The urokinase pathway of plasminogen activation is thought to play an important role in extracellular matrix degradation during the process of breast cancer cell invasion. In this system,
urokinase (uPA) converts the ubiquitous protein proenzyme plasminogen into plasmin which, in turn, can degrade a wide range of proteins. Plasminogen activation is thought to occur on the cell surface, where a specific
urokinase receptor (uPAR) localises and enhances
uPA activity. The activation of plasminogen is also regulated by the specific plasminogen activator inhibitors (PAI)
PAI-1 and
PAI-2 (Li Y. et al., 1999).
Cell lines:
- By flow cytometry, 2500 uPAR/cell were found on normal mammary epithelial cells; in 6 breast cancer cell (BCC) lines (MCF-7, MDA-MB-361, T-47D, MDA-MB-231, BT-20, Hs578T), the cell surface uPAR number ranged between ~13,700 and ~58,800 sites per cell (Li Y. et al., 1999).
- In MDA-MB-231 BCC, both
thrombospondin-1 and transforming growth factor-ß1 up-regulated uPAR expression, and promoted tumour cell invasion (Albo D. et al., 1997), and a monoclonal antibody specific for uPAR has been shown to inhibit invasion by this BCC line (Holst-Hansen C. et al., 1996).
- In MCF-7 BCC, binding of
uPA to uPAR activates extracellular signal-regulated kinases 1 and 2 which are required for increased cellular motility (Nguyen D.H.D. et al., 1998).
- MCF-7 BCC were shown to exhibit a
urokinase-dependent physical association between uPAR and the vitronectin receptor
alphaVbeta5. These BCC responded to
urokinase or to its noncatalytic amino-terminal fragment by exhibiting remarkable cytoskeletal rearrangements that were mediated by
alphaVbeta5 and required protein kinase C activity. On the contrary, binding of vitronectin to
alphaVbeta5 resulted in the protein kinase C-independent formation of F-actin containing microspike-type structures. Furthermore,
alphaVbeta5 was required for
urokinase-directed, receptor-dependent MCF-7 BCC migration (Carriero M.V. et al., 1999).
-
uPA was shown to promote MCF-7 BCC migration on vitronectin-coated surfaces, in an integrin (a
beta1-integrin -probably
alphaVbeta1-, and
alphaVbeta5)-selective manner, by initiating a uPAR-dependent signaling cascade in which Ras, MAP kinase kinase (MEK), extracellular signal-regulated kinase (ERK), and myosin light chain kinase (MLCK) serve as essential downstream effectors (Nguyen D.H. et al., 1999).
- Culture of MDA-MB-231 BCC under hypoxia resulted in increased uPAR mRNA levels. Increased uPAR expression was paralleled by higher cell-associated
uPA levels and lower levels of secreted
uPA as determined by gel zymography performed on cell extracts and culture-conditioned media. In addition, the in vitro invasiveness of MDA-MB-231 breast carcinoma cells was significantly higher when the invasion assay was performed under hypoxic conditions. This effect of hypoxia on invasion was abrogated by including in the assay a monoclonal, function-blocking anti-uPAR antibody or by the presence of 30% carbon monoxide in the hypoxic atmosphere (Graham C.H. et al., 1999).
- Treatment of highly invasive BT549 BCC with a specific p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580 diminished both
uPA/uPAR mRNA and protein expression and abrogated the ability of these BCC to invade matrigel, suggesting that p38 MAPK signaling pathway is involved in the regulation of
uPA/uPAR expression and breast cancer cell invasion. SB203580-induced reduction in
uPA/uPAR mRNA expression was shown to result from the de-stabilization of
uPA and uPAR mRNA. Finally, it was demonstrated that the p38alpha, rather than the p38beta, MAPK isoform activity was essential for
uPA/uPAR expression. Thus, p38alpha MAPK signaling pathway might be important for the maintenance of breast cancer invasive phenotype by promoting the stabilities of
uPA and uPAR mRNA (Huang S. et al., 2000).
Tumors:
-
uPA, its receptor uPAR, and
PAI-1 were measured in breast cancer cytosol from 111 low-risk premenopausal patients and 184 low-risk postmenopausal patients with a median follow-up of 6.0 years.
uPA, uPAR, and
PAI-1 levels were all weakly but significantly correlated with each other in both menopausal groups. There were no significant differences in the median levels of
uPA, uPAR, and
PAI-1 between premenopausal and postmenopausal patients (Grøndahl-Hansen J. et al., 1997).
-
cathepsin D (n=162),
uPA (n=116), uPAR (n=109) and
PAI-1 (n=135) were measured in tumor cytosols obtained from a population of node negative breast cancer patients. A significant correlation was found between levels of
uPA, uPAR, and
PAI-1. Levels of
cathepsin D were directly related to levels of
uPA and uPAR (Kute T.E. et al., 1998).
-
uPA and uPAR levels were semiquantitated by immunocytochemistry in 36 primary breast carcinomas. Both were mostly present in stromal cells in invasive breast carcinomas, suggesting that stromal cells collaborate with malignant cells to mediate metastasis (Kennedy S. et al., 1998).
