Exploring Trisubstituted adenine derivatives as adenosine A1 receptor ligands with antagonist activity: Synthesis, biological evaluation and molecular modelling

The therapeutic potential of adenosine receptor (AR) ligands is becoming increasingly important as our understanding of the physio-pathological functions of ARs advances. This study presents the synthesis and biological screening of six novel trisubstituted adenine analogues, expanding a previously reported series. The AR binding affinity and antiproliferative activity were evaluated for both the new and previously reported compounds, leading to the discovery of derivatives that displaying selective binding affinity towards hA₁AR. Compounds were synthesized using a cyclization approach by combining 4,6-bisalkylamino-5-aminopyrimidines with three different trialkyl/arylorthoesters, thereby generating adenines featuring three different substituents at the 8 position: H, methyl or phenyl. Most promising derivatives presented a phenyl ring at such position and displayed selective antagonistic activity against hA₁AR. N⁶,9-diisopropyl-8-phenyl-9H-purin-6-amine (14c) was identified as the most potent compound with a K_i of 2 nM, motivating the synthesis of new derivatives including N⁶,9-dicyclopentyl-8-phenyl-9H-purin-6-amine (19c). Docking modelling predicted key interactions between the lead compounds and hA₁AR. Determination of their anti-proliferative activity on six cancer cell lines found 19c to be the most potent derivative with low micromolar EC₅₀ values. Our findings support further exploration around the adenine scaffold for cancer research and AR drug development.