Adenosine (Ado) is an endogenous nucleoside ubiquitous in mammals promoting protection and cells repair during metabolic stress conditions. Through interaction with the four Ado receptor subtypes (ARs), AR ligands have shown potential therapeutic interest for many disorders. In this work both new A2AAR agonists and A3AR antagonists were designed, synthesized and tested in vitro. Although Ado 5’-N-ethylcarboxamide derivatives like VT 7 and GCS21680 display a good affinity and selectivity for A2AAR, the development of new agonists for this receptor subtype is still a big challenge in nucleoside chemistry. In this current decade, some papers have reported that a tetrazolyl residue in 4’-position of Ado derivatives led to compounds endowed with good A2AAR affinity. Hence, in this work, compounds bearing the Nethyltetrazoyl moiety in 4’- position of the Ado ribose portion together with different arylalkylthio and arylalkylamino chains in C2-position were designed and synthesized. The new compounds were prepared using a convergent approach. To this purpose, 2,6- dichloropurine was coupled with the suitable modified sugar to afford a nucleoside which was further modified by introducing an amino group at the C6- and the suitable side chain at the C2- position. The modified sugar used in the coupling reaction was synthesized starting from the commercially available D-ribose in seven steps. The binding assay and functional study performed with the new compounds at all AR subtypes transfected on Chinese hamster ovary (CHO) cells revealed that the 2-phenylethylthio derivative was the compound endowed with the better affinity for the A2AAR/A3AR subtypes (17: Ki hA2AR = 5.8 nM; Ki hA3R = 1.2 nM). It is worthwhile to note that the presence of the ethyltetrazolyl substituent in the sugar moiety favors the interaction with the receptor respect to the ethylcarboxamido group. As expected, the new derivatives show a dual behavior at ARs, resulting A2AAR agonists and A3AR antagonists (17: IC50 at hA3AR of 8.4 nM). Furthermore, the wound healing potential of the news nucleosides was evaluated respect to VT 7, CGS21680 and epidermal growth factor (EGF, used as positive control). The compounds, 17, 19, 20, 21 and 22 showed all better wound healing potential respect to VT 7, CGS21680 and EGF. Therefore such compounds are good candidates for further investigation in in vivo model of wound healing. New A3AR antagonists were also prepared based on the observations that, the substitution of the 8-bromine atom of 8-bromo-9-ethyladenine (Ki hA2AAR = 52 nM, Ki hA3AR = 2,800 nM) with phenylacetylene shifts the preference of the resulting compound (Ki hA3AAR = 86 nM, hA2AAR= 600 nM) from A2AAR to A3AR. Hence, from these facts, three series of compounds were prepared. The first one was 8-phenylethynyladenine derivatives substituted at N-9 position with different alkyl/arylalkyl chains. The second one combines substitution on the phenyl ring of 8- phenylethynyladenine with either N-9 cyclopentyl or N9 phenethyl since they resulted being the best N-9 substituents of the first series. The third series combines a fixed para-methoxy- phenylacetylene in C-8 with either N-9 cyclopentyl or N-9 phenethyl, 2-chloro, and with different N6 substituents. The 8-arylethynyladenine derivatives substituted at 9 position with different alkyl/arylalkyl chains, and the corresponding compounds further substituted at the 2 and N6 position, were synthesized starting from commercially available adenine or 2,6- dichlorpurine in three/five steps, respectively. The results of the in vitro test reported that: N-9 cyclopentyl improved affinity while N-9 phenylethyl improved selectivity, the chlorine atom is well tolerated especially when combined with C-8 para-methoxy-phenylacetylene, the N6 substitution gave compounds with maintained selectivity in the same range of the non- substituted derivatives but with a slight decrease of the affinity. Most of the new compounds are endowed with high affinity and different degree of selectivity for the A3AR subtype. In particular, the tetrasubstituted adenine derivative 38 (Ki A3R = 8.4 nM; Ki A1R and Ki A2AR >30,000 nM) resulting the most active and selective ligand and it represents a very good ligand to study the A3AR subtype and its function.
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|Titolo:||Synthesis and Biological Evaluation of new Ligands for the Adenosine Receptors|
|Data di pubblicazione:||22-mar-2019|
|Appare nelle tipologie:||Tesi di dottorato|