Abstract
Purpose
To evaluate the pure prognostic impact of the uPA-receptor splice variant uPAR-del4/5 for lymph node-negative breast cancer patients, and to identify differentially expressed genes associated with high or low uPAR-del4/5 mRNA levels.
Patients and methods
mRNA transcript levels were measured by real-time PCR in tumor samples from 280 node-negative breast cancer patients who had not received adjuvant systemic therapy. Endpoints were distant metastasis-free survival (DMFS) and overall survival (OS). Gene expression analysis was performed with RNA isolated from breast cancer tissue and breast cancer cell lines using Affymetrix U133a GeneChips.
Results
In multivariate analysis, uPAR-del4/5 significantly contributed to the base model of traditional prognostic factors for DMFS (HR = 3.29, P < 0.001) and OS (HR = 2.87, P = 0.002). Using microarrays, seven genes were found to be up-regulated in tumor samples and cancer cell lines with high uPAR-del4/5 mRNA expression. The gene encoding rab31, a member of the Ras oncogene family, was selected for quantitative analysis of mRNA expression in the set of 280 patients. High rab31 values were significantly associated with worse outcome of patients for DMFS (HR = 2.27, P < 0.001) and OS (HR = 2.01, P = 0.008) in multivariate analysis, independent from uPAR-del4/5. The patient subgroup with high uPAR-del4/5 and rab31 levels showed the worst DMFS and OS (P < 0.001, both) compared with tumors with low values of both factors.
Conclusions
Our results suggest that uPAR-del4/5 and rab31 mRNA represent independent prognostic markers in breast cancer and may be components of different, but possibly associated, tumor-relevant signaling pathways.
Similar content being viewed by others
References
Dano K, Behrendt N, Hoyer-Hansen G et al (2005) Plasminogen activation and cancer. Thromb Haemost 93:676–681
Andreasen PA, Egelund R, Petersen HH (2000) The plasminogen activation system in tumor growth, invasion, and metastasis. Cell Mol Life Sci 57:25–40
Reuning U, Magdolen V, Hapke S et al (2003) Molecular and functional interdependence of the urokinase-type plasminogen activator system with integrins. Biol Chem 384:1119–1131
Blasi F, Carmeliet P (2002) uPAR: a versatile signaling orchestrator. Nat Rev Mol Cell Biol 3:932–943
Ragno P (2006) The urokinase receptor: a ligand or a receptor? Story of a sociable molecule. Cell Mol Life Sci 63:1028–1037
Duffy MJ (2004) The urokinase plasminogen activator system: role in malignancy. Curr Pharm Des 10:39–49
Harbeck N, Kates RE, Gauger K et al (2004) Urokinase-type plasminogen activator (uPA) and its inhibitor PAI-1: novel tumor-derived factors with a high prognostic and predictive impact in breast cancer. Thromb Haemost 91:450–456
Foekens JA, Peters HA, Look MP et al (2000) The urokinase system of plasminogen activation and prognosis in 2780 breast cancer patients. Cancer Res 60:636–643
Kotzsch M, Luther T, Harbeck N et al (2000) New ELISA for quantification of human urokinase receptor (CD87) in cancer. Int J Oncol 17:827–834
De Witte JH, Foekens JA, Brünner N et al (2001) Prognostic impact of urokinase-type plasminogen activator receptor (uPAR) in cytosols and pellet extracts derived from primary breast tumours. Br J Cancer 85:85–92
de Bock CE, Wang Y (2004) Clinical significance of urokinase-type plasminogen activator receptor (uPAR) expression in cancer. Med Res Rev 24:13–39
Duffy MJ, Duggan C (2004) The urokinase plasminogen activator system: a rich source of tumour markers for the individualised management of patients with cancer. Clin Biochem 37:541–548
Llinas P, Le Du MH, Gardsvoll H et al (2005) Crystal structure of the human urokinase plasmi-nogen activator receptor bound to an antagonist peptide. EMBO J 24:1655–1663
Huai Q, Mazar AP, Kuo A et al (2006) Structure of human urokinase plasminogen activator in complex with its receptor. Science 311:656–659
Montouri N, Visconte V, Rossi G et al (2005) Soluble and cleaved forms of the urokinase-receptor: degradation products or active molecules? Thromb Haemost 93:192–198
Luther T, Kotzsch M, Meye A et al (2003) Identification of a novel urokinase receptor splice variant and its prognostic relevance in breast cancer. Thromb Haemost 89:705–717
Kotzsch M, Farthmann J, Meye A et al (2005) Prognostic relevance of uPAR-del4/5 and TIMP-3 mRNA expression levels in breast cancer. Eur J Cancer 41:2760–2768
Wang Y, Klijn JG, Zhang Y et al (2005) Gene-expression profiles to predict distant metastasis of lymph-node-negative primary breast cancer. Lancet 365:671–679
Sieuwerts AM, Meijer-van Gelder ME et al (2005) How ADAM-9 and ADAM-11 differentially from estrogen receptor predict response to tamoxifen treatment in patients with recurrent breast cancer: a retrospective study. Clin Cancer Res 11:7311–7321
Tusher VG, Tibshirani R, Chu G (2001) Significance analysis of microarrays applied to the ionizing radiation response. Proc Natl Acad Sci U S A 98:5116–5121
Barlow RE, Bartholomew DJ, Bremmer JM et al (1972) Statistical interference under order restrictions. Wiley, New York
Foekens JF, Schmitt M, van Putten WL et al (1994) Plasminogen activator inhibitor-1 and prognosis in primary breast cancer. J Clin Oncol 12:1648–1658
