rare but very aggressive disease with unknown underlying pathology. Since cathepsins have been shown to play an important role in cancer, their serum levels might provide additional prognostic information. We found a correlation between lower serum cathepsin B levels and poor histologic grade, and observed higher cystatin C levels in postmenopausal women. Serum cystatin C levels indicated that early stage breast carcinomas can be classified into 2 groups: tumours smaller than 2cm and those of 2cm or larger; the latter being associated with higher cystatin C levels. The increase in cystatin C levels may result from a compensation mechanism, although an apparently inefficient one, for elevated cathepsin activity in the tumour. Furthermore, we observed lower levels of cathepsins H and X and a trend for lower cathepsin B and cystatin C levels in inflammatory breast cancer patients; suggesting that cathepsins H and X might be involved in the inflammatory-dependent processes of this aggressive breast disease.
In a fifth study, we studied the impact of single nucleotide polymorphisms (SNPs) in the MMP8 gene on the expression and paradoxical protective role of MMP8 in metastasis. We found that carriers of the minor allele of the rs11225395 promoter SNP had a reduced risk of lymph node metastasis. In vitro functional assays revealed a significantly higher promoter activity in the presence of the minor allele and in support of this, we found binding of nuclear proteins at the polymorphic site for the minor allele but not for the common allele. Moreover, we observed a reduced risk of relapse as well as a better recurrence-free and overall survival among carriers of the minor allele and in particular in those with early stage breast cancer. Hence, our results suggest that MMP8 gene variation may affect breast cancer prognosis and support the notion that MMP8 has an anti-metastatic role in cancer. Doctor in de Medische Wetenschappen Afdeling Experimentele Oncologie Departement Oncologie Faculteit Geneeskunde Doctoral thesis Doctoraatsthesis
H-ras-transformed human bronchial epithelial cells (TBE-1) secrete a single major extracellular matrix metalloprotease which is not found in the normal parental cells. The enzyme is secreted in a latent form of 72 kDa, which can be activated to catalyze the cleavage of the basement membrane macromolecule type IV collagen. The substrates in their order of preference are: gelatin, type IV collagen, type V collagen, fibronectin, and type VII collagen; but the enzyme does not cleave the interstitial collagens or laminin. This protease is identical to gelatinase isolated from normal human skin explants, normal human skin fibroblasts, and SV40-transformed human lung fibroblasts. Based on its ability to initiate the degradation of type IV collagen in a pepsin-resistant portion of the molecule, it will be referred to as type IV collagenase. This enzyme is most likely the human analog of type IV collagenase detected in several rodent tumors, which has the same molecular mass and has been linked to their metastatic potential. Type IV collagenase consists of three domains. Two of them, the amino-terminal domain and the carboxyl-terminal domain, are homologous to interstitial collagenase and human and rat stromelysin. The middle domain, of 175 residues, is organized into three 58-residue head-to-tail repeats which are homologous to the type II motif of the collagen-binding domain of fibronectin. Type IV collagenase represents the third member of a newly recognized gene family coding for secreted extracellular matrix metalloproteases, which includes interstitial fibroblast collagenase and stromelysin.
H-ras-transformed human bronchial epithelial cells (TBE-1) secrete a single major extracellular matrix metalloprotease which is not found in the normal parental cells. The enzyme is secreted in a latent form of 72 kDa, which can be activated to catalyze the cleavage of the basement membrane macromolecule type IV collagen. The substrates in their order of preference are: gelatin, type IV collagen, type V collagen, fibronectin, and type VII collagen; but the enzyme does not cleave the interstitial collagens or laminin. This protease is identical to gelatinase isolated from normal human skin explants, normal human skin fibroblasts, and SV40-transformed human lung fibroblasts. Based on its ability to initiate the degradation of type IV collagen in a pepsin-resistant portion of the molecule, it will be referred to as type IV collagenase. This enzyme is most likely the human analog of type IV collagenase detected in several rodent tumors, which has the same molecular mass and has been linked to their metastatic potential. Type IV collagenase consists of three domains. Two of them, the amino-terminal domain and the carboxyl-terminal domain, are homologous to interstitial collagenase and human and rat stromelysin. The middle domain, of 175 residues, is organized into three 58-residue head-to-tail repeats which are homologous to the type II motif of the collagen-binding domain of fibronectin. Type IV collagenase represents the third member of a newly recognized gene family coding for secreted extracellular matrix metalloproteases, which includes interstitial fibroblast collagenase and stromelysin.
