Design and Synthesis of Fragment Derivatives with a Unique Inhibition Mechanism of the UPAR·uPA Interaction

Overview

Journal ACS Med Chem Lett

Publisher American Chemical Society

Specialty Chemistry

Date 2021 Jan 25

PMID 33488965

Citations 4

Authors

Khuchtumur Bum-Erdene

Degang Liu

David Xu

Mona K Ghozayel

Samy O Meroueh

Affiliations

Soon will be listed here.

Abstract

There is substantial interest in the development of small molecules that inhibit the tight and highly challenging protein-protein interaction between the glycophosphatidylinositol (GPI)-anchored cell surface receptor uPAR and the serine protease uPA. While preparing derivatives of a fragment-like compound that previously emerged from a computational screen, we identified compound (IPR-3242), which inhibited binding of uPA to uPAR with submicromolar ICs. The high inhibition potency prompted us to carry out studies to rule out potential aggregation, lack of stability, reactivity, and nonspecific inhibition. We designed and prepared 16 derivatives to further explore the role of each substituent. Interestingly, the compounds only partially inhibited binding of a fluorescently labeled α-helical peptide that binds to uPAR at the uPAR·uPA interface. Collectively, the results suggest that the compounds bind to uPAR outside of the uPAR·uPA interface, trapping the receptor into a conformation that is not able to bind to uPA. Additional studies will have to be carried out to determine whether this unique inhibition mechanism can occur at the cell surface.

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References

Mani T, Wang F, Knabe W, Sinn A, Khanna M, Jo I . Small-molecule inhibition of the uPAR·uPA interaction: synthesis, biochemical, cellular, in vivo pharmacokinetics and efficacy studies in breast cancer metastasis. Bioorg Med Chem. 2013; 21(7):2145-55. PMC: 3625246. DOI: 10.1016/j.bmc.2012.12.047. View

Huai Q, Zhou A, Lin L, Mazar A, Parry G, Callahan J . Crystal structures of two human vitronectin, urokinase and urokinase receptor complexes. Nat Struct Mol Biol. 2008; 15(4):422-3. PMC: 2443823. DOI: 10.1038/nsmb.1404. View

Berkenblit A, Matulonis U, Kroener J, Dezube B, Lam G, Cuasay L . A6, a urokinase plasminogen activator (uPA)-derived peptide in patients with advanced gynecologic cancer: a phase I trial. Gynecol Oncol. 2005; 99(1):50-7. DOI: 10.1016/j.ygyno.2005.05.023. View

Ignar D, Andrews J, WITHERSPOON S, Leray J, Clay W, Kilpatrick K . Inhibition of establishment of primary and micrometastatic tumors by a urokinase plasminogen activator receptor antagonist. Clin Exp Metastasis. 1998; 16(1):9-20. DOI: 10.1023/a:1006503816792. View

Guo Y, Higazi A, Arakelian A, Sachais B, Cines D, Goldfarb R . A peptide derived from the nonreceptor binding region of urokinase plasminogen activator (uPA) inhibits tumor progression and angiogenesis and induces tumor cell death in vivo. FASEB J. 2000; 14(10):1400-10. DOI: 10.1096/fj.14.10.1400. View

Wei Y, Eble J, Wang Z, Kreidberg J, Chapman H . Urokinase receptors promote beta1 integrin function through interactions with integrin alpha3beta1. Mol Biol Cell. 2001; 12(10):2975-86. PMC: 60149. DOI: 10.1091/mbc.12.10.2975. View

Huai Q, Mazar A, Kuo A, Parry G, Shaw D, Callahan J . Structure of human urokinase plasminogen activator in complex with its receptor. Science. 2006; 311(5761):656-9. DOI: 10.1126/science.1121143. View

Gardsvoll H, Gilquin B, Le Du M, Menez A, Jorgensen T, Ploug M . Characterization of the functional epitope on the urokinase receptor. Complete alanine scanning mutagenesis supplemented by chemical cross-linking. J Biol Chem. 2006; 281(28):19260-72. DOI: 10.1074/jbc.M513583200. View

Xu D, Bum-Erdene K, Si Y, Zhou D, Ghozayel M, Meroueh S . Mimicking Intermolecular Interactions of Tight Protein-Protein Complexes for Small-Molecule Antagonists. ChemMedChem. 2017; 12(21):1794-1809. PMC: 6067920. DOI: 10.1002/cmdc.201700572. View

10.