Cheng JM, Lahad JP, Kuo WL et al (2004) The RAB25 small GTPase determines aggressiveness of ovarian and breast cancers. Nature Med 10:1251–1256
Buyse M, Loi S, van’t Veer L et al (2006) Validation and clinical utility of a 70-gene prognostic signature for women with node-negative breast cancer. J Natl Cancer Inst 98:1183–1192
Braun S, Cevatli BS, Assemi C et al (2001) Comparative analysis of micrometastasis to the bone marrow and lymph nodes of node-negative breast cancer patients receiving no adjuvant therapy. J Clin Oncol 19:1468–1475
Schmitt M, Wilhelm OG, Reuning U et al (2000) The urokinase plasminogen activator system as a novel target for tumour therapy. Fibrinol Proteol 14:114–1132
Bouchet C, Hacene K, Martin PM et al (1999) Dissemination risk index based on plasminogen activator system components in primary breast cancer. J Clin Oncol 17:3048–3057
Guyton DP, Evans DM, Sloan-Stakleff KD (2000) Urokinase plasminogen activator receptor (uPAR): A potential indicator of invasion for in situ breast cancer. Breast J 6:130–136
Casey JR, Petranka JG, Kottra J et al (1994) The structure of the urokinase-type plasminogen activator receptor gene. Blood 84:1151–1156
Pyke C, Eriksen J, Solberg H et al (1993) An alternatively spliced variant of mRNA for the human receptor for urokinase plasminogen activator. FEBS Lett 326:69–74
Stephens RW, Nielsen HJ, Christensen IJ et al (1999) Plasma urokinase receptor levels in patients with colorectal cancer: relationship to prognosis. J Natl Cancer Inst 91:869–874
Riisbro R, Christensen IB, Piironen T et al (2002) Prognostic significance of soluble urokinase plasminogen activator receptor in serum and cytosol of tumor tissue from patients with primary breast cancer. Clin Cancer Res 8:1132–1141
Grebenchtchikov N, Maguire TM, Riisbro R et al (2005) Measurement of plasminogen activator system components in plasma and tumor tissue extracts obtained from patients with breast cancer: an EORTC Receptor and Biomarker Group collaboration. Oncol Rep 14:235–239
Farthmann J, Holzscheiter L, Biermann J et al (2004) Development of quantitative RT-PCR assays for wild-type urokinase receptor (uPAR-wt) and its splice variant uPAR-del5. Radiol Oncol 38:111–119
Pacheco MM, Nishimoto IN, Mourao et al (2001) Prognostic significance of the combined expression of matrix metalloproteinase-9, urokinase type plasminogen activator and its recep-tor in breast cancer as measured by Northern blot analysis. Int J Biol Markers 16:62–68
Bao X, Faris AE, Jang EK et al (2002) Molecular cloning, bacterial expression and properties of Rab31 and Rab32. Eur J Biochem 269:259–271
Zerial M, McBride H (2001) Rab proteins as membrane organizers. Nat Rev Mol Cell Biol 2:107–117
Jones MC, Caswell PT, Norman JC (2006) Endocytic recycling pathways: emerging regulators of cell migration. Curr Opin Cell Biol 18:549–557
Ceresa BP (2006) Regulation of EGFR endocytic trafficking by rab proteins. Histol Histopathol 21:987–993
Pellinen T, Arjonen A, Vuoriluoto K et al (2006) Small GTPase Rab21 regulates cell adhesion and controls endosomal traffic of ß1-integrins. J Cell Biol 173:767–780
Wei Y, Lukashev M, Simon DI et al (1996) Regulation of integrin function by the urokinase receptor. Science 273:1551–1555
Liu D, Aguirre Ghiso J, Estrada Y et al (2002) EGFR is a transducer of the urokinase receptor initiated signal that is required for in vivo growth of human carcinoma. Cancer Cell 1:445–457
Mazzieri R, D’Alessio S, Kenmoe RK et al (2006) An uncleavable uPAR mutant allows dissection of signaling pathways in uPA-dependent cell migration. Mol Biol Cell 17:367–378
Cheng JM, Lahad JP, Gray JW et al (2005) Emerging role of RAB GTPases in cancer and human disease. Cancer Res 65:2516–2519
Calvo A, Xiao N, Kang J et al (2002) Alterations in gene expression profiles during prostate cancer progression: functional correlations to tumorigenicity and down-regulation of selenoprotein-P in mouse and human tumors. Cancer Res 62:5325–5335
Goldenring JR, Ray GS, Lee JR (1999) Rab11 in dysplasia of Barrett’s epithelia. Yale J Biol Med 72:113–120
He H, Dai F, Yu L et al (2002) Identification and characterization of nine novel human small GTPases showing variable expression in liver cancer tissue. Gene Expr 10:231–242
Pellagatti A, Esoof N, Watkins F et al (2004) Gene expression profiling in the myelodysplastic syndromes using microarray technology. Br J Haematol 125:576–583
Amillet JM, Ferbus D, Real FX et al (2006) Characterization of human Rab20 overexpressed in exocrine pancreatic carcinoma. Human Pathol 37:256–263
Abba MC, Hu Y, Sun H et al (2005) Gene expression signature of estrogen receptor status in breast cancer. BMC Genomics 6:37
Acknowledgements
This study was supported by grants from the Deutsche Krebshilfe e.V., Germany (Grant No. 106 185). We thank Antje Zobjack, Annelie Zürich, and Andrea Lohse for their excellent technical assistance.
Author information
Authors and Affiliations
Corresponding author
Electronic supplementary material
Rights and permissions
About this article
Cite this article
Kotzsch, M., Sieuwerts, A.M., Grosser, M. et al. 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, 229–240 (2008). https://doi.org/10.1007/s10549-007-9782-6
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10549-007-9782-6