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Eleftherios P. Diamandis - One of the best experts on this subject based on the ideXlab platform.
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Human tissue Kallikreins: Blood levels and response to radiotherapy in intermediate risk prostate cancer
Radiotherapy and Oncology, 2017Co-Authors: Nicola J Nasser, John Thoms, Ri Wang, Antoninus Soosaipillai, Eleftherios P. Diamandis, Melania Pintilie, Robert G BristowAbstract:Abstract Objectives Kallikreins are serine proteases over expressed in many malignancies. In this study, we measure changes in serum Kallikrein (KLKs) levels during intensity-modulated radiotherapy (IMRT) in prostate cancer patients, and find potential correlations between serum Kallikrein level and normal tissues toxicity during radiation. Methods Forty-nine patients with prostate cancer were recruited as follows: group 1, definitive standard fractionation IMRT (78Gy in 39 fractions, n =15); group 2, definitive hypofractionated IMRT (60Gy in 20 fractions, n =15); and group 3, IMRT postprostatectomy (66Gy in 33 fractions, n =19). Patients treated with definitive radiation therapy were intermediate risk. Blood samples were collected at baseline and quarterly during IMRT and at each follow-up visit. Acute toxicity was graded weekly during radiotherapy using CTC-AE v4.0 criteria. Multiplexed immunoassays were used to quantify total, free, and intact Prostate Specific Antigen (PSA), as well as Kallikreins 2, 4, 6, and 11. Results The serum Kallikreins, PSA (total, free and intact), KLK2, 6, and 11 change significantly after definitive radiotherapy. KLK2 and intact PSA decrease as fast as two weeks after initiation of radiation, while the first significant decrease in total and free PSA is noted only at the completion of radiation. KLK6 and KLK11 surge temporarily during radiation therapy and decrease below baseline levels at 8weeks and 12months, respectively after completion of radiation. KLK4 levels did not change with radiation. There was no correlation between GU or GI toxicities and serum Kallikreins. Conclusions PSA, KLK2, 6, and 11, change significantly after definitive prostate radiotherapy, though KLK2 and PSA decrease by the end of the radiation course while KLK6 and KLK11 decrease significantly starting at 2 and 12months, respectively, after radiation. There was no correlation between GU or GI toxicities and serum Kallikreins.
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Unleashing the therapeutic potential of human Kallikrein-related serine proteases
Nature Reviews Drug Discovery, 2015Co-Authors: Ioannis Prassas, Azza Eissa, Gennadiy Poda, Eleftherios P. DiamandisAbstract:Tissue Kallikreins (KLKs) constitute a family of 15 secreted serine proteases that are encoded by the largest protease gene cluster in the human genome. KLKs were traditionally known for their clinical applicability as cancer markers — for instance, KLK3 (also known as prostate-specific antigen) is a marker for prostate cancer. The field of KLK research has recently blossomed with the development of KLK-knockout models and the elucidation of the 3D structures of several KLKs. Novel pathophysiological roles for these proteases have recently been assigned in various tissues, such as the airway, cardiovascular system, tooth, skin and brain tissues. The promise of KLKs as therapeutic targets in various pathologies — including skin diseases, hereditary angioedema, neurodegeneration, inflammation and cancer — is emerging. Systematic efforts for the development of the first generation of KLK-based inhibitors as candidate therapeutics have already been initiated. Tissue Kallikreins are a family of fifteen secreted serine proteases encoded by the largest protease gene cluster in the human genome. In the past decade, substantial progress has been made in characterizing the natural substrates, endogenous inhibitors and in vivo functions of Kallikreins, and studies have delineated important pathophysiological roles for these proteases in a variety of tissues. Thus, Kallikreins are now considered attractive targets for the development of novel therapeutics for airway, cardiovascular, tooth, brain, skin and neoplastic diseases. In this Review, we discuss recent advances in our understanding of the physiological functions and pathological implications of Kallikrein proteases, and highlight progress in the identification of Kallikrein inhibitors, which together are bringing us closer to therapeutically targeting Kallikreins in selected disease settings. Members of the family of Kallikrein (KLK)-related proteases are found in various tissues — including the airway, prostate and brain — and have a wide range of functions. The authors describe the roles of KLKs in health and disease, and highlight the small-molecule, peptide-based, protein-based, antibody-based and immunotherapeutic strategies that are being used to target KLKs in certain diseases.
