1. 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 α 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. 2. 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. 3. 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. 4. 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. 5. 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.
- Adenosine 5′-O-(1-boranotriphosphate) derivatives
- Green fluorescent protein
- Human embryonic kidney (HEK 293) cells
- Purinergic receptor
- Receptor endocytosis