Research Progress of Targeting uPA-uPAR System in Tumor Therapy / 肿瘤防治研究

The uPA-uPAR system is highly expressed in various tumor tissues. This system can promote the degradation of extracellular matrix proteins, as well as combine with vitronectin and integrin to transmit intracellular signal transduction. Subsequently, it mediates the occurrence and development of tumors. In recent years, a series of therapeutic programs that target this system has achieved notable results in tumor treatment, and some of them have been under the clinical trial stage, thus providing new ideas for tumor targeted therapy. Therefore, this paper intends to provide a review of research progress on the gene therapy, drug therapy, and immunotherapy targeting uPA-uPAR system.

Relevance of the plasminogen activation system in the pathogenesis and progression of gastric cancer / La importancia del sistema activador de plasminógeno en la patogénesis y progresión del cáncer gástrico

Abstract Gastric cancer is ranked as the third death-causing cancer and one of the most incident malignancies worldwide. AlthoughHelicobacter pyloriis the most well-established risk factor for the development of this neoplasm, most of the infected individuals do not develop gastric cancer. Two of the main challenges faced by the world's scientific community in the combat against gastric cancer are the unraveling of its pathogenesis and the identification of novel ways to bring down the mortality. Malignant cell invasion of the non-neoplastic adjacent tissue and metastasis are pivotal events during cancer development and progression. Both processes are facilitated by proteases capable of degrading components of the extracellular matrix, some of which have been associated to clinic-pathological aspects of the disease. Recent studies have suggested the possible connection betweenH. pyloriand the expression of some of these proteases in gastric mucosa. This review summarizes the current knowledge about epidemiological, clinical and biological aspects of gastric cancer; it also discusses the main findings about the involvement of the plasminogen activation system in the development and progression of this disease, as well as its potential repercussions in the clinical setting. Rev. Biol. Trop. 66(1): 28-47. Epub 2018 March 01.

Resumen El cáncer gástrico es la tercera causa de muerte por cáncer a nivel mundial y uno de los más incidentes. A pesar de que la infección porHelicobacter pylories el factor de riesgo más reconocido para el desarrollo de esta neoplasia, la mayoría de personas infectadas con la bacteria no desarrolla la enfermedad. Dos de los principales desafíos a los que actualmente se enfrenta la comunidad científica mundial en la lucha contra el cáncer gástrico son el esclarecimiento de la patogénesis y la identificación de nuevos parámetros que contribuyan a disminuir la mortalidad. La invasión de las células malignas al tejido no neoplásico adyacente y la metástasis son eventos claves durante el desarrollo y progresión del cáncer. Lo anterior es facilitado por proteasas capaces de degradar los componentes de la matriz extracelular, algunas de las cuales han sido asociadas con aspectos clínico-patológicos de la enfermedad. Estudios recientes han sugerido la posible relación entre la bacteriaH. pyloriy la inducción en mucosa gástrica de algunas de estas proteasas. Esta revisión resume conocimientos actuales sobre aspectos epidemiológicos, clínicos y biológicos del cáncer gástrico; también discute los principales hallazgos en torno a la participación del sistema activador de plasminógeno en el desarrollo y progresión del mismo, así como sus potenciales repercusiones en la práctica clínica.

Construction and identification of specific shRNA interference plasmid vector targeted to uPAR gene / 中国癌症杂志

Background and purpose: Urokinase-type plasminogen activator receptor is related to invasion and metastasis of tumor. Inhibition of uPAR expression in tumor cells results in reducing its metastasis. This study was aimed to construct an expression vector with short hairpin RNA (shRNA) of uPAR, which could pave the way for RNAi-mediated tongue squamous cell carcinoma therapy. Methods: Genome sequences of uPAR gene were retrieved from Genhank and cDNA was designed to code expression of shRNA for uPAR gene. The cDNA was synthesized and inserted into the eukaryotic expression vector pWH1, and the recombinant pWH1-uPAR expression vector was identified by enzyme cutting method. Then, pWH1-uPAR expression vector was transfected into tongue squamous cell carcinoma Ts cells by Lipofectomine 2000. At last, the expression of uPAR in Ts cells transfected with pWH1-uPAR expression vector was observed by RT-PCR, immunocytochemistry staining and Western blot. MTT assay was performed to measure the proliferation of Ts cell. Results: The uPAR shRNA eukaryotic expression vector was successfully constructed. Compared with Ts cells and Ts cells transfected with plasmid pWH1, the Ts cells transfected with pWHI-uPAR expression vector showed a lower mRNA and protein expression of uPAR. The inhibition rate of proliferation was 32.9% of Ts cells by transfected with pWHl- uPAR. Conclusion: The constructed uPAR shR.NA expression vector could inhibit the expression of uPAR in tongue squamous cell carcinoma, which may be helpful for further research on the function of uPAR and provide effective methods for therapy of tongue squamous cell carcinoma.

