A side-by-side pharmacological comparison of ribose and (N)-methanocarba (bicyclo[3.1.0]hexane) nucleosides as A 3 AR agonists indicated that the bicyclic pseudoribose ring constraint provided higher affinity/selectivity at human and mouse A 3 AR. The mean affinity enhancement for 5 pairs of 5′methylamides was 11-fold at hA 3 AR and 42-fold at mA 3 AR. Novel C2-(5-fluorothien-2-ylethynyl) substitution enhanced affinity in the methanocarba but not ribose series, with highly hA 3 ARselective 16 (MRS7334) displaying K i 280 pM and favorable pharmacokinetics and off-target activity profile. Molecular dynamics comparison of 16 and its corresponding riboside 8 suggested a qualitative entropic advantage of 16 in hA 3 AR binding. The 5-F substitution tended to increase hA 3 AR affinity (cf. 5-Cl) for methanocarba but not ribose derivatives. A representative methanocarba agonist 4 was shown to interact potently exclusively with A 3 AR, among 240 GPCRs and 466 kinases. Thus, despite added synthetic difficulty, the (N)-methanocarba modification has distinct advantages for A 3 AR agonists, which have translational potential for chronic disease treatment.

Dopamine-derived N 6 -substituents, compared to N 6 -(2-phenylethyl), in truncated (N)-methanocarba (bicyclo[3.1.0]hexyl) adenosines favored high A 3 adenosine receptor (AR) affinity/selectivity, e.g., C2-phenylethynyl analogue 15 (MRS7591, K i = 10.9/17.8 nM, at human/mouse A 3 AR). 15 was a partial agonist in vitro (hA 3 AR, cAMP inhibition, 31% E max ; mA 3 AR, [ 35 S]GTP-γ-S binding, 16% E max ) and in vivo and also antagonized hA 3 AR in vitro. Distal H-bonding substitutions of the N 6 -(2-phenylethyl) moiety particularly enhanced mA 3 AR affinity by polar interactions with the extracellular loops, predicted using docking and molecular dynamics simulation with newly constructed mA 3 AR and hA 3 AR homology models. These hybrid models were based on an inactive antagonistbound hA 1 AR structure for the upper part of TM2 and an agonistbound hA 2A AR structure for the remaining TM portions. These species-independent A 3 AR-selective nucleosides are low efficacy partial agonists and novel, nuanced modulators of the A 3 AR, a drug target of growing interest.

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N)-Methanocarba ([3.1.0]bicyclohexyl) adenosines and corresponding ribosides were synthesized to identify novel A 1 adenosine receptor (A 1 AR) agonists for CNS or peripheral applications. Human and mouse AR binding was determined to assess the constrained ring system's A 1 AR compatibility. N 6 -Dicyclobutylmethyl ribose agonist (9, MRS7469, >2000-fold selective for A 1 AR) and known truncated N 6 -dicyclopropylmethyl methanocarba 7 (MRS5474) were drug-like.The pure diastereoisomer of known riboside 4 displayed high hA 1 AR selectivity. Methanocarba modification reduced A 1 AR selectivity of N 6 -dicyclopropylmethyl and endo-norbornyladenosines, but increased ribavirin selectivity. Most analogues tested (ip.) were inactive or weak in inducing mouse hypothermia, despite mA 1 AR full agonism and variable mA 3 AR efficacy, but

Adenines that incorporate known agonist affinity-enhancing substituents are A3AR-selective antagonists.