Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.73 (urokinase-type plasminogen activator)
 10,685 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

hK4 (prostase, KLK4), a recently cloned prostate-specific serine protease and a member of the tissue kallikrein family, is a zymogen composed of 228 amino acid residues including an amino-terminal propiece, Ser-Cys-Ser-Gln-. A chimeric form of hK4 (ch-hK4) was constructed in which the propiece of hK4 was replaced by that of prostate-specific antigen (PSA) to create an activation site susceptible to trypsin-type proteases. ch-hK4 was expressed in Escherichia coli, isolated from inclusion bodies, refolded, and purified with an overall yield of 25%. The zymogen was readily self-activated during the refolding process to generate an active form (21 kDa) of hK4 (rhK4). rhK4 cleaved the chromogenic substrates Val-Leu-Arg-pNA (S-2266), Pro-Phe-Arg-pNA (S-2302), Ile-Glu-Gly-Arg-pNA (S-2222), and Val-Leu-Lys-pNA (S-2251), indicating that rhK4 has a trypsin-type substrate specificity. The rhK4 was inhibited by aprotinin (6 kDa), forming an equimolar 27 kDa complex. rhK4 readily activated both the precursor of PSA (pro-PSA) and single chain urokinase-type plasminogen activator (scuPA, pro-uPA). rhK4 also completely degraded prostatic acid phosphatase but failed to cleave serum albumin, another protein purified from human seminal plasma. These results indicate that hK4 may have a role in the physiologic processing of seminal plasma proteins such as pro-PSA, as well as in the pathogenesis of prostate cancer through its activation of pro-uPA.
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PMID:Characterization of hK4 (prostase), a prostate-specific serine protease: activation of the precursor of prostate specific antigen (pro-PSA) and single-chain urokinase-type plasminogen activator and degradation of prostatic acid phosphatase. 1173 17

Testing for tumour markers should only be performed if it results in a better patient outcome, increased quality of life or reduced overall cost of care. Ideally, the clinical value of a tumour marker should be validated in a large prospective study or a meta-analysis of small-scale retrospective/prospective studies (i.e. a level 1 evidence study) prior to routine use. Markers that have been validated in such a level 1 evidence study include carcinoembryonic antigen in the surveillance of patients with diagnosed colorectal cancer, alphafetoprotein, human chorionic gonadotrophin and lactate dehydrogenase for evaluating prognosis in patients non-seminomatous germ cell tumours, CA 125 for monitoring therapy in patients with ovarian cancer, oestrogen receptors for predicting response to hormone therapy in breast cancer, HER-2 for predicting response to trastuzumab in patients with advanced breast cancer and urokinase plasminogen activator/plasminogen activator inhibitor type 1 for determining prognosis in breast cancer. Although currently in widespread use, the value of prostate-specific antigen in screening for prostate cancer has yet to be validated in a large prospective randomized trial.
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PMID:Evidence for the clinical use of tumour markers. 1533 88

Activity-based protein profiling (ABPP) is a chemical method that utilizes active-site-directed probes to determine the functional state of enzymes in complex proteomes. Probe-labeled enzymes are typically detected by in-gel fluorescence scanning, a robust technique that nonetheless exhibits some key deficiencies, including limited sensitivity and resolution, as well as ambiguity regarding the molecular identity of the enzymes under investigation. Here, we report a microarray platform for ABPP that addresses these limitations. In this platform, proteomes are treated with ABPP probes in solution, after which labeled enzymes are captured and visualized on glass slides displaying an array of anti-enzyme antibodies. We show that ABPP microarrays exhibit superior sensitivity and resolution compared to gel-based methods, permitting the parallel analysis of several enzyme activities in proteomes, including cancer-associated proteases such as urokinase, matrix metalloproteinase-9, and prostate-specific antigen.
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PMID:Microarray platform for profiling enzyme activities in complex proteomes. 1557 75

