Receptor binding mode and pharmacological characterization of a potent and selective dual CXCR1/CXCR2 non-competitive allosteric inhibitor

Background and Purpose: DF 2156A is a new dual inhibitor of IL-8 receptors CXCR1 and CXCR2 with an optimal pharmacokinetic profile. We characterized its binding mode, molecular mechanism of action and selectivity, and evaluated its therapeutic potential. Experimental Approach: The binding mode, molecular mechanism of action and selectivity were investigated using chemotaxis of L1.2 transfectants and human leucocytes, in addition to radioligand and [ 35S]-GTPγS binding approaches. The therapeutic potential of DF 2156A was evaluated in acute (liver ischaemia and reperfusion) and chronic (sponge-induced angiogenesis) experimental models of inflammation. Key Results: A network of polar interactions stabilized by a direct ionic bond between DF 2156A and Lys99 on CXCR1 and the non-conserved residue Asp 293 on CXCR2 are the key determinants of DF 2156A binding. DF 2156A acted as a non-competitive allosteric inhibitor blocking the signal transduction leading to chemotaxis without altering the binding affinity of natural ligands. DF 2156A effectively and selectively inhibited CXCR1/CXCR2-mediated chemotaxis of L1.2 transfectants and leucocytes. In a murine model of sponge-induced angiogenesis, DF 2156A reduced leucocyte influx, TNF-α production and neovessel formation. In vitro, DF 2156A prevented proliferation, migration and capillary-like organization of HUVECs in response to human IL-8. In a rat model of liver ischaemia and reperfusion (I/R) injury, DF 2156A decreased PMN and monocyte-macrophage infiltration and associated hepatocellular injury. Conclusion and Implications: DF 2156A is a non-competitive allosteric inhibitor of both IL-8 receptors CXCR1 and CXCR2. It prevented experimental angiogenesis and hepatic I/R injury in vivo and, therefore, has therapeutic potential for acute and chronic inflammatory diseases. Fil: Bertini, R.. Dompé; Italia Fil: Barcelos, L. S.. Universidade Federal de Minas Gerais; Brasil Fil: Beccari, A. R.. Dompé; Italia Fil: Cavalieri, B.. Dompé; Italia. Università di Torino; Italia Fil: Moriconi, A.. Dompé; Italia Fil: Bizzarri, C.. Dompé; Italia Fil: Di Benedetto, P.. Dompé; Italia Fil: Di Giacinto, C.. Dompé; Italia Fil: Gloaguen, I.. Dompé; Italia Fil: Galliera, E.. Università degli Studi di Milano; Italia Fil: Corsi, M. M.. Università degli Studi di Milano; Italia. Policlinico San Donato IRCCS; Italia Fil: Russo, R. C.. Universidade Federal de Minas Gerais; Brasil Fil: Andrade, S. P.. Universidade Federal de Minas Gerais; Brasil Fil: Cesta, M. C.. Dompé; Italia Fil: Nano, G.. Dompé; Italia Fil: Aramini, A.. Dompé; Italia Fil: Cutrin, Juan Carlos. Università di Torino; Italia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Cardiológicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Cardiológicas; Argentina Fil: Locati, M.. Università degli Studi di Milano; Italia Fil: Allegretti, M.. Dompé; Italia Fil: Teixeira, M. M.. Universidade Federal de Minas Gerais; Brasil