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Evaluation of the prognostic significance of human Kallikrein 8 protein expression levels in advanced ovarian cancer by using automated quantitative protein analysis
Journal of Clinical Oncology, 2007Co-Authors: Panteleimon Kountourakis, Andreas Scorilas, Eleftherios P. Diamandis, A. Psyrri, S. Markakis, Diane Kowalski, Robert L. Camp, Meletios-athanassios DimopoulosAbstract:5581 Background: Kallikreins, a subgroup of the serine protease enzyme family, are considered important prognostic biomarkers in cancer. Here, we sought to determine the prognostic value of Kallikrein 8 (hkl8) in ovarian cancer using a novel method of compartmentalized in situ protein analysis. Materials and Methods: A tissue array composed of 150 advanced stage ovarian cancers, uniformly treated with surgical debulking followed by platinum-paclitaxel combination chemotherapy, was constructed. For evaluation of Kallikrein 8 protein expression, we used an immunofluorescence-based method of automated in situ quantitative measurement of protein analysis (AQUA). Results: Mean follow-up time of the cohort was 34.35 months. One hundred twenty six of 150 cases had sufficient tissue for AQUA analysis. There was association between tumor mask hk8 protein expression levels and clinicopathological variables including grade (p=0.0011), residual disease (p=0.0063),clinical response to chemotherapy (p=0.0346). In univa...
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Coordinated steroid hormone-dependent and independent expression of multiple Kallikreins in breast cancer cell lines
Breast Cancer Research and Treatment, 2007Co-Authors: Miltiadis Paliouras, Eleftherios P. DiamandisAbstract:The regulation of gene expression by steroid hormones plays an important role in the normal development and function of many organs, as well in the pathogenesis of endocrine-related cancers. Previous experiments have shown that many Kallikrein genes are under steroid hormone regulation in breast cancer cell lines. We here examine the coordinated expression of multiple Kallikrein genes in several breast cancer cell lines after steroid hormone stimulation. Breast cancer cell lines were treated with various steroid hormones and Kallikrein ( KLK/ hK) expression of hK3 (prostate-specific antigen, PSA), hK5, hK6, hK7, hK8, hK10, hK11, hK13, and hK14 was analyzed at the RNA level via RT-PCR and at the protein level by immunofluorometric ELISA assays. We identified several distinct hK hormone-dependent and hormone-independent expression patterns. Hormone-specific modulation of expression was seen for several Kallikreins in BT-474, MCF-7, and T-47D cell lines. hK6 was specifically up-regulated upon estradiol treatment in all three cell lines whereas PSA expression was induced by dihydrotestosterone (DHT) and norgestrel stimulation in BT-474 and T-47D. hK10, hK11, hK13, and hK14 were specifically up-regulated by DHT in T-47D and by estradiol in BT-474 cells. Bioinformatic analysis of upstream proximal promoter sequences for these hKs did not identify any recognizable hormone-response elements (HREs), suggesting that the coordinated activation of these four hKs represents a unique expression “cassette”, utilizing a common hormone-dependent mechanism. We conclude that groups of human hKs are coordinately expressed in a steroid hormone-dependent manner. Our data supports clinical observations linking expression of multiple hKs with breast cancer prognosis.