Prostate-specific antigen (PSA) is a serine protease that is widely used as a surrogate marker in the early diagnosis and management of prostate cancer. The physiological relevance of tissue PSA levels and their role in prostate tumor growth and metastasis are not known. Free-PSA (f-PSA) was purified to homogeneity from human seminal plasma by column chromatography, eliminating hk2 and all known PSA complexes and retaining its protease activity. Confluent monolayers of prostate cancer cell lines, PC-3M and LNCaP, were treated with f-PSA in a series of in vitro experiments to determine the changes in expression of various genes that are known to regulate tumor growth and metastasis. Gene array, quantitative polymerase chain reaction (QPCR), and enzyme-linked immunosorbent assay (ELISA) results show significant changes in the expression of various cancer-related genes in PC-3M and LNCaP cells treated with f-PSA. In a gene array analysis of PC-3M cells treated with 10 muM f-PSA, 136 genes were upregulated and 137 genes were downregulated. In LNCaP cells treated with an identical concentration of f-PSA, a total of 793 genes was regulated. QPCR analysis reveals that the genes for urokinase-type plasminogen activator (uPA), VEGF, and Pim-1 oncogene, known to promote tumor growth, were significantly downregulated, whereas IFN-gamma, known to be a tumor-suppressor gene, was significantly upregulated in f-PSA-treated PC-3M cells. The effect of f-PSA on VEGF and IFN-gamma gene expression and on protein release in PC-3M cells was distinctly dose-dependent. In vivo studies showed a significant reduction (P = .03) in tumor load when f-PSA was administered in the tumor vicinity of PC-3M tumor-bearing BALB/c nude mice. Our data support the hypothesis that f-PSA plays a significant role in prostate tumor growth by regulating various proangiogenic and antiangiogenic growth factors.
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PMID:Prostate-specific antigen modulates the expression of genes involved in prostate tumor growth. 1579 24

The plasminogen activation system is involved in cancer progression and metastasis. Among other proteolytic factors, it includes the serine protease urokinase-type plasminogen activator (uPA) and its three-domain (D1D2D3) receptor uPAR (CD87), which focuses plasminogen activation to the cell surface. The function of uPAR is regulated in part through shedding of domain D1 by proteases, e.g., uPA itself or plasmin. Human tissue kallikrein 4 (hK4), which is highly expressed in prostate and ovarian tumor tissue, was previously shown to cleave and activate the pro-enzyme forms of prostate-specific antigen (PSA, tissue kallikrein hK3) and uPA. Here we demonstrate that uPAR is also a target for hK4, being cleaved in the D1-D2 linker sequence and, to a lesser extent, in its D3 juxtamembrane domain. hK4 may thus modulate the tumor-associated uPA/uPAR-system activity by either activating the pro-enzyme form of uPA or cleaving the cell surface-associated uPA receptor.
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PMID:Interplay of human tissue kallikrein 4 (hK4) with the plasminogen activation system: hK4 regulates the structure and functions of the urokinase-type plasminogen activator receptor (uPAR). 1649 55

Protein C inhibitor (PCI) is a heparin-dependent serine protease inhibitor found in human plasma, urine, and other body fluids. In blood plasma, PCI is present at approximately 0.08 microM and inactivates activated protein C and other coagulation and fibrinolytic enzymes. In seminal plasma, PCI is present at 2.2 to 3.7 microM. The main sources of seminal PCI are the seminal vesicles, where it remains fully active. Following ejaculation, PCI completely looses its activity in approximately 2 hours, when it partially complexes with prostate-specific antigen, two plasminogen activators (urokinase-type and tissue-type plasminogen activators), and tissue kallikrein. PCI is also present in an active form in follicular fluid at approximately 0.1 microM. Purified functionally active human blood plasma-derived as well as inactive semen-derived PCI inhibited both binding and penetration of zona-free hamster oocytes by human sperm. The binding inhibition by PCI was dose dependent. A concentration of 0.04 microM PCI (approximately 100-fold lower than that present in seminal plasma) inhibited 50% of the binding and penetration ability. Given that capacitated sperm used for in vitro fertilization usually contains more than 0.05 microM of PCI, fertilization rates might be significantly reduced. All of these data suggest that PCI is involved in human reproduction at several steps, including the fertilization process.
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PMID:The role of protein C inhibitor in human reproduction. 1725 88

The introduction of prostate-specific antigen (PSA) has revolutionized the detection and management of patients with prostate cancer. Despite this there has always been a concern among clinicians about the usefulness of total PSA levels as a marker for prostate cancer. We discuss the use of calculated variables and molecular forms of PSA. The precursor forms of PSA have been associated with the presence and biological behaviour of prostate cancer. With recent advances in biotechnology, e.g. high-throughput molecular analyses, many potential blood biomarkers have been identified and are currently under investigation. Given the plethora of candidate biomarkers we discuss a selected group of novel blood-based biomarkers, e.g. human glandular kallikrein, early prostate cancer antigen, insulin-like growth factors, urokinase plasminogen activators, transforming growth factor-beta, interleukin-6, chromogranin A, and prostate secretory protein. While these and other markers have shown promise in early-phase studies, no single biomarker is likely to have the appropriate degree of certainty to dictate treatment decisions. Consequently, the future of cancer prognosis might rely on small panels of markers that can accurately predict cancer presence, stage and metastasis, and serve as prognosticators, targets, and/or surrogate endpoints of disease progression and response to therapy.
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PMID:New blood-based biomarkers for the diagnosis, staging and prognosis of prostate cancer. 1794 30

