RESUMEN
Currently, there is an urgent need for biocompatible metal-ion chelators capable of antioxidant activity and disassembly of amyloid beta (Aß)-aggregates as potential therapeutics for Alzheimer's disease (AD). We recently demonstrated the promising antioxidant activity of adenine/guanine 2',3' or 3',5'-bis(thio)phosphate analogues, 2'-dA/G3'5'PO/S and A2'3'PO/S, and their affinity to Zn(ii)-ions. These findings encouraged us to evaluate them as agents for the dissolution of Aß42-Zn(ii)/Cu(ii) aggregates. Specifically, we explored their ability to bind Cu(ii)/Zn(ii)-ions, the geometry and stoichiometry of these complexes, Cu(ii)/Zn(ii)-binding-sites and binding mode, and the ability of these analogues to dissolve Aß42-Zn(ii)/Cu(ii) aggregates, as well as their effect on the secondary structure of those aggregates. Finally, we identified the most promising agents for dissolution of Aß42-Zn(ii)/Cu(ii) aggregates. Specifically, we observed the formation of a 1 : 1 complex between 2'-dG3'5'PO and Cu(ii), involving O4 ligands. Zn(ii) was coordinated by both thiophosphate groups of 2'-dA3'5'PS and A2'3'PS involving O2S2 ligands in a 1 : 1 stoichiometry. A2'3'PS dissolves Aß42-Zn(ii) and Aß42-Cu(ii) aggregates as effectively as, and 2.5-fold more effectively than EDTA, respectively. Furthermore, 2'-dG3'5'PS and A2'3'PS reverted the Aß42-M(ii) structure, back to that of the free Aß42. Finally, cryo-TEM and TEM images confirmed the disassembly of Aß42 and Aß42-M(ii) aggregates by A2'3'PS. Hence, 2'-dG3'5'PS and A2'3'PS may serve as promising scaffolds for new AD therapeutics, acting as both effective antioxidants and agents for solubilization of Aß42-Cu(ii)/Zn(ii) aggregates.
Asunto(s)
Péptidos beta-Amiloides/química , Cobre/química , Nucleósidos/química , Fragmentos de Péptidos/química , Fosfatos/química , Fosfatos/farmacología , Agregado de Proteínas/efectos de los fármacos , Zinc/química , Antioxidantes/química , Antioxidantes/farmacología , Quelantes/química , Quelantes/farmacologíaRESUMEN
We synthesized a series of adenine/guanine 2',3'- or 3',5'-bisphosphate and -bisphosphorothioate analogues, 1-6, as potential Cu(+)/Fe(2+) chelators, with a view to apply them as biocompatible and water-soluble antioxidants. We found that electron paramagnetic resonance (EPR)-monitored inhibition of OH radicals production from H2O2, in an Fe(2+)-H2O2 system, by bisphosphate derivatives 1, 3, and 5 (IC50 = 36, 24, and 40 µM, respectively), was more effective than it was by ethylenediaminetetraacetic acid (EDTA), by a factor of 1.5, 2, and 1.4, respectively. Moreover, 2'-deoxyadenosine-3',5'-bisphosphate, 1, was 1.8- and 4.7-times more potent than adenosine 5'-monophosphate (AMP) and adenosine 5'-diphosphate (ADP), respectively. The bisphosphorothioate derivatives 2, 4, and 6 (IC50 = 92, 50, and 80 µM, respectively), exhibited a dual antioxidant activity, acting as both metal-ion chelators and radical scavengers [2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) assay data indicates IC50 = 50, 70, and 108 µM vs 27 µM for Trolox]. Only 2'-deoxyadenosine-3',5'-bisphosphorothioate, 2, exhibited good inhibition of Cu(+)-induced H2O2 decomposition (IC50 = 78 vs 224 µM for EDTA). Nucleoside-bisphosphorothioate analogues (2, 4, and 6) were weaker inhibitors than the corresponding bisphosphate analogues (1, 3, and 5), due to intramolecular oxidation under Fenton reaction conditions. (1)H- and (31)P NMR monitored Cu(+) titration of 2, showed that Cu(+) was coordinated by both 3',5'-bisphosphorothioate groups, as well as N7-nitrogen atom, while adenosine-2',3'-bisphosphorothioate, 6, coordinated Cu(+) only by 2',3'-bisphosphorothioate groups. In conclusion, an additional terminal phosphate group on AMP/guanosine 5'-monophosphate (GMP) resulted in Fe(2+)-selective chelators highly potent as Fenton reaction inhibitors.