Synthesis, Biological Evaluation, and Molecular Modeling of Ribose-Modified Adenosine Analogues as Adenosine Receptor Agonists

A number of 3‘-C-methyl analogues of selective adenosine receptor agonists such as CPA, CHA, CCPA, 2‘-Me-CCPA, NECA, and IB-MECA was synthesized to further investigate the subdomain of the receptor that binds the ribose moiety of the ligands. Affinity data at A1, A2A, and A3 receptors in bovine brain membranes showed that the 3‘-C-modification in adenosine resulted in a decrease of the affinity at all three receptor subtypes. When this modification was combined with N 6-substitution with groups that induce high potency and selectivity at A1 receptor, the affinity and selectivity were increased. However, all 3‘-C-methyl derivatives proved to be very less active than the corresponding 2‘-C-methyl analogues. The most active compound was found to be 3‘-Me-CPA which displayed a K i value of 0.35 μM at A1 receptor and a selectivity for A1 vs A2A and A3 receptors higher than 28-fold. 2‘-Me-CCPA was confirmed to be the most selective, high affinity agonist so far known also at human A1 receptor with a K i value of 3.3 nM and 2903- and 341-fold selective vs human A2A and A3 receptors, respectively. In functional assay, 3‘-Me-CPA, 3‘-Me-CCPA, and 2-Cl-3‘-Me-IB-MECA inhibited forskolin-stimulated adenylyl cyclase activity with IC50 values ranging from 0.3 to 4.9 μM, acting as full agonists. A rhodopsin-based model of the bovine A1AR was built to rationalize the higher affinity and selectivity of 2‘-C-methyl derivatives of N 6-substituted-adenosine compared to that of 3‘-C-methyl analogues. In the docking exploration, it was found that 2‘-Me-CCPA was able to form a number of interactions with several polar residues in the transmembrane helices TM-3, TM-6, and TM-7 of bA1AR which were not preserved in the molecular dynamics simulation of 3‘-Me-CCPA/bA1AR complex.