TY - JOUR
T1 - uPA-Silica-Particles (SP-uPA)
T2 - A novel analytical system to investigate uPA-uPAR interaction and to test synthetic uPAR antagonists as potential cancer therapeutics
AU - Guthaus, Elke
AU - Bürgle, Marcus
AU - Schmiedeberg, Niko
AU - Hocke, Stefan
AU - Eickler, Alexandra
AU - Kramer, Michael D.
AU - Sweep, C. G.J.Fred
AU - Magdolen, Viktor
AU - Kessler, Horst
AU - Schmitt, Manfred
N1 - Funding Information:
This work was supported by the German Research Association (Deutsche Forschungsgemeinschaft), Sonderforschungsbere-ich SFB 456, B9. We thank Dr. Larry Sklar, University of Albuquerque, USA, for valuable suggestions.
PY - 2002
Y1 - 2002
N2 - The urokinase-type plasminogen activation system, including the serine protease uPA (urokinase-type plasminogen activator) and its cell surface receptor (uPAR, CD87), are important key molecules in tumor invasion and metastasis. Besides its proteolytic function, binding of uPA to uPAR on tumor cells exerts various cell responses such as migration, adhesion, proliferation, and differentiation. Hence, the uPA/uPAR system is a potential target for tumor therapy. We have designed a new generation of uPA-derived synthetic cyclic peptides suited to interfere with the binding of uPA to uPAR and present a new technology involving micro silica particles coated with uPA (SP-uPA) and reacting with recombinant soluble uPAR (suPAR), to rapidly assess the antagonistic potential of uPA-peptides by flow cytofluorometry (FACS). For this, we used silica particles of 10 μm in diameter to which HMW-uPA is coupled using the EDC/NHS method. Soluble, recombinant suPAR was added and the interaction of SP-uPA with suPAR verified by reaction with monoclonal antibody HD13.1 directed to uPAR, followed by a cyan dye (cy5)-labeled antibody directed against mouse IgG. Thereby it was possible to test naturally occurring ligands of uPAR (HMW-uPA, ATF) as well as highly effective, synthetic cyclic uPA-derived peptides cyclo21,29[D-Cys21Nle28Cys29]-uPA21-30, cyclo21,29[D-Cys212-Nal24Cys29]-uPA21-30, and cyclo21,29[D-Cys21Orn23Thi24Thi25Cys29]-uPA21-30. The results obtained with the noncellular SP-uPA/uPAR system are highly comparable to those obtained with a cellular system involving FITC-uPA and the promyeloid cell line U937 as the source of uPAR.
AB - The urokinase-type plasminogen activation system, including the serine protease uPA (urokinase-type plasminogen activator) and its cell surface receptor (uPAR, CD87), are important key molecules in tumor invasion and metastasis. Besides its proteolytic function, binding of uPA to uPAR on tumor cells exerts various cell responses such as migration, adhesion, proliferation, and differentiation. Hence, the uPA/uPAR system is a potential target for tumor therapy. We have designed a new generation of uPA-derived synthetic cyclic peptides suited to interfere with the binding of uPA to uPAR and present a new technology involving micro silica particles coated with uPA (SP-uPA) and reacting with recombinant soluble uPAR (suPAR), to rapidly assess the antagonistic potential of uPA-peptides by flow cytofluorometry (FACS). For this, we used silica particles of 10 μm in diameter to which HMW-uPA is coupled using the EDC/NHS method. Soluble, recombinant suPAR was added and the interaction of SP-uPA with suPAR verified by reaction with monoclonal antibody HD13.1 directed to uPAR, followed by a cyan dye (cy5)-labeled antibody directed against mouse IgG. Thereby it was possible to test naturally occurring ligands of uPAR (HMW-uPA, ATF) as well as highly effective, synthetic cyclic uPA-derived peptides cyclo21,29[D-Cys21Nle28Cys29]-uPA21-30, cyclo21,29[D-Cys212-Nal24Cys29]-uPA21-30, and cyclo21,29[D-Cys21Orn23Thi24Thi25Cys29]-uPA21-30. The results obtained with the noncellular SP-uPA/uPAR system are highly comparable to those obtained with a cellular system involving FITC-uPA and the promyeloid cell line U937 as the source of uPAR.
KW - CD87
KW - Silica particle
KW - Therapeutics
KW - Urokinase
KW - Urokinase receptor
UR - http://www.scopus.com/inward/record.url?scp=0036005974&partnerID=8YFLogxK
U2 - 10.1515/BC.2002.021
DO - 10.1515/BC.2002.021
M3 - Article
C2 - 11930939
AN - SCOPUS:0036005974
SN - 1431-6730
VL - 383
SP - 207
EP - 216
JO - Biological Chemistry
JF - Biological Chemistry
IS - 1
ER -