- Studies have indicated that uPAR could be associated in large molecular complexes with other molecules, such as integrins. In a study of 10 human breast carcinomas, the ability of uPAR to physically associate with the vitronectin receptor
alphaVbeta5 was shown (Carriero M.V. et al., 1999).
- Tumor-associated macrophages of invasive breast carcinomas and of ductal carcinoma
in situ were found to possess significantly elevated uPAR levels compared with macrophages derived from normal breast tissue (Hildenbrand R. et al., 1999).
- uPAR mRNA and protein expression was studied by in-situ hybridization and immunohistochemistry, respectively, in 50 formalin-fixed, paraffin-embedded specimens of ductal carcinoma in situ (DCIS). Three different antibodies were used to stain cell-associated uPAR; chicken polyclonal antibody (pAb) HU277 and monoclonal antibodies (mAb) IID7 and 3936. In all cases, myoepithelial and stromal cells reacted with either antibody. Especially, reaction of macrophage-like cells with mAb 3936 resulted in a well-marked and bright staining. Applying mAb IID7, in 46 of the 50 breast specimens tumour cells showed a positive immunoreaction. Likewise pAb HU277 stained tumour cells in 40 of the 50 cases, whereas mAb 3936 reacted with only 24 of the 50 tissue sections. Endothelial cells were marked by both mAb IID7 and pAb HU277 (46/50 and 35/50, respectively); mAb 3936 did not label at all. All of the cell types stained by mAb IID7 and pAb HU277 also displayed reactivity with uPAR mRNA-specific antisense oligonucleotides in in-situ hybridization (Hildenbrand R. et al., 2000).
Albo D. et al. (1997)
Thrombospondin-1 and transforming growth factor-ß1 promote breast tumour cell invasion through up-regulation of the plasminogen/plasmin system. Surgery 122, 493-499.
Andreasen P.A. et al. (1997) The
urokinase-type plasminogen activator system in cancer metastasis: a review. Int J. Cancer 72, 1-22 (
Review).
Carriero M.V. et al. (1999)
Urokinase receptor interacts with
alpha(v)beta5 vitronectin receptor, promoting
urokinase-dependent cell migration in breast cancer. Cancer Res. 59, 5307-5314.
Graham C.H. et al. (1999) Hypoxia-mediated stimulation of carcinoma cell invasiveness via upregulation of urokinase receptor expression. Int. J. Cancer 80, 617-623.
Grøndahl-Hansen J. et al. (1997)
Plasminogen activator inhibitor type 1 in cytosolic tumor extracts predicts prognosis in low-risk breast cancer patients. Clin. Cancer Res. 3, 233-239.
Hildenbrand R. et al. (1999) Urokinase plasminogen activator receptor (CD87) expression of tumor-associated macrophages in ductal carcinoma in situ, breast cancer, and resident macrophages of normal breast tissue. J. Leukoc. Biol. 66, 40-49.
Hildenbrand R. et al. (2000) Validation of immunolocalization of the urokinase receptor expression in ductal carcinoma in situ of the breast: comparison with detection by non-isotopic in-situ hybridization. Histopathology 36, 499-504.
Holst-Hansen C. et al. (1996)
Urokinase-type plasminogen activation in three human breast cancer cell lines correalte with their
in vitro invasiveness. Clin. Exp. Metastasis 14, 297-307.
Huang S. et al. (2000)
Urokinase plasminogen activator/urokinase-specific surface receptor expression and matrix invasion by breast cancer cells requires constitutive p38alpha mitogen-activated protein kinase activity. J. Biol. Chem. 275, 12266-12272.
Kennedy S. et al. (1998) Semi-quantitation of
urokinase plasminogen activator and its receptor in breast carcinomas by immunocytochemistry. Br. J. Cancer 77, 1638-1641.
Kute T.E. et al. (1998) Low
cathepsin D and low plasminogen activator type 1 inhibitor in tumor cytosols defines a group of node negative breast cancer patients with low risk of recurrence. Breast Cancer Res. Treat. 47, 9-16.
Li Y. et al. (1999) Cell surface expression of urokinase receptor in normal mammary epithelial cells and breast cancer cell lines. Anticancer Res. 19, 1223-1228.
Nguyen D.H.D. et al. (1998) Binding of
urokinase-type plasminogen activator to its receptor in MCF-7 cells activates extracellular signal-regulated kinases 1 and 2 which is required for cellular motility. J. Biol. Chem. 273, 8502-8507.
Nguyen D.H. et al. (1999) Myosin light chain kinase functions downstream of Ras/ERK to promote migration of
urokinase-type plasminogen activator-stimulated cells in an integrin-selective manner. J. Cell. Biol. 146, 149-164.
Roldan A.L. et al. (1990) Cloning and expression of the receptor for human
urokinase plasminogen activator, a central molecule in cell surface, plasmin dependent proteolysis. EMBO J. 9, 467-474.
Vagnarelli P. et al. (1992) Assignment of the human urokinase receptor gene (PLAUR) to 19q13. Cytogenet. Cell Genet. 60, 197-199.
