Subtype specific internalization of P2Y 1 and P2Y 2 receptors induced by novel adenosine 5′‐O‐(1‐boranotriphosphate) derivatives

P2Y‐nucleotide receptors represent important targets for drug development. The lack of stable and receptor specific agonists, however, has prevented successful therapeutic applications. A novel series of P‐boronated ATP derivatives (ATP‐ α ‐B) were synthesized by substitution of a nonbridging O at P with a BH 3 group. This introduces a chiral center, thus resulting in diastereoisomers. In addition, at C2 of the adenine ring a further substitution was made (Cl‐ or methylthio‐). The pairs of diastereoisomers were denoted here as A and B isomers. Here, we tested the receptor subtype specificity of these analogs on HEK 293 cells stably expressing rat P2Y 1 and rat P2Y 2 receptors, respectively, both attached to the fluorescent marker protein GFP (rP2Y 1 ‐GFP, rP2Y 2 ‐GFP). We investigated agonist‐induced receptor endocytosis, [Ca 2+ ] i rise and arachidonic acid (AA) release. Agonist‐induced endocytosis of rP2Y 1 ‐GFP was more pronounced for the A isomers than the corresponding B counterparts for all ATP‐ α ‐B analogs. Both 2‐MeS‐substituted diastereoisomers induced a greater degree of agonist‐induced receptor endocytosis as compared to the 2‐Cl‐substituted derivatives. Endocytosis results are in accordance with the potency to induce Ca 2+ release by these compounds in HEK 293 cells stably transfected with rP2Y 1 . In case of rP2Y 2 ‐GFP, the borano‐nucleotides were very weak agonists in comparison to UTP and ATP in terms of Ca 2+ release, AA release and in inducing receptor endocytosis. The different ATP‐ α ‐B derivatives and also the diastereoisomers were equally ineffective. Thus, the new agonists may be considered as potent and highly specific agonist drug candidates for P2Y 1 receptors. The difference in activity of the ATP analogs at P2Y receptors could be used as a tool to investigate structural differences between P2Y receptor subtypes.British Journal of Pharmacology (2004) 142 , 869–878. doi: 10.1038/sj.bjp.0705859