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01-06-2011 | Preclinical study

Overexpression of the urokinase receptor mRNA splice variant uPAR-del4/5 affects tumor-associated processes of breast cancer cells in vitro and in vivo

Authors: Sumito Sato, Charlotte Kopitz, Bettina Grismayer, Nathalie Beaufort, Ute Reuning, Manfred Schmitt, Thomas Luther, Matthias Kotzsch, Achim Krüger, Viktor Magdolen

Published in: Breast Cancer Research and Treatment | Issue 3/2011

Abstract

uPAR, the three-domain membrane receptor of the serine protease urokinase, plays a crucial role in tumor growth and metastasis. Several mRNA splice variants of this receptor have been reported. One of these, uPAR-del4/5, lacking exons 4 and 5, and thus encoding a uPAR form lacking domain DII, is specifically overexpressed in breast cancer and represents a statistically independent prognostic factor for distant metastasis-free survival in breast cancer patients. The aim of the present study was to examine the molecular and cellular properties of the encoded uPAR-del4/5 protein. To investigate the impact of the uPAR-del4/5 overexpression on in vitro and in vivo aspects of tumor progression (e.g., proliferation, adhesion, invasion, metastatic seeding, and/or metastatic growth), we combined the analysis of transfected cancer cell lines with a murine xenograft tumor model. Increased expression of uPAR-del4/5 in human cancer cells led to reduced adhesion to several extracellular matrix proteins and decreased invasion through Matrigel^TM, while cell proliferation was not affected in vitro. Moreover, invasion of uPAR-del4/5 overexpressing cells was not altered by addition of urokinase, while that of uPAR-wild-type overexpressing cells was drastically increased. Accordingly, we observed that, in contrast to uPAR-wild-type, uPAR-del4/5 does not interact with urokinase. On the other hand, when overexpressed in human breast cancer cells, uPAR-del4/5 distinctly impaired metastatic dissemination and growth in vivo. We demonstrate that the uPAR-del4/5 mRNA splice variant mediates tumor-relevant biological processes in vitro and in vivo. Our results thus illustrate how tumor-specific alternative splicing can distinctly impact the biology of the tumor.

Blasi F, Sidenius N (2010) The urokinase receptor: focused cell surface proteolysis, cell adhesion and signaling. FEBS Lett 584:1923–1930PubMedCrossRef

Blasi F, Carmeliet P (2002) uPAR: a versatile signalling orchestrator. Nat Rev Mol Cell Biol 3:932–943PubMedCrossRef

Reuning U, Magdolen V, Wilhelm O, Fischer K, Lutz V, Graeff H, Schmitt M (1998) Multifunctional potential of the plasminogen activation system in tumor invasion and metastasis. Int J Oncol 13:893–906PubMed

Harbeck N, Kates RE, Gauger K, Willems A, Kiechle M, Magdolen V, Schmitt M (2004) Urokinase-type plasminogen activator (uPA) and its inhibitor PAI-I: novel tumor-derived factors with a high prognostic and predictive impact in breast cancer. Thromb Haemost 91:450–456PubMed

Høyer-Hansen G, Lund IK (2007) Urokinase receptor variants in tissue and body fluids. Adv Clin Chem 44:65–102PubMedCrossRef

Harris L, Fritsche H, Mennel R, Norton L, Ravdin P, Taube S, Somerfield MR, Hayes DF, Bast RC (2007) American Society of Clinical Oncology 2007 update of recommendations for the use of tumor markers in breast cancer. J Clin Oncol 25:5287–5312PubMedCrossRef

Ploug M (2003) Structure-function relationships in the interaction between the urokinase-type plasminogen activator and its receptor. Curr Pharm Des 9:1499–1528PubMedCrossRef

Smith HW, Marshall CJ (2010) Regulation of cell signalling by uPAR. Nat Rev Mol Cell Biol 11:23–36PubMedCrossRef

Reuning U, Magdolen V, Hapke S, Schmitt M (2003) Molecular and functional interdependence of the urokinase-type plasminogen activator system with integrins. Biol Chem 384:1119–1131

10.