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Human tissue Kallikreins: The cancer biomarker family
Cancer Letters, 2007Co-Authors: Miltiadis Paliouras, Carla Borgono, Eleftherios P. DiamandisAbstract:Human tissue Kallikreins (KLKs) are attracting increased attention due to their role as biomarkers for the screening, diagnosis, prognosis, and monitoring of various cancers including those of the prostate, ovarian, breast, testicular, and lung. Human tissue Kallikrein genes represent the largest contiguous group of proteases within the human genome. Originally thought to consist of three genes, the identification of the human Kallikrein locus has expanded this number to fifteen. These genes, and their encoded proteins, share a high degree of homology and are expressed in different tissues. Prostate-specific antigen (PSA), the most commonly known Kallikrein, is a useful biomarker for prostate cancer. Several other Kallikreins, including Kallikreins 2 (KLK2) and 11 (KLK11) are emerging as complementary prostate cancer biomarkers. Along with these Kallikreins, several others have been implicated in the other cancers. For example, KLK5, 6, 7, 10, 11, and 14 are emerging biomarkers for ovarian cancer. The identification of Kallikrein substrates and the development of proteolytic cascade models implicate Kallikrein proteins in cancer progression. This review describes the current status of Kallikreins as cancer biomarkers. © 2006 Elsevier Ireland Ltd. All rights reserved.
Hans Lilja - One of the best experts on this subject based on the ideXlab platform.
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Prostate-specific Kallikrein-related peptidases and their relation to prostate cancer biology and detection
Thrombosis and Haemostasis, 2020Co-Authors: Daniel L. J. Thorek, Sigrid V. Carlsson, David Ulmert, Michael J Evans, Hans LiljaAbstract:summaryKallikreins are a family of serine proteases with a range of tissue-specific and essential proteolytic functions. Among the best studied are the prostate tissue-specific KLK2 and KLK3 genes and their secreted protease products, human Kallikrein 2, hk2, and prostate-specific antigen (PSA). Members of the so-called classic Kallikreins, these highly active trypsin-like serine proteases play established roles in human reproduction. Both hK2 and PSA expression is regulated by the androgen receptor which has a fundamental role in prostate tissue development and progression of disease. This feature, combined with the ability to sensitively detect different forms of these proteins in blood and biopsies, result in a crucially important biomarker for the presence and recurrence of cancer. Emerging evidence has begun to suggest a role for these Kallikreins in critical vascular events. This review discusses the established and developing biological roles of hK2 and PSA, as well as the historical and advanced use of their detection to accurately and non-invasively detect and guide treatment of prostatic disease.
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improving the specificity of screening for lethal prostate cancer using prostate specific antigen and a panel of Kallikrein markers a nested case control study
European Urology, 2015Co-Authors: Par Stattin, Andrew J Vickers, Daniel D Sjoberg, Robert Johansson, Torvald Granfors, Mattias Johansson, Kim Pettersson, Peter T Scardino, Goran Hallmans, Hans LiljaAbstract:Background: A disadvantage of prostate-specific antigen (PSA) for the early detection of prostate cancer (PCa) is that many men must be screened, biopsied, and diagnosed to prevent one death. Objective: To increase the specificity of screening for lethal PCa at an early stage. Design, setting, and participants: We conducted a case-control study nested within a population-based cohort. PSA and three additional Kallikreins were measured in cryopreserved blood from a population-based cohort in Vasterbotten, Sweden. Of 40 379 men providing blood at ages 40, 50, and 60 yr from 1986 to 2009, 12 542 men were followed for >15 yr. From this cohort, the Swedish Cancer Registry identified 1423 incident PCa cases, 235 with distant metastasis. Outcome measurements and statistical analysis: Risk of distant metastasis for different PSA levels and a prespecified statistical model based on the four Kallikrein markers. Results and limitations: Most metastatic cases occurred in men with PSA in the top quartile at age 50 yr (69%) or 60 yr (74%), whereas 20-yr risk of metastasis for men with PSA below median was low (� 0.6%). Among men with PSA >2 ng/ml, a prespecified model based on four Kallikrein markers significantly enhanced the prediction of metastasis compared with PSA alone. About half of all men with PSA >2 ng/ml were defined as low risk by this model and had a � 1% 15-yr risk of metastasis. Conclusions: Screening at ages 50-60 yr should focus on men with PSA in the top quartile. A marker panel can aid biopsy decision making. Patient summary: For men in their fifties, screening should focus on those in the top 10% to 25% of PSA values because the majority of subsequent cases of distant metastasis are found among these men. Testing of four Kallikrein markers in men with an elevated PSA could aid biopsy decision making. # 2015 European Association of Urology. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by- nc-nd/4.0/).
