Synthesis and pharmacological characterization of [125I]MRS5127, a high affinity, selective agonist radioligand for the A3 adenosine receptor

A recently reported selective agonist of the human A(3) adenosine receptor (hA(3)AR), MRS5127 (1'R,2'R,3'S,4'R,5'S)-4'-[2-chloro-6-(3-iodobenzylamino)-purine]-2',3'-O-dihydroxy-bicyclo-[3.1.0]hexane, was radioiodinated and characterized pharmacologically. It contains a rigid bicyclic ring system in place of a 5'-truncated ribose moiety, and was selected for radiolabeling due to its nanomolar binding affinity at both human and rat A(3)ARs. The radioiodination of the N-6-3-iodobenzyl substituent by iododestannylation of a 3-(trimethylstannyl)benzyl precursor was achieved in 73% yield, measured after purification by HPLC. [I-125]MRS5127 bound to the human A(3)AR expressed in membranes of stably transfected HEK 293 cells. Specific binding was saturable, competitive, and followed a one-site binding model, with a K-d value of 5.74 +/- 0.97 nM. At a concentration equivalent to its K-d, non-specific binding comprised 27 +/- 2% of total binding. In kinetic studies, [I-125]MRS5127 rapidly associated with the hA(3)AR (t(1/2) =0.514 +/- 0.014 min), and the affinity calculated from association and dissociation rate constants was 3.50 +/- 1.46 nM. The pharmacological profile of ligands in competition experiments with [I-125]MRS5127 was consistent with the known structure-activity-relationship profile of the hA(3)AR. [I-125]MRS5127 bound with similar high affinity (K-d, nM) to recombinant A(3)ARs from mouse (4.90 +/- 0.77), rabbit (2.53 +/- 0.11), and dog (3.35 +/- 0.54). For all of the species tested, MRS5127 exhibited A(3)AR agonist activity based on negative coupling to cAMP production. Thus, [I-125]MRS5127 represents a new species-independent agonist radioligand for the A3AR. The major advantage of [I-125]MRS5127 compared with previously used A(3)AR radioligands is its high affinity, low degree of non-specific binding, and improved A(3)AR selectivity. (C) 2009 Elsevier Inc. All rights reserved.