Steric influence of 4′-position substituents and C2-Hexynyl group on A₃AR antagonism in truncated 4′-Thioadenosine derivatives

A₃ adenosine receptor (A₃AR) has attracted significant interest due to its therapeutic potential in inflammation, neurodegenerative disorders, and cancer. This study investigates the structure-activity relationships (SAR) of truncated 4′-thioadenosine derivatives with modifications at the 4′- and C2-positions. Alkyl substituents of varying sizes (methyl, ethyl, isopropyl) were introduced at the 4′-position to evaluate steric effects, while C2-propynyl and hexynyl groups were incorporated to explore binding enhancement. The 4′-ethyl derivatives exhibit potent A₃AR antagonistic effects, compared to the 4′-methyl series which exhibits partial agonist activity. C2-hexynyl substitution significantly enhanced binding affinity and antagonistic properties. In contrast, 4′-isopropyl derivatives lacked measurable binding, highlighting steric constraints at 4′-position. Docking studies revealed that none of the compounds formed interactions with Thr94, a residue critical for agonistic activity. This lack of interaction likely explains their observed antagonistic behavior, with the 4′-alkyl and C2-hexynyl substitutions appearing to play a significant role in this effect. This work underscores the role of steric bulk at the 4′-position and hydrophobic modifications at the C2-position in modulating A₃AR activity, providing valuable insights for designing selective A₃AR antagonists.