Antagonistic Anti-urokinase Plasminogen Activator Receptor (uPAR) Antibodies Significantly Inhibit uPAR-mediated Cellular Signaling and Migration

Department of Pharmaceutical Chemistry, University of California, San Francisco, California 94158-2517, USA.
Journal of Biological Chemistry (Impact Factor: 4.57). 08/2010; 285(35):26878-88. DOI: 10.1074/jbc.M109.077677
Source: PubMed

ABSTRACT Interactions between urokinase plasminogen activator receptor (uPAR) and its various ligands regulate tumor growth, invasion, and metastasis. Antibodies that bind specific uPAR epitopes may disrupt these interactions, thereby inhibiting these processes. Using a highly diverse and naïve human fragment of the antigen binding (Fab) phage display library, we identified 12 unique human Fabs that bind uPAR. Two of these antibodies compete against urokinase plasminogen activator (uPA) for uPAR binding, whereas a third competes with beta1 integrins for uPAR binding. These competitive antibodies inhibit uPAR-dependent cell signaling and invasion in the non-small cell lung cancer cell line, H1299. Additionally, the integrin-blocking antibody abrogates uPAR/beta1 integrin-mediated H1299 cell adhesion to fibronectin and vitronectin. This antibody and one of the uPAR/uPA antagonist antibodies shows a significant combined effect in inhibiting cell invasion through Matrigel/Collagen I or Collagen I matrices. Our results indicate that these antagonistic antibodies have potential for the detection and treatment of uPAR-expressing tumors.

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Available from: Charles S Craik, Sep 26, 2015
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    • "Cell Line Studies The clonogenic survival and matrigel invasion assays were conducted as previously described 19, 20. For the qPCR analysis RNA was prepared from each cell line (~ 2 x 106 cells/cell line) using an RNEasy kit (Qiagen). "
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    Theranostics 01/2014; 4(3):267-79. DOI:10.7150/thno.7323 · 8.02 Impact Factor
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    • "Recently, 12 unique human Fabs that bind human uPAR were identified through a highly diverse and naive human fragment of the antigen binding (Fab) phage display library 36. Among them, two Fabs (2E9 and 2G10) could compete with uPA for uPAR binding and one Fab (3C6) competes α5β1 integrin for uPAR binding 36. Both ATN-658 and these Fabs are suitable for uPAR-targeted imaging and therapy. "
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    ABSTRACT: Urokinase-type plasminogen activator receptor (uPAR) is a glycosylphosphatidylinositol (GPI)-anchored protein. Besides regulating proteolysis, uPAR could also activate many intracellular signaling pathways that promote cell motility, invasion, proliferation, and survival through cooperating with transmembrane receptors. uPAR is overexpressed across a variety of tumors and is associated with cancer invasion and metastasis. In order to meet the demand for a rapid development and potential clinical application of anti-cancer therapy based on uPA/uPAR system, it is desirable to develop non-invasive imaging methods to visualize and quantify uPAR expression in vivo. In this review, we will discuss recent advances in the development of uPAR-targeted nuclear imaging and radionuclide therapy agents. The successful development of molecular imaging probes to visualize uPAR expression in vivo would not only assist preclinical researches on uPAR function, but also eventually impact patient management.
    Theranostics 06/2013; 3(7):507-515. DOI:10.7150/thno.5557 · 8.02 Impact Factor
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    • "Despite these encouraging findings, the full theranostic potential of this targeting system still needs further validation, since the species-selectivity inherent to this targeting peptide 52 precludes bona fide toxicity assessment as it leaves the host stromal compartment essentially untouched in these xenograft mouse models thereby attenuating the general toxicity of this targeted radiotherapy. A similar precaution should, nonetheless, also have been applied to a recent study, where the efficacy and translational potential of a 177Lu-conjugated recombinant antibody, which binds human uPAR with moderate affinity (KD ~ 10-40 nM) but not mouse uPAR, was evaluated as a uPAR-targeting radiotherapeutic in an orthotopic mammary carcinoma xenograft model in nude mice 62, 63. Unfortunately, this study also lacks the imperative control using an irrelevant 177Lu-labelled mAb to demonstrate the specificity of the targeted ionization-induced insult in vivo. "
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    ABSTRACT: Research performed during the last two decades has provided a wealth of information to highlight the role of the urokinase-type plasminogen activator receptor (uPAR) in the progression and dissemination of invasive and metastatic cancer. In parallel, our perception of the structure-function relationships in uPAR has been refined to such a level that a rational design of uPAR function as well as compounds specifically targeting defined functions of uPAR are now realistic options. This knowledge opens new avenues for developing therapeutic intervention regimens targeting uPAR as well as for monitoring the effects of such treatments by non-invasive imaging using e.g. positron emission tomography. This mini-review will focus on recent advancements in translational research devoted to non-invasive targeting of uPAR, with a view to molecular imaging of its expression in live individuals as well as specific eradication of these cells by targeted radiotherapy.
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