Mani T, Liu D, Zhou D, Li L, Knabe W, Wang F . Probing binding and cellular activity of pyrrolidinone and piperidinone small molecules targeting the urokinase receptor. ChemMedChem. 2013; 8(12):1963-77. PMC: 4058332. DOI: 10.1002/cmdc.201300340. View

11.

Gardsvoll H, Ploug M . Mapping of the vitronectin-binding site on the urokinase receptor: involvement of a coherent receptor interface consisting of residues from both domain I and the flanking interdomain linker region. J Biol Chem. 2007; 282(18):13561-72. DOI: 10.1074/jbc.M610184200. View

12.

Irwin J, Duan D, Torosyan H, Doak A, Ziebart K, Sterling T . An Aggregation Advisor for Ligand Discovery. J Med Chem. 2015; 58(17):7076-87. PMC: 4646424. DOI: 10.1021/acs.jmedchem.5b01105. View

13.

Madsen C, Sarra Ferraris G, Andolfo A, Cunningham O, Sidenius N . uPAR-induced cell adhesion and migration: vitronectin provides the key. J Cell Biol. 2007; 177(5):927-39. PMC: 2064291. DOI: 10.1083/jcb.200612058. View

14.

Mertens H, Kjaergaard M, Mysling S, Gardsvoll H, Jorgensen T, Svergun D . A flexible multidomain structure drives the function of the urokinase-type plasminogen activator receptor (uPAR). J Biol Chem. 2012; 287(41):34304-15. PMC: 3464537. DOI: 10.1074/jbc.M112.398404. View

15.

Zhao B, Gandhi S, Yuan C, Luo Z, Li R, Gardsvoll H . Stabilizing a flexible interdomain hinge region harboring the SMB binding site drives uPAR into its closed conformation. J Mol Biol. 2015; 427(6 Pt B):1389-1403. DOI: 10.1016/j.jmb.2015.01.022. View

16.

Wei Y, Lukashev M, Simon D, Bodary S, Rosenberg S, Doyle M . Regulation of integrin function by the urokinase receptor. Science. 1996; 273(5281):1551-5. DOI: 10.1126/science.273.5281.1551. View

17.

Bum-Erdene K, Zhou D, Gonzalez-Gutierrez G, Ghozayel M, Si Y, Xu D . Small-Molecule Covalent Modification of Conserved Cysteine Leads to Allosteric Inhibition of the TEAD⋅Yap Protein-Protein Interaction. Cell Chem Biol. 2018; 26(3):378-389.e13. DOI: 10.1016/j.chembiol.2018.11.010. View

18.

Khanna M, Wang F, Jo I, Eric Knabe W, Wilson S, Li L . Targeting multiple conformations leads to small molecule inhibitors of the uPAR·uPA protein-protein interaction that block cancer cell invasion. ACS Chem Biol. 2011; 6(11):1232-43. PMC: 3220747. DOI: 10.1021/cb200180m. View

19.

Llinas P, Le Du M, Gardsvoll H, Dano K, Ploug M, Gilquin B . Crystal structure of the human urokinase plasminogen activator receptor bound to an antagonist peptide. EMBO J. 2005; 24(9):1655-63. PMC: 1142576. DOI: 10.1038/sj.emboj.7600635. View

20.

Wang F, Eric Knabe W, Li L, Jo I, Mani T, Roehm H . Design, synthesis, biochemical studies, cellular characterization, and structure-based computational studies of small molecules targeting the urokinase receptor. Bioorg Med Chem. 2012; 20(15):4760-73. PMC: 3437670. DOI: 10.1016/j.bmc.2012.06.002. View