NF546 [4,4′-(Carbonylbis(imino-3,1-phenylene-carbonylimino-3,1-(4-methyl-phenylene)-carbonylimino))-bis(1,3-xylene-α,α′-diphosphonic Acid) Tetrasodium Salt] Is a Non-Nucleotide P2Y11 Agonist and Stimulates Release of Interleukin-8 from Human Monocyte-Derived Dendritic Cells

• Published in: The Journal of pharmacology and experimental therapeutics 2010-01, Vol.332 (1), p.238-247
• Main Authors: Meis, Sabine, Hamacher, Alexandra, Hongwiset, Darunee, Marzian, Claudia, Wiese, Michael, Eckstein, Niels, Royer, Hans-Dieter, Communi, Didier, Boeynaems, Jean-Marie, Hausmann, Ralf, Schmalzing, Günther, Kassack, Matthias U.
• Format: Article
• Language: English
• Subjects: Calcium - metabolism; Cell Culture Techniques; Cell Line, Tumor; Cloning, Molecular; Cyclic AMP - metabolism; Dendritic Cells - drug effects; Dendritic Cells - immunology; Dendritic Cells - metabolism; Diphosphonates - pharmacology; Dose-Response Relationship, Drug; Humans; Interleukin-8 - secretion; Ligands; Naphthalenesulfonates - pharmacology; Protein Binding; Purinergic P2 Receptor Agonists; Receptors, Purinergic P2 - metabolism; Receptors, Purinergic P2X; Recombinant Proteins; Transfection
• ISSN: 0022-3565; 1521-0103
• DOI: 10.1124/jpet.109.157750

The G protein-coupled P2Y11 receptor is involved in immune system modulation. In-depth physiological evaluation is hampered, however, by a lack of selective and potent ligands. By screening a library of sulfonic and phosphonic acid derivatives at P2Y11 receptors recombinantly expressed in human 1321N1 astrocytoma cells (calcium and cAMP assays), the selective non-nucleotide P2Y11 agonist NF546 [4,4′-(carbonylbis(imino-3,1-phenylene-carbonylimino-3,1-(4-methyl-phenylene)carbonylimino))-bis(1,3-xylene-α,α′-diphosphonic acid) tetrasodium salt] was identified. NF546 had a pEC50 of 6.27 and is relatively selective for P2Y11 over P2Y1, P2Y2, P2Y4, P2Y6, P2Y12, P2X1, P2X2, and P2X2-X3. Adenosine-5′-O-(3-thio)triphosphate (ATPγS), a nonhydrolyzable analog of the physiological P2Y11 agonist ATP, and NF546 use a common binding site as suggested by molecular modeling studies and their competitive behavior toward the nanomolar potency antagonist NF340 [4,4′-(carbonylbis(imino-3,1-(4-methyl-phenylene)carbonylimino))bis(naphthalene-2,6-disulfonic acid) tetrasodium salt] in Schild analysis. The pA2 of NF340 was 8.02 against ATPγS and 8.04 against NF546 (calcium assays). NF546 was further tested for P2Y11-mediated effects in monocyte-derived dendritic cells. Similarly to ATPγS, NF546 led to thrombospondin-1 secretion and inhibition of lipopolysaccharide-stimulated interleukin-12 release, whereas NF340 inhibited these effects. Further, for the first time, it was shown that ATPγS or NF546 stimulation promotes interleukin 8 (IL-8) release from dendritic cells, which could be inhibited by NF340. In conclusion, we have described the first selective, non-nucleotide agonist NF546 for P2Y11 receptors in both recombinant and physiological expression systems and could show a P2Y11-stimulated IL-8 release, further supporting the immunomodulatory role of P2Y11 receptors.