Effect of Arsenic Trioxide on Human Hepatoma Cell Proliferation and Its uPAR Expression / 中国药房

OBJECTIVE:To investigate the effect of arsenic trioxide(As2O3)on human hepatoma cells SMMC-7721 viability and the expression of urokinase plasminogen activator receptor(uPAR).METHODS:SMMC-7721 cells were cultured and treated with As2O3 at different concentrations for different time with untreated SMMC-7721 cells(without As2O3 treatment)served as control.The proliferation rate of the SMMC-7721 cells was measured by MTT assay;the change of cell cycle was detected by flow cytometry analysis;the effect of As2O3 on uPAR mRNA levels was analyzed by real-time fluorescent quantitative polymerase chain reaction.RESULTS:As2O3 had significant inhibitory effect on SMMC-7721 proliferation in time-dependent and concentration-dependent manner.As compared with control group,As2O3 significantly decreased the proportion of cells in G0/G1 phase but increased the cell proportion in G2/M phase,and it markedly down-regulated the level of uPAR mRNA expression(P

Hepatocyte Growth Factor/c-Met Signaling in Regulating Urokinase Plasminogen Activator in Human Stomach Cancer: A Potential Therapeutic Target for Human Stomach Cancer

BACKGROUND: Up-regulation of the hepatocyte growth factor (HGF), its transmembrane tyrosine kinase receptor (c-Met), and urokinase type plasminogen activator (uPA), is associated with the development and metastasis of various types of cancers. However, the mechanisms by which HGF/c-Met signaling mediates cancer progression and metastasis are unclear. METHODS: We investigated the roles of HGF/c-Met in tumor progression and metastasis in NUGC-3 and MKN-28 stomach cancer cell lines. RESULTS: Treatment with HGF increased c-Met phosphorylation in a dose-dependent manner, as well as increasing cell proliferation. HGF treatment also increased the protein level and the activity of uPA in NUGC-3 and MKN-28 cells. A monoclonal antibody against human uPA receptor (uPAR), mAb 3936, inhibited HGF-mediated tumor cell invasion in a dose-dependent manner. Down-regulation of uPA using uPA-shRNA induced a decrease in in vitro cell invasion in NUGC-3 cells. CONCLUSIONS: These results suggest that NUGC-3 and MKN-28 cells express functional c-Met, which may provide a therapeutic target for interfering with metastases of cancer cells by inhibiting uPA and uPAR-mediated proteolysis.

mRNA Expression Differences of uPA, uPAR in Eutopic Endometrium of Advanced Stage Endometriosis Patients / 대한불임학회지

OBJECTIVE: We investigated the expression of uPA and uPAR in eutopic endometrium of advanced stage endometriosis and control patients. METHODS: The 33 endometriosis patients and 32 controls were enrolled. Endometrial samples were obtained from 65 premenopausal women aged 29~44 years, undergoing laparoscopic surgery or hysterectomy for non-malignant lesions. Sufficient samples were collected from 33 patients with endometriosis stage III and IV and 32 controls without endometriosis confirmed by laparoscopic surgery. The mRNA expression of uPA and uPAR from eutopic endometrium were analyzed by RT-QC PCR. RESULTS: The mRNAs of uPA and uPAR were expressed in eutopic endometrium from endometriosis and normal controls throughout the menstrual cycle. Uterine endometrium from women with endometriosis expresses significantly (p<0.05) higher levels of u-PA mRNA than endometrium from normal women without endometriosis in the proliferative phase. There were no significant differences in expression of uPAR in eutopic endometrium between controls and endometriosis patients. CONCLUSION: These results suggest that eutopic endometrium from endometriosis patients may be more invasive and prone to peritoneal implantation because of greater u-PA mRNA expression than endometrium from women without endometriosis. Thus, increased proteolytic activity may be one etiology for the invasive properties of the endometrium resulting in the development of endometriosis.