The matrix metalloproteinases (MMPs) are a large family of proteolytic enzymes, which are involved in the degradation of many different components of the extracellular matrix. The MMPs have been classified into different groups including collagenases, gelatinases, stromelysins, and others, particularly membrane‐type MMPs, based mainly on the in vitro substrate specificity of individual MMPs. There is increasing evidence to indicate that individual MMPs have important roles in tumour invasion and metastasis. However, the current concept of the role of MMPs in tumour invasion is that they not only have a direct role in tumour invasion by facilitating extracellular matrix degradation, but as a consequence they also have an important role in maintaining the tumour micro‐environment and thus promoting tumour growth. Inhibiting the action of MMPs represents a new therapeutic approach for the treatment of individual types of cancer and several broad‐spectrum, low‐molecular‐weight MMP inhibitors are currently being assessed for clinical use. This review examines the role of MMPs in tumour invasion and metastasis, with an emphasis on studies of clinical relevance. Copyright © 1999 John Wiley & Sons, Ltd.
Objective:
To evaluate the prognostic significance of proliferating cell nuclear antigen (PCNA) in patients with lymph node–positive primary breast cancer.
Design:
A retrospective study.
Setting:
A tertiary care hospital.
Study Participants:
A consecutive series of 123 patients with lymph node–positive primary breast cancer.
Intervention:
The PCNA-labeling index [(PCNA-positive cells/1000 cells) × 100] was quantified in paraffin-embedded tissue specimens from 123 patients with lymph node–positive primary breast cancer by immunohistochemical staining. Other important clinicopathological variables, including estrogen receptor status, histological grade, lymph node status, primary tumor status, ploidy pattern, S-phase fraction, and TNM staging, were also identified and evaluated. Main Outcome Measures: The influence of the PCNA-labeling index on the disease-free survival rate and overall survival rate.
Results:
The PCNA-labeling index of the tissue specimens tested from 123 patients ranged from 11% to 82%. The PCNA-labeling index was closely related to primary tumor status, histological grade, TNM staging, and S-phase fraction. Between patients with a high PCNA-labeling index (>35%) and those with a low PCNA-labeling index (≤35%), there were significant (P<.01) differences in both 5-year disease-free survival rates (2% vs 85%) and 5-year overall survival rates (2% vs 92%). When the PCNA-labeling index and all the clinicopathologic variables were entered into a multivariate analysis for either disease-free survival or overall survival by the Cox proportional hazards model, the PCNA-labeling index emerged as an independent prognostic factor.
Conclusion:
Based on our results, the PCNA-labeling index potentially is a useful prognostic factor for lymph node-positive primary breast cancer.Arch Surg. 1997;132:264-267
Background. Matrix metalloproteinases (MMP) are a gene family of zinc enzymes capable of degrading almost all of the extracellular matrix macromolecules in vivo. Their enzymic activities are believed to be responsible for tumor invasion and metastasis.
Methods. In this study, using peroxidase-antiperoxidase method, monospecific antisera against MMP-1 (tissue collagenase), MMP-2 (type IV collagenase/72-kilo-dalton [KD] gelatinase), and MMP-3 (stromelysin) were applied to 29 squamous cell carcinomas and normal epithelium of the esophagus to identify cells synthesizing and secreting these enzymes.
Results. Immunoreactivity of MMP-1, −2, and −3 was observed in small cancer nests of the deeply invasive or marginal portion of the tumor. Among the 29 patients studied, the presence of at least one MMP was observed in 17 (58.6%). All three enzymes were observed in six (20.6%) patients, MMP-2 and −3 in five (17.2%) patients, only MMP-2 in three (10.3%) patients, and MMP-3 alone in three (10.3%) patients. There was a good correlation among histologic stage and tumor invasion, lymph node metastasis, and MMP expression. In particular, expression of MMP-2 and −3 was closely related to lymph node metastasis and vascular invasion.
