A3 adenosine receptors:: Synthesis and biological evaluation of new potent and selective ligands

New polysubstituted adenosine derivatives were synthesized and tested at the four human adenosine receptor stably transfected into Chinese hamster ovary (CHO) cells, utilizing radioligand binding studies (A(1,), A(2A), A(3)) or adenylyl cyclase activity assay (A(2B)). Binding studies demonstrated that the presence of a phenylethynyl group in the 2 position of adenosine favoured the interaction with A(3) receptors leading to compounds endowed with high affinity and selectivity for the A(3) subtype. Further substitution of the N-6- and 4'-positions enhances the A(3) affinity and selectivity. In particular, the N-6-methoxy-2-phenyl-ethynyl-5'-N-methylcarboxamidoadenosine (5), showing K(i)A(3) = 1.9 nm and a selectivity A(1)/A(3) and A(2A)/A(3) of 4862 and 8670 fold, respectively, results one of the most potent and selective agonist at the human A(3) adenosine receptor subtype reported so far.