The introduction of prostate-specific antigen (PSA) revolutionized prostate cancer (PCa) screening and ushered the PSA era. However, its use as a screening tool remains controversial and changes in the epidemiology of PCa have strongly limited its prognostic role. Therefore, we need novel approaches to improve our ability to detect PCa and foretell the course of the disease. To improve the specificity of total PSA, several approaches based on PSA derivatives have been investigated such as age-specific values, PSA density (PSAD), PSAD of the transition zone, PSA velocity and assessment of various isoforms of PSA. With recent advances in biotechnology such as high-throughput molecular analyses, many potential blood biomarkers have been identified and are currently under investigation. Given the plethora of candidate PCa biomarkers, we have chosen to discuss a select group of candidate blood-based biomarkers including human glandular kallikrein, early prostate cancer antigens, insulin-like growth factor-I (IGF-I) and its binding proteins (IGFBP-2 and IGFBP-3), urokinase plasminogen activation system, transforming growth factor-beta1, interleukin-6, chromogranin A, prostate secretory protein, prostate-specific membrane antigen, PCa-specific autoantibodies and alpha-methylacyl-CoA racemase. While these and other markers have shown promise in early phase studies, no single biomarker is likely to have the appropriate degree of certainty to dictate treatment decisions. Consequently, the future of cancer prognosis may rely on small panels of markers that can accurately predict PCa presence, stage, metastasis, and serve as prognosticators, targets and/or surrogate end points of disease progression and response to therapy.
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PMID:New circulating biomarkers for prostate cancer. 1799 18

OBJECTIVE To evaluate the expression of urokinase-plasminogen-activator receptor (uPA-R) in disseminated tumour cells (DTC) in bone marrow (BM) and peripheral blood (PB) of patients with clinically localized prostate cancer before radical prostatectomy (RP), and to assess the associations with pathological variables and prognosis. PATIENTS AND METHODS In all, 52 patients (47 with clinically localized cancer and five with benign prostatic hyperplasia, BPH, as controls) were prospectively enrolled. BM and PB samples were drawn before surgery. DTC were enriched using a commercial system, cytokeratin (CK) 8/18 was used to detect DTC, and uPA-R expression was detected by dual-immunostaining of the DTC. The final pathology of the RP specimen was compared with the results of immunostaining. Follow-up was initiated to detect tumour relapse (defined as a prostate-specific antigen (PSA) level of > or =0.2 ng/mL). RESULTS Overall, there was expression of 'CK + uPA-R' in 60% of the BM and in 19% of the PB specimens. Expression of this marker in BM was most significantly increased in those with unfavourable Gleason scores (P = 0.004), followed by high-risk cancer (P = 0.005). The relative risk for CK + uPA-R expression in the BM was 3.1 times higher in high-risk than in low-risk prostate cancer. No relevant expression rates were detected for PB. In the control group, no patient showed CK or uPA-R expression in BM or PB. The PSA-recurrence free survival was significantly lower in patients with CK + uPA-R-positive BM cells (P = 0.01). CONCLUSION In this pilot study, the preoperative detection rate of CK + uPAR expression in BM of patients with prostate cancer increased with Gleason score and in those with high-risk disease. All patients with a later PSA relapse had had uPA-R expression in their DTC from the BM. DTC with uPA-R expression was an adverse prognostic factor for prostate cancer.
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PMID:Urokinase-plasminogen-activator receptor expression in disseminated tumour cells in the bone marrow and peripheral blood of patients with clinically localized prostate cancer. 1915 51

We present the rapid prototyping of electrochemical sensor arrays integrated to microfluidics towards the fabrication of integrated microsystems prototypes for point-of-care diagnostics. Rapid prototyping of microfluidics was realised by high-precision milling of polycarbonate sheets, which offers flexibility and rapid turnover of the desired designs. On the other hand, the electrochemical sensor arrays were fabricated using standard photolithographic and metal (gold and silver) deposition technology in order to realise three-electrode cells comprising gold counter and working electrodes as well as silver reference electrode. The integration of fluidic chips and electrode arrays was realised via a laser-machined double-sided adhesive gasket that allowed creating the microchannels necessary for sample and reagent delivery. We focused our attention on the reproducibility of the electrode array preparation for the multiplexed detection of tumour markers such as carcinoembryonic antigen and prostate-specific antigen as well as genetic breast cancer markers such as estrogen receptor-alpha, plasminogen activator urokinase receptor, epidermal growth factor receptor and erythroblastic leukemia viral oncogene homolog 2. We showed that by carefully controlling the electrode surface pre-treatment and derivatisation via thiolated antibodies or short DNA probes that the detection of several key health parameters on a single chip was achievable with excellent reproducibility and high sensitivity.
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PMID:Design and testing of a packaged microfluidic cell for the multiplexed electrochemical detection of cancer markers. 1973 40


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