Identification of Highly Promising Antioxidants/Neuroprotectants Based on Nucleoside 5′-Phosphorothioate Scaffold. Synthesis, Activity, and Mechanisms of Action

With a view to identify novel and biocompatible neuroprotectants, we designed nucleoside 5′-thiophosphate analogues, 6-11. We identified 2-SMe-ADP(α-S), 7A, as a most promising neuroprotectant. 7A reduced ROS production in PC12 cells under oxidizing conditions, IC₅₀ of 0.08 vs 21 μM for ADP. Furthermore, 7A rescued primary neurons subjected to oxidation, EC₅₀ of 0.04 vs 19 μM for ADP. 7A is a most potent P2Y₁-R agonist, EC₅₀ of 0.0026 μM. Activity of 7A in cells involved P2Y_1/12-R as indicated by blocking P2Y₁₂-R or P2Y₁-R. Compound 7A inhibited Fenton reaction better than EDTA, IC₅₀ of 37 vs 54 μM, due to radical scavenging, IC₅₀ of 12.5 vs 30 μM for ADP, and Fe(II)-chelation, IC₅₀ of 80 vs >200 μM for ADP (ferrozine assay). In addition, 7A was stable in human blood serum, t_1/2 of 15 vs 1.5 h for ADP, and resisted hydrolysis by NPP1/3, 2-fold vs ADP. Hence, we propose 7A as a highly promising neuroprotectant.