Characterization of Dual-Acting A 3 Adenosine Receptor Positive Allosteric Modulators That Preferentially Enhance Adenosine-Induced Gα i3 and Gα oA Isoprotein Activation.

The A 3 adenosine receptor (A 3 AR) is a promising therapeutic target for inflammatory diseases, cancer, and chronic neuropathic pain, with agonists already in advanced clinical trials. Here we report an in-depth comparison of the pharmacological properties and structure-activity relationships of existing and expanded compound libraries of 2-substituted 1 H -imidazo[4,5- c ]quinolin-4-amine and 4-amino-substituted quinoline derivatives that function as A 3 AR positive allosteric modulators (PAMs). We also show that our lead compound from each series enhances adenosine-induced A 3 AR signaling preferentially toward activation of Gα i3 and Gα oA isoproteins, which are coexpressed with the A 3 AR in immune cells and spinal cord neurons. Finally, utilizing an extracellular/intracellular chimeric A 3 AR approach composed of sequences from a responding (human) and a nonresponding (mouse) species, we provide evidence in support of the idea that the imidazoquinolin-4-amine class of PAMs variably interacts dually with the orthosteric ligand binding site as well as with a separate allosteric site located within the inner/intracellular regions of the receptor. This study has advanced both structural and pharmacological understanding of these two classes of A 3 AR PAMs, which includes leads for future pharmaceutical development.