Delineating the mode of action of adenosine A1 receptor allosteric modulators.
Despite the identification of 2-amino-3-benzoylthiophenes (2A3BTs) as the first example of small-molecule allosteric potentiators of agonist function at a G protein-coupled receptor (GPCR)-the adenosine A(1) receptor-their mechanism of action is still not fully understood. We now report the mechanistic basis for the complex behaviors noted for 2A3BTs at A(1) receptors. Using a combination of membrane-based and intact-cell radioligand binding, multiple signaling assays, and a native tissue bioassay, we found that the allosteric interaction between 2A3BTs and the agonists 2-chloro-N(6)-[(3)H]cyclopentyladenosine or (-)-N(6)-(2-phenylisopropyl)adenosine (R-PIA) or the antagonist [(3)H]8-cyclopentyl-1,3-dipropylxanthine is consistent with a ternary complex model involving recognition of a single extracellular allosteric site. However, when allowed access to the intracellular milieu, 2A3BTs have a secondary action as direct G protein inhibitors; this latter property is receptor-independent as it is observed in nontransfected cells and also after stimulation of another GPCR. In addition, we found that 2A3BTs can signal as allosteric agonists in their own right but show bias toward certain pathways relative to the orthosteric agonist, R-PIA. These results indicate that 2A3BTs have a dual mode of action when interacting with the A(1) receptor and that they can engender novel functional selectivity in A(1) signaling. These mechanisms need to be factored into allosteric ligand structure-activity studies.
Valant
Celine
C
Drug Discovery Biology & Department of Pharmacology, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, Victoria, Australia.
Aurelio
Luigi
L
Urmaliya
Vijay B
VB
White
Paul
P
Scammells
Peter J
PJ
Sexton
Patrick M
PM
Christopoulos
Arthur
A
D000241:Adenosine / Q000031:analogs & derivatives / Q000494:pharmacology; D000495:Allosteric Site; D008024:Ligands; D043682:Receptor, Adenosine A1 / Q000187:drug effects / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D014970:Xanthines
D008024:Ligands; D043682:Receptor, Adenosine A1; D043562:Receptors, G-Protein-Coupled; D014970:Xanthines; C047235:N-(1-methyl-2-phenylethyl)adenosine; C051360:1,3-dipropyl-8-cyclopentylxanthine; D000241:Adenosine; C046489:1,3-dipropylxanthine