Structural Determinants of A3 Adenosine Receptor Activation:  Nucleoside Ligands at the Agonist/Antagonist Boundary

• Published in: Journal of medicinal chemistry Vol. 45; no. 20; pp. 4471 - 4484
• Main Authors: Gao, Zhan-Guo, Kim, Soo-Kyung, Biadatti, Thibaud, Chen, Wangzhong, Lee, Kyeong, Barak, Dov, Kim, Seong Gon, Johnson, Carl R, Jacobson, Kenneth A
• Format: Journal Article
• Language: English
• Published: Washington, DC American Chemical Society 26.09.2002
• Subjects: Biological and medical sciences; Medical sciences; Neuropharmacology; Neurotransmitters. Neurotransmission. Receptors; Peptidergic system (neuropeptide, opioid peptide, opiates...). Adenosinergic and purinergic systems; Pharmacology. Drug treatments; Agonist; Purine nucleoside; Rat; Lactam; Oxygen heterocycle; Modeling; Structure activity relation; Molecular model; Bicyclic compound; Antagonist; Oxygen nitrogen heterocycle; Adenine derivatives; Human; Molecular rigidity; Adenosine; Rodentia; Tricyclic compound; In vitro; A3 adenosine receptor; Spiran; Vertebrata; Mammalia; Animal; Epoxide; Affinity; Mutation
• ISSN: 0022-2623; 1520-4804
• DOI: 10.1021/jm020211+

Mutagenesis of the human A3 adenosine receptor (AR) suggested that certain amino acid residues contributed differently to ligand binding and activation processes. Here we demonstrated that various adenosine modifications, including adenine substitution and ribose ring constraints, also contributed differentially to these processes. The ligand effects on cyclic AMP production in intact CHO cells expressing the A3AR and in receptor binding were compared. Notably, the simple 2-fluoro group alone or 2-chloro in combination with N 6-substitution dramatically diminished the efficacy of adenosine derivatives, even converting agonist into antagonist. Other affinity-increasing substitutions, including N 6-(3-iodobenzyl) 4 and the (Northern)-methanocarba 15, also reduced efficacy, except in combination with a flexible 5‘-uronamide. 2-Cl-N 6-(3-iodobenzyl) derivatives, both in the (N)-methanocarba (i.e., of the Northern conformation) and riboside series 18 and 5, respectively, were potent antagonists with little residual agonism. Ring-constrained 2‘,3‘-epoxide derivatives in both riboside and (N)-methanocarba series 13 and 21, respectively, and a cyclized (spiral) 4‘,5‘-uronamide derivative 14 were synthesized and found to be human A3AR antagonists. 14 bound potently at both human (26 nM) and rat (49 nM) A3ARs. A rhodopsin-based A3AR model, containing all domains except the C-terminal region, indicated separate structural requirements for receptor binding and activation for these adenosine analogues. Ligand docking, taking into account binding of selected derivatives at mutant A3ARs, featured interactions of TM3 (His95) with the adenine moiety and TMs 6 and 7 with the ribose 5‘-region. The 5‘-OH group of antagonist N 6-(3-iodobenzyl)-2-chloroadenosine 5 formed a H-bond with N274 but not with S271. The 5‘-substituent of nucleoside antagonists moved toward TM7 and away from TM6. The conserved Trp243 (6.48) side chain, involved in recognition of the classical (nonnucleoside) A3AR antagonists but not adenosine-derived ligands, displayed a characteristic movement exclusively upon docking of agonists. Thus, A3AR activation appeared to require flexibility at the 5‘- and 3‘-positions, which was diminished in (N)-methanocarba, spiro, and epoxide analogues, and was characteristic of ribose interactions at TM6 and TM7.