Conclusions. These results suggest that MMP, especially MMP-2 and −3, play an important role in tumor invasion and metastasis and that analysis of MMP-2 and −3 production is useful for evaluation of malignant potential in esophageal carcinoma. Cancer 1992; 702747–53.
BACKGROUND
Previous studies have shown that matrix metalloproteinase-2 (MMP-2) (a 72-kilodalton Type IV collagenase/gelatinase A) is associated with breast carcinoma, but to the authors' knowledge there are no reports showing that it is prognostic for overall survival.METHODS
Expression of the immunoreactive protein for MMP-2 was evaluated in tissue sections from primary breast carcinomas of 177 patients with a monoclonal antibody to MMP-2 using an immunohistochemical technique.RESULTSApproximately 84% of the samples were MMP-2 positive, with 22% being strongly positive. Positive MMP-2 immunostaining was prognostic for shortened survival. After 10 years 56% of the patients with tumors that were strongly positive for MMP-2 were alive, whereas 88% of patients with an MMP-2 negative tumor and 70% of patients with weakly or moderately positive tumors were still alive (chi-square test = 7.4; P < 0.01, log rank analysis). MMP-2 positivity was linked with an unfavorable prognosis regardless of the age of the patient, tumor grade, receptor status of the tumor, and stage of disease. These results were confirmed by a multivariate analysis in which MMP-2 positivity emerged as an independent prognostic factor for poor survival.CONCLUSIONS
To the authors' knowledge this study is the first time that MMP-2 immunoreactive protein has been associated strongly with a shortened survival independent of major prognostic indicators in patients with primary breast carcinoma, increasing the risk of death 3.6-fold during the first 10 years of follow-up. Cancer 1998;83:1153-1162. © 1998 American Cancer Society.
BACKGROUND
The relationship between serum levels of matrix metalloproteinase-2 (MMP-2) and MMP-3 and recurrence in patients with urothelial carcinoma after complete resection was studied to determine whether the enzymes could be a new predictor of recurrence.METHODS
Serum levels of MMP-2 and MMP-3 in 146 healthy controls, 52 patients with superficial (noninvasive) and 35 patients with advanced (invasive or metastatic) bladder carcinoma, and 30 patients with advanced upper urothelial carcinoma (renal pelvis and ureter) were measured by a one-step sandwich enzyme assay.RESULTSAmong the 53 patients with advanced urothelial carcinoma who underwent complete resection, the 1- and 3-year disease free survival rates of patients with elevated serum levels of either or both of the enzymes were 51% and 19%, respectively, whereas those of patients with normal serum levels of these enzymes were much higher, 86% and 79%, respectively (P < 0.005). In pT2N0M0 and pT3N0M0 patients who underwent complete resection, the recurrence rate in those with preoperative elevated serum levels of either or both of the enzymes was significantly higher than that in patients with normal levels (75% vs. 8%; P < 0.001). The 1- and 3-year disease free survival rates of pT2N0M0 and pT3N0M0 patients with elevated serum levels of either or both of the enzymes were 55% and 21%, respectively, which was significantly lower than that of patients with normal levels (92%; P < 0.001).CONCLUSIONS
The results of this study indicate that the elevation of serum levels of either MMP-2 or MMP-3 or both could be new predictors of recurrence in patients with advanced urothelial carcinoma after complete resection. Cancer 1996;78:2379-87.