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Prostate-specific Kallikrein-related peptidases and their relation to prostate cancer biology and detection. Established relevance and emerging roles.
Thrombosis and Haemostasis, 2013Co-Authors: Daniel L. J. Thorek, Sigrid Carlsson, David Ulmert, Michael J Evans, Hans LiljaAbstract:Kallikreins are a family of serine proteases with a range of tissue-specific and essential proteolytic functions. Among the best studied are the prostate tissue-specific KLK2 and KLK3 genes and their secreted protease products, human Kallikrein 2, hk2, and prostate-specific antigen (PSA). Members of the so-called classic Kallikreins, these highly active trypsin-like serine proteases play established roles in human reproduction. Both hK2 and PSA expression is regulated by the androgen receptor which has a fundamental role in prostate tissue development and progression of disease. This feature, combined with the ability to sensitively detect different forms of these proteins in blood and biopsies, result in a crucially important biomarker for the presence and recurrence of cancer. Emerging evidence has begun to suggest a role for these Kallikreins in critical vascular events. This review discusses the established and developing biological roles of hK2 and PSA, as well as the historical and advanced use of their detection to accurately and non-invasively detect and guide treatment of prostatic disease.
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a four Kallikrein panel predicts prostate cancer in men with recent screening data from the european randomized study of screening for prostate cancer rotterdam
Clinical Cancer Research, 2010Co-Authors: Andrew J Vickers, Kim Pettersson, Peter T Scardino, Angel M Cronin, Monique J Roobol, Caroline Savage, Mari T Peltola, Fritz H Schroder, Hans LiljaAbstract:Purpose—We have developed a statistical prediction model for prostate cancer based on four Kallikrein markers in blood: total, free, and intact prostate specific antigen (PSA) and Kallikreinrelated peptidase 2 (hK2). Although this model accurately predicts the result of biopsy in unscreened men, its properties for men with a history of PSA screening have not been fully characterized. Experimental Design—1501 previously screened men with elevated PSA underwent initial biopsy during rounds 2 and 3 of the European Randomized Study of Prostate Cancer Screening, Rotterdam, with 388 cancers diagnosed. Biomarker levels were measured in serum samples taken before biopsy. The prediction model developed on the unscreened cohort was then applied and predictions compared to biopsy outcome. Results—The previously developed four-Kallikrein prediction model had much higher predictive accuracy than PSA and age alone (area-under-the-curve of 0.711 vs. 0.585 and 0.713 vs. 0.557 with and without digital rectal exam, respectively; both p<0.001). Similar statistically significant enhancements were seen for high-grade cancer. Applying the model with a cut-off of 20% cancer risk as the criterion for biopsy would reduce the biopsy rate by 362 for every 1000 men with elevated PSA. Although diagnosis would be delayed for 47 cancers, these would be predominately low stage and low grade (83% Gleason 6 T1c). Conclusions—A panel of four Kallikreins can help predict the result of initial biopsy in previously screened men with elevated PSA. Use of a statistical model based on the panel would substantially decrease rates of unnecessary biopsy.