Llinas P, Le Du MH, Gårdsvoll H, Danø K, Ploug M, Gilquin B, Stura EA, Ménez A (2005) Crystal structure of the human urokinase plasminogen activator receptor bound to an antagonist peptide. EMBO J 24:1655–1663PubMedCrossRef

11.

Huai Q, Mazar AP, Kuo A, Parry GC, Shaw DE, Callahan J, Li Y, Yuan C, Bian C, Chen L, Furie B, Furie BC, Cines DB, Huang M (2006) Structure of human urokinase plasminogen activator in complex with its receptor. Science 311:656–659PubMedCrossRef

12.

Gårdsvoll H, Ploug M (2007) Mapping of the vitronectin-binding site on the urokinase receptor: involvement of a coherent receptor interface consisting of residues from both domain I and the flanking interdomain linker region. J Biol Chem 282:13561–13572PubMedCrossRef

13.

Huai Q, Zhou A, Lin L, Mazar AP, Parry GC, Callahan J, Shaw DE, Furie B, Furie BC, Huang M (2008) Crystal structures of two human vitronectin, urokinase and urokinase receptor complexes. Nat Struct Mol Biol 15:422–423PubMedCrossRef

14.

Stewart CE, Sayers I (2009) Characterisation of urokinase plasminogen activator receptor variants in human airway and peripheral cells. BMC Mol Biol 10:75PubMedCrossRef

15.

Luther T, Kotzsch M, Meye A, Langerholc T, Fussel S, Olbricht N, Albrecht S, Ockert D, Muehlenweg B, Friedrich K, Grosser M, Schmitt M, Baretton G, Magdolen V (2003) Identification of a novel urokinase receptor splice variant and its prognostic relevance in breast cancer. Thromb Haemost 89:705–717PubMed

16.

Pyke C, Eriksen J, Solberg H, Nielsen BS, Kristensen P, Lund LR, Danø K (1993) An alternatively spliced variant of mRNA for the human receptor for urokinase plasminogen activator. FEBS Lett 326:69–74PubMedCrossRef

17.

Kotzsch M, Farthmann J, Meye A, Fuessel S, Baretton G, Tjan-Heijnen VC, Schmitt M, Luther T, Sweep FC, Magdolen V, Span PN (2005) Prognostic relevance of uPAR-del4/5 and TIMP-3 mRNA expression levels in breast cancer. Eur J Cancer 41:2760–2768PubMedCrossRef

18.

Kotzsch M, Sieuwerts AM, Grosser M, Meye A, Fuessel S, Meijer-van Gelder ME, Smid M, Schmitt M, Baretton G, Luther T, Magdolen V, Foekens JA (2008) Urokinase receptor splice variant uPAR-del4/5-associated gene expression in breast cancer: identification of rab31 as an independent prognostic factor. Breast Cancer Res Treat 111:229–240PubMedCrossRef

19.

Möbus V, Gerharz CD, Press U, Moll R, Beck T, Mellin W, Pollow K, Knapstein PG, Kreienberg R (1992) Morphological, immunohistochemical and biochemical characterization of 6 newly established human ovarian carcinoma cell lines. Int J Cancer 52:76–84PubMedCrossRef

20.

Luther T, Magdolen V, Albrecht S, Kasper M, Riemer C, Kessler H, Graeff H, Müller M, Schmitt M (1997) Epitope-mapped monoclonal antibodies as tools for functional and morphological analyses of the human urokinase receptor in tumor tissue. Am J Pathol 150:1231–1244PubMed

21.

Goretzki L, Bognacki J, Koppitz M, Rettenberger P, Magdolen V, Creutzburg S, Hammelburger J, Weidle UH, Wilhelm O, Kessler H, Graeff H, Schmitt M (1997) Quantitative assessment of interaction of urokinase-type plasminogen activator and its receptor (CD87) by use of a solid-phase uPA-ligand binding assay. Fibrinol Proteol 11:11–19CrossRef

22.

Krüger A, Soeltl R, Sopov I, Kopitz C, Arlt M, Magdolen V, Harbeck N, Gänsbacher B, Schmitt M (2001) Hydroxamate-type matrix metalloproteinase inhibitor batimastat promotes liver metastasis. Cancer Res 61:1272–1275PubMed

23.