Involvement of MAPK pathway in hypoxia-induced up-regulation of urokinase plasminogen activator receptor in a human prostatic cancer cell line, PC3MLN4

Clinical studies have shown that tumor hypoxia is associated with invasive growth and metastasis and may be an important prognostic factor adversely influencing survival in patients with tumors. To investigate the mechanisms involved in hypoxia-induced invasive growth and metastasis, hypoxia-mediated urokinase plasmalogen activator receptor (uPAR) expression, cellular invasiveness, and mitogen activated protein kinase (MAPK) activation were measured in a prostate cancer cell line, PC3MLN4. The levels of uPAR expression and cellular invasiveness were increased in hypoxic cells. Hypoxia-induced cellular invasiveness was blocked by an anti-uPAR monoclonal antibody. Phosphorylations of ERK and p38 kinases were also more extensive in hypoxic cells than in normoxic cells. Hypoxia-induced uPAR up-regulation was inhibited by pre-treatments with a specific inhibitor of MEK, PD98059 and a specific inhibitor of p38 MAP kinase, SB203580. Cell growth also increased in hypoxic cells. From these results, hypoxia increased tumor cell invasion by up-regulating uPAR expression, which might be mediated through ERK and p38 kinase signaling pathways in PC3MLN4 prostate cancer cell line.

Urokinase Plasminogen Activator Receptor Gene Expression and Clinico-Pathologic Feature in Gastric Cancer Patients

PUPOSE: Invasion and metastasis in solid tumors require the action of tumor-associated proteases. The serine protease urokinase-type plasminogen (uPA) and receptor (uPAR) appear to have a major function in these processes. Expression of the uPAR is elevated in breast and colon carcinomas, and this is often associated with invasiveness and poor prognosis. The purpose of this study was to determine whether the expression of the uPAR gene correlates with clinico-pathological parameters in human gastric carcinomas. MATERIALS AND METHODS: We examined the expression of uPAR mRNA by using northern blot analysis and RT-PCR in 35 gastric carcinomas and the surrounding normal mucosa. Macroscopic and histopathological tumor findings and survival rates were obtained from the patient records and from endoscopic, surgical, and pathological reports. RESULTS: The expression of uPAR and was higher in most neoplasms than in the corresponding normal mucosal tissue. uPAR mRNA expression in tumors correlated well with lymph-node metastasis (P<0.02) and tumor stage (P<0.01). The survival rate of patients with tumors displaying high uPAR expression levels was significantly lower (P<0.04) than that of patients without uPAR expression, but IL-8 showed only the tendency of survival difference. CONCLUSION: These results suggest that uPAR may be an important prognostic factor in human gastric carcinomas.

Construction and expression of recombinant plasmid pEGFPC1uPAR in Pam 212 cells / 第三军医大学学报

Objective To construct the eukaryotic expression plasmid of pEGFPC1uPAR gene and explore the effect on the proliferation and invasion ability of Pam 212 cells. Methods The human uPAR cDNA was cloned by PCR, and inserted into the eukaryotic expression plasmid pEGFPC1. After identification of sequencing, the reconstructive plasmid was transformed transiently into Pam 212 cells, then the cell growth and the invasion ability were evaluated. Results The reconstructive plasmid of pEGFPC1uPAR was validated by sequencing. The reconstructive plasmid can promote the growth of Pam 212 cells and enhance the invasion ability. Conclusion The pEGFPC1uPAR plasmid was constructed successfully and uPAR was confirmed to promote the growth and the invasion ability of Pam 212 cells, which lay the foundation for further studies of uPAR in vivo.