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New nomenclature for the human tissue Kallikrein gene family
Clinical Chemistry, 2000Co-Authors: Eleftherios P. Diamandis, Shigetaka Yoshida, Linda K. Ashworth, Torbjörn Egelrud, Sheila P. Little, Judith A. Clements, Sadao Shiosaka, Peter S. Nelson, George M Yousef, Hans LiljaAbstract:The human Kallikrein gene family is important to the discipline of clinical chemistry because it contains genes that encode for valuable cancer biomarkers, including the best tumor marker available today, prostate-specific antigen (PSA). Despite reports of numerous Kallikrein-like genes in the mouse (1), until 2–3 years ago, only three human Kallikrein genes were recognized: pancreatic/renal Kallikrein (KLK1) , human glandular Kallikrein 2 (KLK2) , and prostate-specific antigen (KLK3) (1)(2). The proteins encoded by the three Kallikrein genes are now known as hK1, hK2, and hK3 (PSA). These three genes encode for serine proteases with either trypsin-like (hK1, hK2) or chymotrypsin-like (hK3) activity. Traditionally, Kallikreins have been defined as enzymes that can act on high-molecular weight substrates and release bioactive peptides, known as kinins (3). Among the known Kallikrein enzymes, only plasma Kallikrein (encoded by a single gene localized on human chromosome 4q35; official symbol KLKB1 ) and pancreatic/renal Kallikrein (hK1) have significant kininogenase activity. The proteins encoded by the KLK2 and the KLK3 genes have minimal or no kininogenase activity (4)(5). Why then are the KLK1 , KLK2 , and KLK3 genes classified together into one gene family (tissue Kallikrein gene family), when two of the three enzymes have no significant Kallikrein enzymatic activity? This grouping is justified, based on the extensive homologies between the three genes at both the DNA …
Jean Dube - One of the best experts on this subject based on the ideXlab platform.
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human Kallikrein hk2 has low kininogenase activity while prostate specific antigen hk3 has none
Biochimica et Biophysica Acta, 1997Co-Authors: David Deperthes, Francois Marceau, Gilles Frenette, Roland R Tremblay, Claude Lazure, Jean DubeAbstract:Abstract In the present paper, we determined the kinin-releasing activity of human prostatic Kallikrein hK2 and compared it to one of the Kallikreins hK1 and prostate specific antigen (hK3). Kinin-like substances active on the rabbit jugular vein were progressively produced when nanomolar concentrations of hK2 were incubated with heated plasma. However in these experiments, hK1 appeared much more potent than hK2 while hK3 was totally inactive. When hK2 was incubated with purified high molecular weight kininogen, several peptides were generated as shown by the analysis on C18 reverse-phase HPLC. Kinin activity was localized exclusively in a small peak having an elution time identical to that of bradykinin while the only important peak obtained with hK1 corresponded to Lys-bradykinin. Finally, the rate of kinin production of hK2 was found to be more than a thousandfold lower than that of hK1. These experiments show that Kallikreins hK2 has only a low kininogenase activity. However, it is not excluded that some of the peptides produced by hK2 action could have other types of biological activity.
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prostatic Kallikrein hk2 but not prostate specific antigen hk3 activates single chain urokinase type plasminogen activator
International Journal of Cancer, 1997Co-Authors: Gilles Frenette, Roland R Tremblay, Claude Lazure, Jean DubeAbstract:Our work was undertaken to compare the relative efficiency of 2 purified prostatic Kallikreins, namely, hK2 and prostate-specific antigen (PSA or hK3), in the activation of single-chain urokinase (scuPA). We found that hK2 converts scuPA into an active enzyme with an efficiency equal to approximately 1/50 that of plasmin. During the activation of scuPA by hK2, two fragments of 33 and 22 kDa were generated. The NH2-terminal amino acid sequence of the 33 kDa fragment showed that hK2 cleaved scuPA between Lys158 and Ile159. In contrast to a previous report by another group, our purified hK3 preparation containing no trypsin-like contaminants was totally unable to activate scuPA. Our results show that Kallikrein hK2 has plasmin-like activity and suggest that it could be the initiator of a proteolytic cascade leading to prostatic cancer invasion. Int. J.Cancer 71: 897-899, 1997. © 1997 Wiley-Liss Inc.