The matrix metalloproteinases (MMPs) are a large family of proteolytic enzymes, which are involved in the degradation of many different components of the extracellular matrix. The MMPs have been classified into different groups including collagenases, gelatinases, stromelysins, and others, particularly membrane-type MMPs, based mainly on the in vitro substrate specificity of individual MMPs. There is increasing evidence to indicate that individual MMPs have important roles in tumour invasion and metastasis. However, the current concept of the role of MMPs in tumour invasion is that they not only have a direct role in tumour invasion by facilitating extracellular matrix degradation, but as a consequence they also have an important role in maintaining the tumour micro-environment and thus promoting tumour growth. Inhibiting the action of MMPs represents a new therapeutic approach for the treatment of individual types of cancer and several broad-spectrum, low-molecular-weight MMP inhibitors are currently being assessed for clinical use. This review examines the role of MMPs in tumour invasion and metastasis, with an emphasis on studies of clinical relevance. Copyright © 1999 John Wiley & Sons, Ltd.
Background: The outcome of breast cancer is usuallydetermined by multiple factors. Serum tumor necrosis factoralpha concentration has been found to be increasedin the circulation of patients with malignancy. Thisstudy was designed with the aim to investigateany correlation between the serum tumor necrosis factoralpha and the clinicopathological fetures and furthermore evaluatethe prognostic significance of serum tumor necrosis factoralpha concentration in breast cancer. Methods: Forty consecutivepatients with invasive breast cancer undergoing modified radicalmastectomy were prospectively included and evaluated. Venous bloodsamples were collected before the surgery. Sera wereobtained by centrifugation, and stored at − 70°C until assayed. The control group consisted 30healthy, age-matched subjects. Serum concentrations of tumor necrosisfactor alpha were measured by the quantitative sandwichenzyme immunoassay technique. The data on tumor size,age, estrogen receptor status, lymph node status andTNM staging were reviewed and recorded.Results: The mean value of serum tumor necrosis factor alphain patients with invasive breast cancer was 1.47± 0.58 pg/ml and that of the controlgroup was 0.98 ± 0.37 pg/ml, and thedifference was significant (P < 0.01). With univariableanalysis, patients with maximum tumor size of 5cm or larger (P=0.03), more advancedTNM staging (P < 0.01); and more advancedlymph node status (P < 0.01) were shownto have significantly higher serum concentrations of tumornecrosis factor alpha. However, with multivariable analysis, TNMstaging appeared as the only independent factor (P< 0.01) predicting the significant, higher serum concentrationsof tumor necrosis factor alpha. Conclusion: Preoperative evaluationof serum tumor necrosis factor alpha concentrations maybe a valuable parameter for reflecting the severityof staging for invasive breast cancer.
Lance A. Liotta, Patricia S. Steeg, and William G. Stetler-Stevenson Laboratory of Pathology National Cancer Institute National Institutes of Health Bethesda, Maryland 20892 The most life-threatening aspects of the oncogenic pro- cess are invasion and metastasis. Even though the clinical significance of such expression of the malignant pheno- type has been well appreciated, advances in understand- ing the molecular mechanisms involved in metastasis have lagged behind other developments in the cancer field. Progress has been hindered by the sheer complexity of this multistep, tumor-host interaction that also encom- passes angiogenic and immunologic mechanisms. To tackle the problem, investigators have separated invasion and metastasis into a series of defined, sequential steps, and focused on one step at a time. For each step, new ex- perimental models had to be developed, and a combined effort using the disciplines of cell biology, protein bio- chemistry, and molecular genetics has now resulted in a surge of new information. General themes are emerging that yield new strategies for prognosis and therapy of hu- man metastatic cancer. A group of coordinated cellular processes is responsi- ble for metastasis. Furthermore, it is now clear that nega- tive regulatory processes may be just as important as positive ones. Some genetic changes result in an im- balance of growth