Small-molecule Inhibition of the Interactions of the Urokinase Receptor: A Targe

Project: Research project

Description

DESCRIPTION (provided by applicant): The urokinase receptor (uPAR) is a GPI-anchored protein that serves as a receptor to the serine protease urokinase (uPA). Upon docking to its receptor, uPA is activated, an event that leads to pericellular proteolysis and degradation of the extracellular matrix, a primary barrier between tumor and vasculature. But it is now widely recognized that while uPAR is not capable of signaling, the uPAR/uPA complex interacts and activates a number of cell surface receptors such as integrins, receptor tyrosine kinases, and GPCRs. The uPAR/uPA interaction has been implicated with nearly every step of tumor invasion and metastasis and is therefore a highly suitable target for the development of anti-cancer therapeutics. Previous efforts have concentrated on inhibiting the serine protease activity of uPA with small molecules. But this strategy neglects interactions of the uPAR/uPA complex with cell signaling proteins that are independent of the proteolytic activity of uPA. In this application, we follow a unique approach that seeks to target the uPAR/uPA protein-protein interaction using small molecules with the expectation that these molecules will exhibit the dual effect of blocking proteolysis and signaling. Our initial efforts have been highly rewarding. A preliminary structure-based computational screen has led to 8 active compounds. Cell culture-based studies reveal that a number of these compounds block MDA-MB-231 tumor cell adhesion, migration, and invasion. Compounds were also found to inhibit lung H1299 tumor cell proliferation. Biochemical analyses reveal that that these compounds also block integrin binding to the uPAR/uPA complex. Our objective in this grant application is to optimize the pharmacokinetic properties and potency of three of these compounds to block tumor invasion and metastasis in vivo. To that end, a multidisciplinary research program involving computational biology, synthetic chemistry, biophysical chemistry, structural biology, and cell biology will be followed. The first aim will consist of creating a virtual combinatorial library of compounds based on our lead molecules, ranking these compounds based on predicted potency, and predicting pharmacokinetic properties of the most potent compounds. The second aim will involve the chemical synthesis of the most promising compounds, followed by an assessment of their activity using a fluorescence polarization assay. The most potent compounds are further characterized with isothermal titration calorimetry. We then solve the three-dimensional structure of these compounds in complex with the urokinase receptor using x-ray diffraction. Finally, the third aim will consist of assessing the cellular efficacy of the most selective and potent inhibitors in MDA-MB-231 and other tumor cell lines, and to perform preliminary in vivo dosage studies to set the stage for future studies in mice xenografts. PUBLIC HEALTH RELEVANCE: The urokinase receptor is a cell surface receptor that binds and activates various proteins. These interactions lead, among other things, to the degradation of a protective layer between a primary tumor and the vasculature, as well as the adhesion and migration of cells. These processes are hallmarks of cancer metastasis. Small molecules with drug-like properties that prevent the urokinase receptor from interacting with its binding partners could lead to highly effective therapeutics to prevent tumor metastasis.
StatusFinished
Effective start/end date5/15/092/28/14

Funding

  • National Institutes of Health: $255,640.00
  • National Institutes of Health: $255,640.00
  • National Institutes of Health: $247,971.00
  • National Institutes of Health: $247,971.00

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Urokinase-Type Plasminogen Activator
Neoplasms
Neoplasm Metastasis
Tumors
Cell Surface Receptors
Serine Proteases
Proteins
Cell Adhesion
Integrins
Proteolysis
Molecules
Cell Movement
Pharmacokinetics
Digital Libraries
Calorimetry
Fluorescence Polarization
Organized Financing
Receptor Protein-Tyrosine Kinases
Computational Biology
Tumor Cell Line

ASJC

  • Medicine(all)