Roland R Tremblay - One of the best experts on this subject based on the ideXlab platform.
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serpin derived peptide substrates for investigating the substrate specificity of human tissue Kallikreins hk1 and hk2
Journal of Biological Chemistry, 1997Co-Authors: Luc Bourgeois, David Deperthes, Roland R Tremblay, Michele Brillardbourdet, Jean Y Dube, Maria Aparecida Juliano, Luiz Juliano, Francis GauthierAbstract:Abstract The third human tissue Kallikrein to be identified, hK2, could be an alternate or complementary marker to Kallikrein hK3 (prostate-specific antigen) for prostate diseases. Most of the hK2 in seminal plasma forms an inactive complex with protein C inhibitor (PCI), a serpin secreted by seminal vesicles. As serpin inhibitors behave as suicide substrates that are cleaved early in the interaction with their target enzyme, and Kallikreins have different sensitivities to serpin inhibitors, we prepared a series of substrates with intramolecularly quenched fluorescence based on the sequences of the serpin reactive loops. They were used to compare the substrate specificities of hK1 and hK2, which both have trypsin-like specificity, and thus differ from chymotrypsin-like hK3. The serpin-derived peptides behaved as Kallikrein substrates whose sensitivities reflected the specificity of the parent inhibitory proteins. Substrates derived from PCI were the most sensitive for both hK1 and hK2 with specificity constants of about 107 m −1. s−1. Those derived from antithrombin III and α2-antiplasmin were more specific for hK2 while a kallistatin-derived substrate was specifically cleaved by hK1. hK1 and hK2 substrates of greater specificity were obtained using chimeric peptides based on the sequence of serpin reactive loops. The main difference between specificities of hK1 and hK2 arise because hK2 can accommodate positively charged as well as small residues at P2 and requires an arginyl residue at P1. Thus, unlike hK1, hK2 does not cleave kininogen-derived substrates overlapping the region of N-terminal insertion of bradykinin in human kininogens.
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human Kallikrein hk2 has low kininogenase activity while prostate specific antigen hk3 has none
Biochimica et Biophysica Acta, 1997Co-Authors: David Deperthes, Francois Marceau, Gilles Frenette, Roland R Tremblay, Claude Lazure, Jean DubeAbstract:Abstract In the present paper, we determined the kinin-releasing activity of human prostatic Kallikrein hK2 and compared it to one of the Kallikreins hK1 and prostate specific antigen (hK3). Kinin-like substances active on the rabbit jugular vein were progressively produced when nanomolar concentrations of hK2 were incubated with heated plasma. However in these experiments, hK1 appeared much more potent than hK2 while hK3 was totally inactive. When hK2 was incubated with purified high molecular weight kininogen, several peptides were generated as shown by the analysis on C18 reverse-phase HPLC. Kinin activity was localized exclusively in a small peak having an elution time identical to that of bradykinin while the only important peak obtained with hK1 corresponded to Lys-bradykinin. Finally, the rate of kinin production of hK2 was found to be more than a thousandfold lower than that of hK1. These experiments show that Kallikreins hK2 has only a low kininogenase activity. However, it is not excluded that some of the peptides produced by hK2 action could have other types of biological activity.
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prostatic Kallikrein hk2 but not prostate specific antigen hk3 activates single chain urokinase type plasminogen activator
International Journal of Cancer, 1997Co-Authors: Gilles Frenette, Roland R Tremblay, Claude Lazure, Jean DubeAbstract:Our work was undertaken to compare the relative efficiency of 2 purified prostatic Kallikreins, namely, hK2 and prostate-specific antigen (PSA or hK3), in the activation of single-chain urokinase (scuPA). We found that hK2 converts scuPA into an active enzyme with an efficiency equal to approximately 1/50 that of plasmin. During the activation of scuPA by hK2, two fragments of 33 and 22 kDa were generated. The NH2-terminal amino acid sequence of the 33 kDa fragment showed that hK2 cleaved scuPA between Lys158 and Ile159. In contrast to a previous report by another group, our purified hK3 preparation containing no trypsin-like contaminants was totally unable to activate scuPA. Our results show that Kallikrein hK2 has plasmin-like activity and suggest that it could be the initiator of a proteolytic cascade leading to prostatic cancer invasion. Int. J.Cancer 71: 897-899, 1997. © 1997 Wiley-Liss Inc.