regulation, leading to uncontrolled proliferation. However, unrestrained growth does not, by itself, result in invasion and metastasis. The latter pheno- type may therefore require additional genetic changes. Thus, tumorigenicity and metastatic potential have both overlapping and separate features. Invasion and metasta- sis can be facilitated by proteins that stimulate tumor cell attachment to host cellular or extracellular matrix deter- minants, tumor cell proteolysis of host barriers such as the basement membrane, tumor cell locomotion, and tumor cell colony formation in the target organ for metastasis. Facilitory proteins may act at many levels intracellularly or extracellularly, but are counterbalanced by factors that can block their production, regulation, or action. A com- mon theme has emerged: in addition to loss of growth control, an imbalanced regulation of motility and proteoly- sis appears to be required for invasion and metastasis. Moreover, these same functions are also necessary for angiogenesis. Angiogenesis by normal endothelial cells and metastasis by tumor cells are functionally similar but differ in their regulation. Metastatic Cells Do Not Constitute a Minor Subpopulation The process of metastasis involves a cascade of linked, sequential steps involving multiple host-tumor interac- tions (Fidler and Hart, 1982; Schirrmacher, 1985; Liotta et
We have identified a novel membrane-type matrix metalloproteinase (MT-MMP) expressed on the cell surface and inducing activation of pro-gelatinase A in vitro. In this study, we further examined the possibility that MT-MMP is the activator of pro-gelatinase A in tumors as well as in vitro. Expression of MT-MMP mRNA was analyzed by Northern blotting in 58 cases of human lung carcinomas. MT-MMP mRNA expression was increased in tumor tissues compared with adjacent normal tissues. The ratio of MT-MMP mRNA levels in tumor/normal tissues (T/N ratio) was 3.19 +/- 1.62 in 29 cases of adenocarcinoma, 3.09 +/- 1.44 in 24 cases of squamous cell carcinoma, 4.40 +/- 0.47 in 3 cases of large cell carcinoma and 3.63 +/- 2.11 in 2 cases of small cell carcinoma, respectively. Activated gelatinase A, as detected by gelatin zymography, was also predominant in tumors compared with normal tissue counterparts, though the difference in mRNA levels was not significant. The activation ratio of gelatinase A in tumor vs. normal tissues correlated well with that of MT-MMP mRNA expression and with lymph node metastases. Our findings suggest that MT-MMP is indeed the tumor-specific activator of pro-gelatinase A in lung carcinomas and is important to initiate invasion of basement membranes.
Adjuvant treatment for node-negative breast cancer remains controversial. It is important to pick out the high-risk groups who may benefit from adjuvant systemic therapy and avoid the unnecessary additional therapy for the favorable prognostic groups.
Retrospective study of immunohistochemical staining for HLA-DR on tumor cells from paraffin-embedded tissue specimens of 32 patients with node-negative invasive breast cancer at this hospital from 1986 to 1991 was performed with the aim to investigate its prognostic significance.
HLA-DR staining was positive in nine (28%) patients and negative in twenty-three (72%) patients. One (11%) was a recurrence, and no (0%) death occurred in the positive group, compared with nine (39%) recurrences and five (22%) deaths in the negative group. The multivariate analysis failed to show that HLA-DR expression is an independent prognostic factor. However, with univariate analysis, the 5-year disease-free survival rate (87%) of the positive group was significantly better than that (35%) of the negative group (p = 0.04). The 5-year overall survival rate (100%) of the positive group was also better than that (66%) of the negative group, but the difference was not statistically significant. Furthermore, when combination of the HLA-DR expression and estrogen receptor status was used, both the 5-year disease-free and overall survival rate (81% and 100%, respectively) of group A (positive staining for either HLA-DR or estrogen receptor and positive staining for both) were significantly better than those (33% and 58%, respectively) of group B (negative staining for both HLA-DR and estrogen receptor).
We believe HLA-DR expression may be a promising, additive predictive factor to node-negative breast cancer and deserves further investigation based on these preliminary results.
