Enhancement of inosine-mediated A 2A R signaling through positive allosteric modulation

Cell Signal. 2018 Jan;42:227-235. doi: 10.1016/j.cellsig.2017.11.002. Epub 2017 Nov 8.

Abstract

Inosine is an endogenous nucleoside that is produced by metabolic deamination of adenosine. Inosine is metabolically more stable (half-life 15h) than adenosine (half-life <10s). Inosine exerts anti-inflammatory and immunomodulatory effects similar to those observed with adenosine. These effects are mediated in part through the adenosine A2A receptor (A2AR). Relative to adenosine inosine exhibits a lower affinity towards the A2AR. Therefore, it is generally believed that inosine is incapable of activating the A2AR through direct engagement, but indirectly activates the A2AR upon metabolic conversion to higher affinity adenosine. A handful of studies, however, have provided evidence for direct inosine engagement at the A2AR leading to activation of downstream signaling events and inhibition of cytokine production. Here, we demonstrate that under conditions devoid of adenosine, inosine as well as an analog of inosine 6-S-[(4-Nitrophenyl)methyl]-6-thioinosine selectively and dose-dependently activated A2AR-mediated cAMP production and ERK1/2 phosphorylation in CHO cells stably expressing the human A2AR. Inosine also inhibited LPS-stimulated TNF-α, CCL3 and CCL4 production by splenic monocytes in an A2AR-dependent manner. In addition, we demonstrate that a positive allosteric modulator (PAM) of the A2AR enhanced inosine-mediated cAMP production, ERK1/2 phosphorylation and inhibition of pro-inflammatory cytokine and chemokine production. The cumulative effects of allosteric enhancement of adenosine-mediated and inosine-mediated A2AR activation may be the basis for the sustained anti-inflammatory and immunomodulatory effects observed in vivo and thereby provide insights into potential therapeutic interventions for inflammation- and immune-mediated diseases.

Keywords: A(2A)R; Cytokines; ERK1/2; Inosine; PAM; cAMP.

MeSH terms

  • Allosteric Regulation
  • Animals
  • CHO Cells
  • Cell Membrane / drug effects
  • Cell Membrane / metabolism*
  • Chemokine CCL3 / genetics
  • Chemokine CCL3 / immunology
  • Chemokine CCL4 / genetics
  • Chemokine CCL4 / immunology
  • Cricetulus
  • Cyclic AMP / biosynthesis
  • Gene Expression Regulation / immunology*
  • HEK293 Cells
  • Humans
  • Inosine / metabolism*
  • Inosine / pharmacology
  • Lipopolysaccharides / pharmacology
  • Mice
  • Mice, Inbred BALB C
  • Mice, Knockout
  • Mitogen-Activated Protein Kinase 1 / genetics
  • Mitogen-Activated Protein Kinase 1 / immunology
  • Mitogen-Activated Protein Kinase 3 / genetics
  • Mitogen-Activated Protein Kinase 3 / immunology
  • Monocytes / cytology
  • Monocytes / drug effects
  • Monocytes / immunology
  • Phosphorylation
  • Receptor, Adenosine A2A / genetics*
  • Receptor, Adenosine A2A / immunology
  • Signal Transduction*
  • Thioinosine / analogs & derivatives
  • Thioinosine / metabolism
  • Thioinosine / pharmacology
  • Tumor Necrosis Factor-alpha / genetics
  • Tumor Necrosis Factor-alpha / immunology

Substances

  • Chemokine CCL3
  • Chemokine CCL4
  • Lipopolysaccharides
  • Receptor, Adenosine A2A
  • Tumor Necrosis Factor-alpha
  • Thioinosine
  • Inosine
  • Cyclic AMP
  • MAPK1 protein, human
  • Mitogen-Activated Protein Kinase 1
  • Mitogen-Activated Protein Kinase 3