Culture of outer root sheath cells of murine vibrissa follicles in vitro and expression of uPA/uPAR / 第三军医大学学报

Objective To establish a method for the cultivation of hain follicle outer root sheath (ORS) cells of murine vibrissa and to study their expression of uPA/uPAR. Methods The ORS cells of murine vibrissa were cultured by combination of enzyme digestion and tissue culture with the feeder layer of dermal sheath cells of the hair follicle. The cells and their uPA/uPAR expression were identified by immunocytochemistry (ICC). Results The feeder layer of DS was suitable for the culture of ORS cells. ORS cells expressed uPA/uPAR protein. Conclusion The DS of hair follicle is a better feeder layer for the growth of ORS cells. ORS cells have the ability to migrate and proliferate.

Invasion-Metastasis by Hepatocyte Growth Factor/c-Met Signaling Concomitant with Induction of Urokinase Plasminogen Activator in Human Pancreatic Cancer: Role as Therapeutic Target / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: Increased expression of the hepatocytes growth factor (HGF) receptor (c-Met) and urokinase type plasminogen activator (uPA) correlate with the development and metastasis of cancers. However, the mechanisms by which HGF/c-Met signaling mediate cancer progression and metastasis are unclear. Therefore, we investigated the roles of HGF/c-Met in tumor progression and metastasis in pancreatic cancer cell lines, L3.6PL and IMIN-PC2. MATERIALS AND METHODS: To see the functional c-Met protein, we were performed immunoprecipitation for functional c-Met protein. And also performed western bolot analysis and gel zymography for the functional uPA protein. To see the inhibition effects of uPAR monoclonal antibody on invasiveness of two pancreatic cancer cell lines, we were carried out standard two chamber invasion assay. RESULTS: At first, we observed the HGF-mediated c-Met phosphorylation and cell growth. c-Met phosphorylation was increased in the HGF-treated cells in a dose dependent manner. HGF resulted in increments of cell growth and ERK phosphorylation. HGF treatment increased the uPA expression and the uPA activity. A monoclonal antibody 3936, specific to uPAR receptor, inhibited HGF- mediated tumor cell invasion in a dose dependent manner. CONCLUSION: These results suggest that functional c- Met and HGF/c-Met signaling up-regulate the activity of uPA and result in increments of invasion-metastasis in the pancreatic cancer cells.

Transfection of maspin/PCR2.1 vector inhibits uPA and uPAR expressions in gastric cancer cells in vitro / 第三军医大学学报

Objective To investigate the inhibitory effect of maspin gene on metastasis and invasion of gastric cancer and its mechanism.Methods Maspin gene was ligated to the expression vector PCR2.1 with T4 DNA ligase after the PCR2.1 was digested by HindⅢ/XbaⅠ.The recombinant vector maspin/PCR2.1 was transfected into human gastric cancer cell lines MKN28 and SGC7901,then the mRNA and protein expression changes of maspin gene,uPA and uPAR were detected respectively by PT-PCR and Western blotting.Results After identified by digestion and sequencing,the reconstructed plasmid was confirmed to contain the correct and full nucleotide sequence of maspin gene.The expressions of maspin gene were up-regulated in MKN28 and SGC7901 cell lines,while the expressions of uPA and uPAR were down-regulated.Conclusion The expression vector maspin/PCR2.1 is constructed successfully and can be expressed in eukaryotic cells.Expressions of uPA and uPAR can be inhibited by maspin gene.

Expression of urokinase-type plasminogen activator (uPA) and its receptor (uPAR) in hepatocellular carcinoma and its clinical and pathological significance / 中国微创外科杂志

0.05). Positive stains were located in cytoplasm of cancer and stromal cells, especially in invasive margin of cancer. Slight positive stains were observed in non-cancer tissues. But in the control samples, there wasn't specific stain. In ELISA, the uPA antigen levels in cancer and non- cancer tissues were (4.23?0.57)ng/mg protein and (1.26?0.14)ng/mg protein; the uPAR antigen levels in caner and non-cancer tissues were(4.25?0.21)ng/mg protein and (3.15?0.23)ng/mg protein respectively. Significant difference existed in both uPA and uPAR antigen levels in these two kinds of tissue ( t=18.963,P=0.000;t=13 693,P=0 000 ). Furthermore, these antigen levels in progressive and aggressive tumors significantly higher than those in non-progressive and non-aggressive tumors ( P

Plasma levels of soluble urokinase-type plasminogen activator receptor in lung cancer and its clinical significance / 中国实用内科杂志