Using quantitative zymography, we measured activity of the type IV collagenases metalloprotease 2 (MMP-2) and MMP-9 in 192 biopsies from colorectal carcinomas, adenomas, and normal bowel. The median level of MMP-9 in samples from Dukes' stage A (n = 18) or C (n = 48) tumors was significantly higher than in stage B carcinomas (n = 65), adenomas (n = 25), and normals (n = 36; P = 0.0001). The median level of active MMP-2 was significantly higher in stage A or C compared with adenomas (P = 0.0001) and normals (P = 0.0001). The median level of inactive MMP-2 was higher in all Dukes' stages compared with normals and adenomas (P = 0.0001). There was a significant increase in inactive MMP-2 from Jass prognostic groups I-IV (P = 0.006) but no correlation with the active enzyme. MMP activity was not related to tumor differentiation, colon versus rectal location, or disease-free, 5-year survival. All groups expressed mRNA for both enzymes, but there were quantitative and locational differences in MMP-2 mRNA expression between normal, benign, and malignant tissues. Thus MMP-2 is controlled at the level of mRNA and protein production and activation in colorectal cancer, and active MMP-2 and MMP-9 enzymes are associated strongly with Dukes' A and C stages of the disease. Variations in MMP levels with the stage or prognostic group of colorectal cancer reflect their differing stromal content.
The relationship between the serum matrix metalloproteinase-2 (MMP-2):tissue inhibitor of metalloproteinases-2 (TIMP-2) ratio and disease recurrence was examined in 53 urothelial cancer patients with muscular invasion or with lymph node metastasis who underwent complete resection. The mean MMP-2:TIMP-2 ratio in 31 patients with recurrence was significantly higher than that in 22 patients without recurrence (P < 0.05). Disease-free survival of patients with high MMP-2:TIMP-2 ratios was extremely poor compared with that of patients with lower ratios (P < 0.01). Cox's multivariate analysis suggests that the serum MMP-2:TIMP-2 ratio would be a new independent prognostic indicator of urothelial cancer recurrence.
The relationship between serum levels of matrix metalloproteinase-2 (MMP-2) and MMP-3 and recurrence in patients with urothelial carcinoma after complete resection was studied to determine whether the enzymes could be a new predictor of recurrence.
Serum levels of MMP-2 and MMP-3 in 146 healthy controls, 52 patients with superficial (noninvasive) and 35 patients with advanced (invasive or metastatic) bladder carcinoma, and 30 patients with advanced upper urothelial carcinoma (renal pelvis and ureter) were measured by a one-step sandwich enzyme assay.
Among the 53 patients with advanced urothelial carcinoma who underwent complete resection, the 1- and 3-year disease free survival rates of patients with elevated serum levels of either or both of the enzymes were 51% and 19%, respectively, whereas those of patients with normal serum levels of these enzymes were much higher, 86% and 79%, respectively (P < 0.005). In pT2N0M0 and pT3N0M0 patients who underwent complete resection, the recurrence rate in those with preoperative elevated serum levels of either or both of the enzymes was significantly higher than that in patients with normal levels (75% vs. 8%; P < 0.001). The 1- and 3-year disease free survival rates of pT2N0M0 and pT3N0M0 patients with elevated serum levels of either or both of the enzymes were 55% and 21%, respectively, which was significantly lower than that of patients with normal levels (92%; P < 0.001).
The results of this study indicate that the elevation of serum levels of either MMP-2 or MMP-3 or both could be new predictors of recurrence in patients with advanced urothelial carcinoma after complete resection.
To evaluate the prognostic significance of proliferating cell nuclear antigen (PCNA) in patients with lymph node-positive primary breast cancer.
A retrospective study.
A tertiary care hospital.
A consecutive series of 123 patients with lymph node-positive primary breast cancer.
The PCNA-labeling index [(PCNA-positive cells/1000 cells) x 100] was quantified in paraffin-embedded tissue specimens from 123 patients with lymph node-positive primary breast cancer by immunohistochemical staining. Other important clinicopathological variables, including estrogen receptor status, histological grade, lymph node status, primary tumor status, ploidy pattern, S-phase fraction, and TNM staging, were also identified and evaluated.
The influence of the PCNA-labeling index on the disease-free survival rate and overall survival rate.
The PCNA-labeling index of the tissue specimens tested from 123 patients ranged from 11% to 82%. The PCNA-labeling index was closely related to primary tumor status, histological grade, TNM staging, and S-phase fraction. Between patients with a high PCNA-labeling index (> 35%) and those with a low PCNA-labeling index (< or = 35%), there were significant (P < .01) differences in both 5-year disease-free survival rates (2% vs 85%) and 5-year overall survival rates (2% vs 92%). When the PCNA-labeling index and all the clinicopathologic variables were entered into a multivariate analysis for either disease-free survival or overall survival by the Cox proportional hazards model, the PCNA-labeling index emerged as an independent prognostic factor.