Kopitz C, Anton M, Gänsbacher B, Krüger A (2005) Reduction of experimental human fibrosarcoma lung metastasis in mice by adenovirus-mediated cystatin C overexpression in the host. Cancer Res 65:8608–8612PubMedCrossRef

24.

Krüger A, Schirrmacher V, Khokha R (1998) The bacterial lacZ gene: an important tool for metastasis research and evaluation of new cancer therapies. Cancer Metastasis Rev 17:285–294PubMedCrossRef

25.

Beaufort N, Leduc D, Eguchi H, Mengele K, Hellmann D, Masegi T, Kamimura T, Yasuoka S, Fend F, Chignard M, Pidard D (2007) The human airway trypsin-like protease modulates the urokinase receptor (uPAR, CD87) structure and functions. Am J Physiol Lung Cell Mol Physiol 292:1263–1272CrossRef

26.

Wei Y, Lukashev M, Simon DI, Bodary SC, Rosenberg S, Doyle MV, Chapman HA (1996) Regulation of integrin function by the urokinase receptor. Science 273:1551–1555PubMedCrossRef

27.

Xue W, Mizukami I, Todd RF, Petty HR (1997) Urokinase-type plasminogen activator receptors associate with beta1 and beta3 integrins of fibrosarcoma cells: dependence on extracellular matrix components. Cancer Res 57:1682–1689PubMed

28.

Krüger A, Soeltl R, Lutz V, Wilhelm OG, Magdolen V, Rojo EE, Hantzopoulos PA, Graeff H, Gänsbacher B, Schmitt M (2000) Reduction of breast carcinoma tumor growth and lung colonization by overexpression of the soluble urokinase-type plasminogen activator receptor (CD87). Cancer Gene Ther 7:292–299PubMedCrossRef

29.

Setyono-Han B, Sturzebecher J, Schmalix WA, Muehlenweg B, Sieuwerts AM, Timmermans M, Magdolen V, Schmitt M, Klijn JG, Foekens JA (2005) Suppression of rat breast cancer metastasis and reduction of primary tumour growth by the small synthetic urokinase inhibitor WX-UK1. Thromb Haemost 93:779–786PubMed

30.

Ward AJ, Cooper TA (2010) The pathobiology of splicing. J Pathol 220:152–163PubMed

31.

Fackenthal JD, Godley LA (2008) Aberrant RNA splicing and its functional consequences in cancer cells. Dis Model Mech 1:37–42PubMedCrossRef

32.

Bjarnadóttir TK, Geirardsdóttir K, Ingemansson M, Mirza MA, Fredriksson R, Schiöth HB (2007) Identification of novel splice variants of Adhesion G protein-coupled receptors. Gene 387:38–48PubMedCrossRef

33.

Alfano D, Franco P, Vocca I, Gambi N, Pisa V, Mancini A, Caputi M, Carriero MV, Iaccarino I, Stoppelli MP (2005) The urokinase plasminogen activator and its receptor: role in cell growth and apoptosis. Thromb Haemost 93:205–211PubMed

34.

Durand MK, Bodker JS, Christensen A, Dupont DM, Hansen M, Jensen JK, Kjelgaard S, Mathiasen L, Pedersen KE, Skeldal S, Wind T, Andreasen PA (2004) Plasminogen activator inhibitor-I and tumour growth, invasion, and metastasis. Thromb Haemost 91:438–449PubMed

35.

Steeg PS (2003) Metastasis suppressors alter the signal transduction of cancer cells. Nat Rev Cancer 3:55–63PubMedCrossRef

Title: Overexpression of the urokinase receptor mRNA splice variant uPAR-del4/5 affects tumor-associated processes of breast cancer cells in vitro and in vivo
Authors: Sumito Sato
Charlotte Kopitz
Bettina Grismayer
Nathalie Beaufort
Ute Reuning
Manfred Schmitt
Thomas Luther
Matthias Kotzsch
Achim Krüger
Viktor Magdolen
Publication date: 01-06-2011
Publisher: Springer US
Published in: Breast Cancer Research and Treatment / Issue 3/2011
Print ISSN: 0167-6806
Electronic ISSN: 1573-7217
DOI: https://doi.org/10.1007/s10549-010-1042-5

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