Objective To study the expression of urokinase-type plasminogen activator receptor (uPAR) in the peripheral blood and investigate its value for the metastasis and prognosis of lung cancer.Methods The plasma levels of suPAR in 26 normal adults,54 patients with lung cancer were measured by using enzyme linked immunosorbent assay (ELISA).Results The plasma suPAR levels of lung cancer patients were increased in comparison with that of normal adults (P

Clinical Relevance of Urokinase-type Plasminogen Activator ( uPA ) , uPA Receptor , Plasminogen Activator Inhibitor-1 Co-expression from Tissue and Serum of Breast Cancer as Targets of Biotherapy / 대한암학회지

PURPOSE: We measured and compared the uPA, plasminogen activator inhibitor-1 (PAI-1) and uPA receptor (uPAR) levels in breast cancer tissues and blood of the patients to evaluate their clinical relevance for biotherapy. MATERIALS AND METHODS: uPA, PAI-1 (Monozyme, Netherland), uPAR (American Diagnostics, USA) levels were measured by ELISA assay in 192 breast cancer tissues, in 18 normal breast tissues and in 163 blood from breast cancer patients. RESULTS: There was a tendency of uPA increment from ductal carcinoma in situ while increment of PAI-1 and uPAR occurred from Ti. With the progression of cancer, uPA, PAI-1, uPAR tended to decrease; however, the uPA/uPAR, uPA/PAI-1 ratios remained unchanged. There was a correlation of uPA expression between normal and cancer tissues ( r(2)= 0.49). Correlation of uPA and PAI-1 was found in normal tissue and stage I cancer tissue while correlation of uPAR and PAI-1 was found with cancer progression. Between cancer tissue and blood significant correlations were found in uPA, PAI-1, uPAR levels. CONCLUSION: uPA, PAI-1, uPAR levels in cancer tissue elevated from the early stage maintaining correlative expressions with cancer progression. A positive correlation between cancer tissue and blood level suggested the applicability of the levels of uPA, PAI-1 or uPAR for detecting patients for biotherapy.

Study on expression and clinical significance of uPAR and VEGF-C in ovarian cancer / 中国癌症杂志

Background and purpose:urokinase-typeplasminoogen activator receptor(uPAR) plays important roles in terms of fibrolysis in tissue,cell adhering prolifration and signal transduction uPAR can the detected in both cancer and matrix cells.It has been reorted that uPAR is associated with invasion and metastasis of cancer cells.This paper is to study the effects of the gene expression of uPAR and VEGF-C on onco-genesis and progression of ovarian cancer.Methods:The expression of uPAR gene protein and VEGF-C were examined immunohistochemically in 58 cases of ovarian serous cystadenocarcinoma,10 cases of borderline serous cystadenomas and 13 cases of serous cystadenocarcinoma.The relationships between the results and pathological features were studied.Results:① The expressions of uPAR in benign and malignant tumors were different significantly(P

The Inhibitory Effects of an Antisense u-PAR Vector on Invasion by Highly Invasive Human Prostate Carcinoma PC-3M Cell Subclones / 中国肿瘤生物治疗杂志

Objective: To observe the inhibiting effects of an antisense u-PAR vector on invasion by highly invasive PC-3M cell subclones. Methods: The effects of an antisense vector on invasion by highly invasive PC-3M cell subclones were observed and compared in vitro by monolayer invasion assay and soft agar clone. Then, both a quantitative RT-PCR and zymography were used to exam the effects of the antisense u-PAR on activity of MMP-9 in those highly invasive cell subclones. Furthermore, the tumorigenesis rate and invasions by the cell subclones with or without the antisense u-PAR were observed in nude mice. Results: It is found that the speed of growth in vitro was slowing down by highly invasive PC-3M cell subclones transfected with the antisense u-PAR, and the ability of anchorage-independent growth of those cell subclones was also decreasing sharply,and the inhibiting rate was 79% and 60%, respectively. Although the antisense u-PAR didn′t change MMP-9 gene transcription, but they could inhibit the activation of MMP-9 of highly invasive PC-3M cell subclones. Moreover, the tumorigenesis rate of the cell subclones with the antisense u-PAR decreased and the growth of a neoplasm also slowed down. The t tests showed the difference between experimental and control groups reached statistical significance ( P