title: Orthogonal activation of the reengineered A3 adenosine receptor (neoceptor) using tailored nucleoside agonists.
abstract: An alternative approach to overcome the inherent lack of specificity of conventional agonist therapy can be the reengineering of the GPCRs and their agonists. A reengineered receptor (neoceptor) could be selectively activated by a modified agonist, but not by the endogenous agonist. Assisted by rhodopsin-based molecular modeling, we pinpointed mutations of the A(3) adenosine receptor (AR) for selective affinity enhancement following complementary modifications of adenosine. Ribose modifications examined included, at 3': amino, aminomethyl, azido, guanidino, ureido; and at 5': uronamido, azidodeoxy. N(6)-Variations included 3-iodobenzyl, 5-chloro-2-methyloxybenzyl, and methyl. An N(6)-3-iodobenzyl-3'-ureido adenosine derivative 10 activated phospholipase C in COS-7 cells (EC(50) = 0.18 microM) or phospholipase D in chick primary cardiomyocytes, both mediated by a mutant (H272E), but not the wild-type, A(3)AR. The affinity enhancements for 10 and the corresponding 3'-acetamidomethyl analogue 6 were >100-fold and >20-fold, respectively. 10 concentration-dependently protected cardiomyocytes transfected with the neoceptor against hypoxia. Unlike 10, adenosine activated the wild-type A(3)AR (EC(50) of 1.0 microM), but had no effect on the H272E mutant A(3)AR (100 microM). Compound 10 was inactive at human A(1), A(2A), and A(2B)ARs. The orthogonal pair comprising an engineered receptor and a modified agonist should be useful for elucidating signaling pathways and could be therapeutically applied to diseases following organ-targeted delivery of the neoceptor gene.
authors.affiliation: Molecular Recognition Section, Laboratory of Bioorganic Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Zhan-Guo
authors.initials: ZG
authors.lastname: Gao
authors.forename: Heng T
authors.initials: HT
authors.lastname: Duong
authors.forename: Tatiana
authors.initials: T
authors.lastname: Sonina
authors.forename: Soo-Kyung
authors.initials: SK
authors.lastname: Kim
authors.forename: Philippe
authors.initials: P
authors.lastname: Van Rompaey
authors.forename: Serge
authors.initials: S
authors.lastname: Van Calenbergh
authors.forename: Liaman
authors.initials: L
authors.lastname: Mamedova
authors.forename: Hea Ok
authors.initials: HO
authors.lastname: Kim
authors.forename: Myong Jung
authors.initials: MJ
authors.lastname: Kim
authors.forename: Ae Yil
authors.initials: AY
authors.lastname: Kim
authors.forename: Bruce T
authors.initials: BT
authors.lastname: Liang
authors.forename: Lak Shin
authors.initials: LS
authors.lastname: Jeong
authors.forename: Kenneth A
authors.initials: KA
authors.lastname: Jacobson
mesh_terms: D058909:Adenosine A3 Receptor Agonists; D000818:Animals; D001665:Binding Sites; D002478:Cells, Cultured; D002642:Chick Embryo; D002522:Chlorocebus aethiops; D006224:Cricetinae; D005818:Genetic Engineering; D006801:Humans; D008024:Ligands; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D015428:Myocardial Reperfusion Injury / Q000378:metabolism; D009705:Nucleosides / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology*; D017434:Protein Structure, Tertiary; D043684:Receptor, Adenosine A3 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship
chemical_list: D058909:Adenosine A3 Receptor Agonists; D008024:Ligands; D009705:Nucleosides; D043684:Receptor, Adenosine A3