George M Yousef - One of the best experts on this subject based on the ideXlab platform.
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Cloning of a Kallikrein pseudogene
Clinical Biochemistry, 2004Co-Authors: George M Yousef, Carla Borgono, Iacovos P Michael, Eleftherios P. DiamandisAbstract:Objectives: Kallikreins are a group of serine proteases clustered together on a small region of chromosome 19q13.4. Recent reports suggest that Kallikreins are differentially expressed in malignancy and have potential as cancer biomarkers. The human Kallikrein gene locus has now been fully characterized and 15 functional Kallikreins were identified. Although many Kallikrein pseudogenes have already been characterized in rodents, none have been identified in humans. Methods and results: In the current study, we identified the first human Kallikrein pseudogene named WKLK1 and mapped it between the KLK2 and KLK4 genes. This pseudogene shares a moderate degree of similarity with the adjacent functional Kallikreins. It has a conserved histidine residue of the catalytic triad of serine proteases and its surrounding motif, but lacks the aspartate and serine residues. Positions of some cysteine residues are also conserved in the pseudogene. This pseudogene lacks intronic sequences and should thus be classified as a processed pseudogene. EST and PCR analyses indicate that this pseudogene may be transcriptionally active, because mRNA was detected in many tissues including the prostate, testis, pituitary, and adrenal glands, as well as in tissues of the female genital organs. Discussion: The mRNA sequence of the gene is, however, defective and is not predicted to code for a protein. Highly conserved sequences were found in the flanking region of the pseudogene, thus supporting the view that it evolved by retrotransposition. We also identified another serine protease fragment that has only the conserved histidine residue. The functional significance of the pseudogene and the other fragment is yet to be identified. D 2004 The Canadian Society of Clinical Chemists. All rights reserved.
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An update on human and mouse glandular Kallikreins.
Clinical Biochemistry, 2004Co-Authors: Eleftherios P. Diamandis, George M Yousef, A. Yvonne OlssonAbstract:Human glandular Kallikreins are secreted serine proteases, involved in many biological processes. Recently, the complete organization of the human and mouse genomic loci has been elucidated. These loci harbor the largest clusters of serine proteases within the human and mouse genomes. Mouse orthologs to all human Kallikrein genes, except for KLK2 and KLK3 genes, have now been identified. Here, we describe an update of the genomic organization of these families in human and mouse, and provide some thoughts for future research directions.
