Structural determinants of efficacy at A₃ adenosine receptors: Modification of the ribose moiety

We have found previously that structural features of adenosine derivatives, particularly at the N⁶- and 2-positions of adenine, determine the intrinsic efficacy as A₃ adenosine receptor (AR) agonists. Here, we have probed this phenomenon with respect to the ribose moiety using a series of ribose-modified adenosine derivatives, examining binding affinity and activation of the human A₃ AR expressed in CHO cells. Both 2′- and 3′-hydroxyl groups in the ribose moiety contribute to A₃ AR binding and activation, with 2′-OH being more essential. Thus, the 2′-fluoro substitution eliminated both binding and activation, while a 3′-fluoro substitution led to only a partial reduction of potency and efficacy at the A₃ AR. A 5′-uronamide group, known to restore full efficacy in other derivatives, failed to fully overcome the diminished efficacy of 3′-fluoro derivatives. The 4′-thio substitution, which generally enhanced A₃ AR potency and selectivity, resulted in 5′-CH₂OH analogues (10 and 12) which were partial agonists of the A₃ AR. Interestingly, the shifting of the N⁶-(3- iodobenzyl)adenine moiety from the 1′- to 4′-position had a minor influence on A₃ AR selectivity, but transformed 15 into a potent antagonist (16) (K_i=4.3nM). Compound 16 antagonized human A ₃ AR agonist-induced inhibition of cyclic AMP with a K_B value of 3.0nM. A novel apio analogue (20) of neplanocin A, was a full A ₃ AR agonist. The affinities of selected, novel analogues at rat ARs were examined, revealing species differences. In summary, critical structural determinants for human A₃ AR activation have been identified, which should prove useful for further understanding the mechanism of receptor activation and development of more potent and selective full agonists, partial agonists and antagonists for A₃ ARs.