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.
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.
|Zhou, Donghui; Springer, Maya Z; Xu, David et al. (2017) Small molecules inhibit STAT3 activation, autophagy, and cancer cell anchorage-independent growth. Bioorg Med Chem 25:2995-3005|
|Xu, David; Li, Liwei; Zhou, Donghui et al. (2017) Structure-Based Target-Specific Screening Leads to Small-Molecule CaMKII Inhibitors. ChemMedChem 12:660-677|
|Liu, Degang; Xu, David; Liu, Min et al. (2017) Small Molecules Engage Hot Spots through Cooperative Binding To Inhibit a Tight Protein-Protein Interaction. Biochemistry 56:1768-1784|
|Xu, David; Jalal, Shadia I; Sledge, George W et al. (2016) Small-molecule binding sites to explore protein-protein interactions in the cancer proteome. Mol Biosyst 12:3067-87|
|Xu, David; Meroueh, Samy O (2016) Effect of Binding Pose and Modeled Structures on SVMGen and GlideScore Enrichment of Chemical Libraries. J Chem Inf Model 56:1139-51|
|Liu, Degang; Zhou, Donghui; Wang, Bo et al. (2015) A new class of orthosteric uPAR·uPA small-molecule antagonists are allosteric inhibitors of the uPAR·vitronectin interaction. ACS Chem Biol 10:1521-34|
|Wang, Bo; Buchman, Cameron D; Li, Liwei et al. (2014) Enrichment of chemical libraries docked to protein conformational ensembles and application to aldehyde dehydrogenase 2. J Chem Inf Model 54:2105-16|
|Peng, Xiaodong; Wang, Fang; Li, Liwei et al. (2014) Exploring a structural protein-drug interactome for new therapeutics in lung cancer. Mol Biosyst 10:581-91|
|Mani, Timmy; Wang, Fang; Knabe, William Eric et al. (2013) Small-molecule inhibition of the uPAR·uPA interaction: synthesis, biochemical, cellular, in vivo pharmacokinetics and efficacy studies in breast cancer metastasis. Bioorg Med Chem 21:2145-55|
|Mani, Timmy; Liu, Degang; Zhou, Donghui et al. (2013) Probing binding and cellular activity of pyrrolidinone and piperidinone small molecules targeting the urokinase receptor. ChemMedChem 8:1963-77|
Showing the most recent 10 out of 20 publications