2-Aryl-8-chloro-1,2,4-triazolo[1,5-a]quinoxalin-4-amines as highly potent A1 and A3 adenosine receptor antagonists

Some 2-aryl-8-chloro-1,2,4-triazolo[1,5-a]quinoxaline derivatives 2–18, obtained by introducing different substituents on either the 4-amino moiety (acyl or carbamoyl groups) or the 2-phenyl ring (4-OCH3) of previously reported 8-chloro-2-phenyl-1,2,4- triazolo[1,5-a]quinoxalin-4-amine (1), have been synthesized and tested in radioligand binding assays at bovine A1 and A2A and at cloned human A1 and A3 adenosine receptors. The rationally designed 8-chloro-2-(4-methoxy-phenyl)-1,2,4-triazolo[1,5-a]quinoxalin- 4-acetylamine (14) can be considered one of the most potent and hA3 versus hA1 selective AR antagonists reported till now. The structure–activity relationships of compounds 2–18 are in agreement with those of previously reported 2-aryl-1,2,4-triazolo[4,3- a]quinoxalines (series A) and 2-arylpyrazolo[3,4-c]quinolines (series B), thus suggesting a similar AR binding mode. In fact, the importance for the A3 receptor–ligand interaction of both a strong acidic NH proton donor and a C@O proton acceptor at position- 4, able to engage hydrogen-bonding interactions with specific sites on the A3 AR, has been confirmed. Using our recently published hA3 receptor model, to better elucidate our experimental results, we decided to theoretically depict the putative TM binding motif of the herein reported 1,2,4-triazolo[1,5-a]quinoxaline derivatives on human A3 receptor. Structure–activity relationships have been explained analyzing the three-dimensional structure of the antagonist-receptor models obtained by molecular docking simulation.