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the human Kallikrein protein 5 hk5 is enzymatically active glycosylated and forms complexes with two protease inhibitors in ovarian cancer fluids
Biochimica et Biophysica Acta, 2003Co-Authors: George M Yousef, Carl Kapadia, Maryellen Polymeris, Carla Borgoňo, Shirley Hutchinson, G Wasney, Antoninus SoosaipillaiAbstract:The Kallikrein family is a group of 15 serine protease genes clustered on chromosome 19q13.4. Binding of Kallikreins to protease inhibitors is an important mechanism for regulating their enzymatic activity and may have potential clinical applications. Human Kallikrein gene 5 (KLK5) is a member of this family and encodes for a secreted serine protease (hK5). This Kallikrein was shown to be differentially expressed at the mRNA and protein levels in diverse malignancies. Our objective was to study the enzymatic activity and the interaction of recombinant hK5 protein with protease inhibitors. Recombinant hK5 protein was produced in yeast and mammalian expression systems and purified by chromatography. HPLC fractionation, followed by ELISA-type assays, immunoblotting and radiolabeling experiments were performed to detect the possible interactions between hK5 and proteinase inhibitors in serum. Enzymatic deglycosylation was performed to examine the glycosylation pattern of the protein. The enzymatic activity of hK5 was tested using trypsin and chymotrypsin-specific synthetic fluorogenic substrates. In serum and ascites fluid, in addition to the free (f40 kDa) form, hK5 forms complexes with a1-antitrypsin and a2macroglobulin. These complexes were detected by hybrid ELISA-type assays using hK5-specific coating antibodies and inhibitor detection antibodies. The ability of hK5 to bind to these inhibitors was further verified in vitro. Spiking of serum samples with 125 I-labeled hK5 results in the distribution of the protein in two higher molecular mass (bound) forms, in addition to the unbound form. The hK5 mature enzyme is active and shows trypsin, but not chymotrypsin-like, activity. The pro-form of hK5 is not active. Recombinant hK5 shows a higher than predicted molecular mass due to glycosylation. hK5 is partially complexed with a1-antitrypsin and a2-macroglobulin in serum and ascites fluid of ovarian cancer patients. The recombinant protein is glycosylated and its mature form shows trypsin-like activity. D 2003 Elsevier B.V. All rights reserved.
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Down-regulation of the human Kallikrein gene 5 (KLK5) in prostate cancer tissues.
The Prostate, 2002Co-Authors: George M Yousef, Albert Chang, Laura M. Rendl, Maria Diamandis, Andreas Scorilas, Klaus Jung, Eleftherios P. DiamandisAbstract:BACKGROUND. Kallikreins are a subgroup of serine proteases with diverse physiological functions. Many Kallikrein genes are differentially expressed in various malignancies and prostate specific antigen (PSA; encoded by the KLK3 gene) is the best tumor marker for prostate cancer. Human glandular Kallikrein (hK2; encoded by the KLK2 gene) is an emerging tumor marker for prostate cancer. KLK5 is a newly discovered human Kallikrein gene which shares a high degree of homology and is located adjacent to KLK2 and KLK3 genes on chromosome 19q13.4. Like KLK2 and KLK3, the KLK5 gene is regulated by steroid hormones in the BT-474 breast cancer cell line. We have previously shown that KLK5 is differentially expressed in ovarian and breast cancer. METHODS. We compared the expression of KLK5 in 29 pairs of histologically confirmed normal and prostate cancer tissues by quantitative RT-PCR using the LightCycler technology. RESULTS. KLK5 expression was significantly lower in cancer tissues compared to their normal counterparts. Lowest levels of expression were found in T3 stage tumors compared with T1 and T2. Also, a significant negative correlation was found between Gleason score and KLK5 expression. CONCLUSIONS. KLK5 should be further studied as a potential new prognostic marker in prostate cancer, whose expression is negatively correlated with cancer aggressiveness. Prostate
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the new human tissue Kallikrein gene family structure function and association to disease
Endocrine Reviews, 2001Co-Authors: George M Yousef, Eleftherios P. DiamandisAbstract:The human tissue Kallikrein gene family was, until recently, thought to consist of only three genes. Two of these human Kallikreins, prostate-specific antigen and human glandular Kallikrein 2, are currently used as valuable biomarkers of prostatic carcinoma. More recently, new Kallikrein-like genes have been discovered. It is now clear that the human tissue Kallikrein gene family contains at least 15 genes. All genes share important similarities, including mapping at the same chromosomal locus (19q13.4), significant homology at both the nucleotide and protein level, and similar genomic organization. All genes encode for putative serine proteases and most of them are regulated by steroid hormones. Recent data suggest that at least a few of these Kallikrein genes are connected to malignancy. In this review, we summarize the recently accumulated knowledge on the human tissue Kallikrein gene family, including gene and protein structure, predicted enzymatic activities, tissue expression, hormonal regulation, and alternative splicing. We further describe the reported associations of the human Kallikreins with various human diseases and identify future avenues for research.