Based on our results, the PCNA-labeling index potentially is a useful prognostic factor for lymph node-positive primary breast cancer.
The outcome of breast cancer is usually determined by multiple factors. Serum tumor necrosis factor alpha concentration has been found to be increased in the circulation of patients with malignancy. This study was designed with the aim to investigate any correlation between the serum tumor necrosis factor alpha and the clinicopathological features and furthermore evaluate the prognostic significance of serum tumor necrosis factor alpha concentration in breast cancer.
Forty consecutive patients with invasive breast cancer undergoing modified radical mastectomy were prospectively included and evaluated. Venous blood samples were collected before the surgery. Sera were obtained by centrifugation, and stored at -70 degrees C until assayed. The control group consisted 30 healthy, age-matched subjects. Serum concentrations of tumor necrosis factor alpha were measured by the quantitative sandwich enzyme immunoassay technique. The data on tumor size, age, estrogen receptor status, lymph node status and TNM staging were reviewed and recorded.
The mean value of serum tumor necrosis factor alpha in patients with invasive breast cancer was 1.47 +/- 0.58 pg/ml and that of the control group was 0.98 +/- 0.37 pg/ml, and the difference was significant (P < 0.01). With univariable analysis, patients with maximum tumor size of 5 cm or larger (P = 0.03), more advanced TNM staging (P < 0.01); and more advanced lymph node status (P < 0.01) were shown to have significantly higher serum concentrations of tumor necrosis factor alpha. However, with multivariable analysis, TNM staging appeared as the only independent factor (P < 0.01) predicting the significant, higher serum concentrations of tumor necrosis factor alpha.
Preoperative evaluation of serum tumor necrosis factor alpha concentrations may be a valuable parameter for reflecting the severity of staging for invasive breast cancer.
The outcome of breast cancer is usually determined by multiple factors. Circulating intercellular adhesion molecule-1 has been found to be increased in the circulation of patients with malignancy. This study was designed with the aim of evaluating the prognostic significance of circulating intercellular adhesion molecule-1 in breast cancer.
From December 1994 to May 1995, 50 patients with invasive breast carcinoma were included. Venous blood samples were collected before surgery and the serum levels of circulating intercellular adhesion molecule-1 were measured with an enzyme immunoassay method. The data of maximum tumor size, age, estrogen receptor status, lymph node status and TNM staging were collected and evaluated simultaneously with the serum levels of circulating intercellular adhesion molecule-1. Fifteen healthy subjects were used as control group.
The mean value of circulating intercellular adhesion molecule-1 in patient group was 463 +/- 92 ng/ml and that of the control group was 346 +/- 68 ng/ml and the difference was significant (p < 0.01). In univariable analysis, patients with maximum tumor size of 5 cm or greater (p < 0.01), more advanced lymph node status (p < 0.01) and more advanced TNM staging (p < 0.01) were shown to have significantly higher serum levels of circulating intercellular adhesion molecule-1. However, in multivariable analysis, TNM staging was demonstrated to be the only independent factor (p < 0.01) related to significant, higher serum level of circulating intercellular adhesion molecule-1.
Preoperative serum levels of circulating intercellular adhesion molecule-1 may reflect the severity of staging for invasive breast cancer and may be a promising, additive predictor which deserves further investigation.
The serum-soluble interleukin-2 receptor has been claimed to be a marker of host biological response in patients with solid malignancies. This study was designed to evaluate the biological significance of the preoperative serum-soluble interleukin-2 receptor concentration in patients with invasive breast cancer.
Venous blood samples were collected from 66 patients with invasive breast carcinoma and the serum concentrations of soluble interleukin-2 receptor were measured with an enzyme immunoassay method. Data regarding maximum tumor diameter, age, estrogen receptor status, lymph node status, distant metastasis status, histologic grade, ploidy pattern and S-phase fraction were also collected and evaluated simultaneously with the serum concentration levels of soluble interleukin-2 receptor. Fifteen age-matched healthy subjects were used as a control group.
