Identification of a novel urokinase receptor splice variant and its prognostic relevance in breast cancer

The cellular receptor for urokinase-type plasminogen activator, uPAR, plays a central role in both cell surface-associated proteolysis and cellular adhesion. In the present study, we systematically searched for splice variants of uPAR mRNA in human cells and tumor tissues by qualitative RT-PCR using...

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Bibliographic Details
Published in:	Thrombosis and haemostasis Vol. 89; no. 4; p. 705
Main Authors:	Luther, Thomas, Kotzsch, Matthias, Meye, Axel, Langerholc, Thomaz, Füssel, Susanne, Olbricht, Natalie, Albrecht, Sybille, Ockert, Detlev, Muehlenweg, Bernd, Friedrich, Katrin, Grosser, Marianne, Schmitt, Manfred, Baretton, Gustavo, Magdolen, Viktor
Format:	Journal Article
Language:	English
Published:	Germany 01-04-2003
Subjects:	Adult Aged Aged, 80 and over Alternative Splicing Animals Antibodies, Monoclonal - metabolism Blotting, Western Breast Neoplasms - diagnosis Breast Neoplasms - genetics Cell Line, Tumor CHO Cells Cricetinae Disease-Free Survival Enzyme-Linked Immunosorbent Assay Exons Female Flow Cytometry Humans Middle Aged Models, Genetic Plasmids - metabolism Plasminogen Activator Inhibitor 1 - metabolism Prognosis Protein Processing, Post-Translational Protein Structure, Tertiary Receptors, Cell Surface - biosynthesis Receptors, Cell Surface - genetics Receptors, Urokinase Plasminogen Activator Reverse Transcriptase Polymerase Chain Reaction RNA, Messenger - metabolism Time Factors Transfection
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Summary:	The cellular receptor for urokinase-type plasminogen activator, uPAR, plays a central role in both cell surface-associated proteolysis and cellular adhesion. In the present study, we systematically searched for splice variants of uPAR mRNA in human cells and tumor tissues by qualitative RT-PCR using specific primers for uPAR exons 1 and 6. Beside the wild-type (wt) uPAR mRNA and the previously described splice variant lacking exon 5 (uPAR-del5), a novel splice variant lacking both exons 4 and 5 (uPAR-del4/5) was found predominantly in various cancer cell lines. To elucidate whether alternatively spliced uPAR mRNA may be translated and post-translationally processed, we generated stably transfected Chinese hamster ovary cells, which harbor expression plasmids of wt uPAR and various uPAR variants including uPAR-del5 and uPAR-del4/5. By ELISA, flow cytofluorometry, and Western blot analysis, we confirmed synthesis and secretion of wt uPAR and the uPAR variants by the use of domain-specific monoclonal antibodies against uPAR. For quantification of uPAR mRNA variants, we established two highly sensitive real-time RT-PCR assays based on LightCycler technology. Study of their expression in a representative set of breast cancer tissues indicated that the novel mRNA variant uPAR-del4/5 was expressed very frequently and independently of uPAR mRNA variants covering exon 4 (uPAR-wt and uPAR-del5). Higher uPAR-del4/5 expression was significantly associated with shorter disease-free survival (p = 0.0004) of breast cancer patients. These results suggest that uPAR-del4/5 mRNA may serve as a novel prognostic marker in breast cancer.
ISSN:	0340-6245
DOI:	10.1055/s-0037-1613577