The mean value of serum-soluble interleukin-2 receptor in patients with invasive breast cancer was 621 +/- 145 units/ml and that of the control group was 308 +/- 59 units/ml; and the difference was significant (p = 0.000). With multivariate analysis, lymph node status (p = 0.000), distant metastasis status (p = 0.000) and maximum tumor diameter (p = 0.000) appeared as independent factors in regards to the significantly, higher serum concentrations of soluble interleukin-2 receptor.
Based on our preliminary results, the preoperative serum-soluble interleukin-2 receptor concentration is closely related to lymph node status, distant metastasis status and tumor diameter in invasive breast carcinoma. This may be an additional valuable predictive factor for the diagnosis of invasive breast cancer.
Previous studies have shown that matrix metalloproteinase-2 (MMP-2) (a 72-kilodalton Type IV collagenase/gelatinase A) is associated with breast carcinoma, but to the authors' knowledge there are no reports showing that it is prognostic for overall survival.
Expression of the immunoreactive protein for MMP-2 was evaluated in tissue sections from primary breast carcinomas of 177 patients with a monoclonal antibody to MMP-2 using an immunohistochemical technique.
Approximately 84% of the samples were MMP-2 positive, with 22% being strongly positive. Positive MMP-2 immunostaining was prognostic for shortened survival. After 10 years 56% of the patients with tumors that were strongly positive for MMP-2 were alive, whereas 88% of patients with an MMP-2 negative tumor and 70% of patients with weakly or moderately positive tumors were still alive (chi-square test = 7.4; P < 0.01, log rank analysis). MMP-2 positivity was linked with an unfavorable prognosis regardless of the age of the patient, tumor grade, receptor status of the tumor, and stage of disease. These results were confirmed by a multivariate analysis in which MMP-2 positivity emerged as an independent prognostic factor for poor survival.
To the authors' knowledge this study is the first time that MMP-2 immunoreactive protein has been associated strongly with a shortened survival independent of major prognostic indicators in patients with primary breast carcinoma, increasing the risk of death 3.6-fold during the first 10 years of follow-up.
The outcome of breast carcinoma is usually determined by multiple factors. Aberrant expression of the cell adhesion molecule CD 44 has been claimed to be associated with poor prognosis in various human malignancies. This study was designed to investigate any correlation between the soluble adhesion molecule CD 44 and the clinicopathologic variables and to evaluate the possible prognostic significance of soluble CD 44. Venous blood samples were preoperatively collected from 100 patients with invasive breast carcinoma. The serum levels of different soluble CD 44 molecules (CD 44 standard form and CD 44 splice variant V6) were measured with an enzyme immunoassay method. The data of primary tumor status, age, estrogen receptor status, lymph node status, histologic grading, distant metastases status, TNM staging, S-phase fraction, and ploidy pattern were collected and evaluated simultaneously with the serum levels of soluble CD 44 st and CD 44 V6. Twenty healthy subjects were used as the control group. The serum levels of soluble CD 44 st showed no significant elevation in patient group. The mean value of soluble CD 44 V6 in patient group was 269.2 +/- 94.3 ng/ml and that of the control group was 179.5 +/- 50.7 ng/ml; the difference was significant (p < 0.01). In multivariate analysis, distant metastasis (p < 0.05) and TNM staging (p < 0.01) appeared as independent factors regarding the significant higher serum levels of soluble CD 44 V6. Based on our preliminary results, preoperative serum soluble CD 44 V6 is closely related to distant metastases and TNM staging. The possible role of soluble CD 44 V6 in the prognostic value of breast carcinoma deserves further elucidation and evaluation with long-term patient follow-up.
Cancer metastasis and angiogenesis: an imbalance of positive and negative regulation
- Liotta
Matrix metalloproteinase-2 immunoreactive protein. A marker of aggressiveness in breast carcinoma
- Talvensaari-Mattila