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  • Further modifications on the position

    2019-11-02

    Further modifications on the 8-position of compound led to compound 4 (8-pCPT-2′--Me-cAMP, a.k.a. 007, ), which is a potent (half-maximal activation of EPAC1 at 2.2µM) and selective (about 100-fold EPAC/PKA selectivity) EPAC agonist. Since the discovery of compound , it has been widely used as a powerful tool for elucidating the EPAC functions., , , In rat models, injection of compound can provoke mechanical hyperalgesia through the mechanism of activating the ε isoform of PKC. When 5-HT at the threshold and subthreshold concentrations, compound remarkably increased the frequency of Ca oscillations. Human peripheral blood lymphocytes (PBL) cultures, treated with compound , at a 100µM level, decreased approximately 40% HIV-1 viral replication. According to the most recent studies, compound recapitulated the cocaine-induced increase in GluA2-lacking α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic Parathyroid Hormone (1-34), bovine receptors (AMPARs) in dopamine neurons of the ventral tegmental area (VTA), indicating its therapeutic potential for drug additions.Based on its aforementioned advantages, compound 4 has been widely employed as a useful chemical probe for EPAC function studies, but its biological application is limited due to its poor plasma membrane penetrating capability. Esterification of the negatively charged singly bonded oxygen of the compound 4 phosphate group led to compound 5 (8-pCPT-2′--Me-cAMP-AM, a.k.a. 007-AM, ), which contains a mixture of the equatorial and the axial isomers. Compound works as a prodrug of , and can be hydrolyzed to release quickly after crossing the plasma membrane. Compound displays an effect of over 100-fold more potent than compound without affecting its EPAC/PKA selectivity, and it has greatly facilitated the studies of EPAC signaling pathways., However, it is a good substrate for phosphodiesterases (PDE) and also acts as a PDE inhibitor. On the other hand, introducing a phenyl group to compound at the -6 position led to compound (-phenyl-cAMP, ). Biological studies suggest that compound acts as a full PKA agonist instead of EPAC agonist. This finding indicates that modification on the -6 position of compound 1 may lead to PKA/EPAC selective agonists. Recently, several other analogues of compound as EPAC modulators were reported. For example, compound (), a highly specific EPAC activator, has better lipophilicity and membrane permeability than compound . Nevertheless, it still suffers from PDE inhibition side effects. Rehmann and co-workers developed a series of compound analogues as EPAC2-selective agonists through a structure-guided approach. The represented compound exhibits over 100-fold EPAC2 active potency toward EPAC1. Although cAMP analogues (e.g. compounds and ) have been widely used as the powerful chemical probes for the study of EPAC functions, more attention should be paid to their selectivity against other targets beyond PKA and EPAC. According to recent studies, most cAMP analogues (e.g. compounds and ) have multiple cellular targets, thereby leading to cross-target activities and off-target effects., , , , A series of non-cyclic nucleotide EPAC selective inhibitors were reported by Cheng and colleagues., They established a sensitive and robust fluorescence-based high throughput (HTS) assay for screening EPAC specific inhibitors that compete with compound (). Compound is suitable for screening EPAC specific inhibitors because it can induce more than 100-fold fluorescent change when binding to purified full-length EPAC2, and can be reversed by addition of excessive cAMP. Three compounds from the NCI diversity set library were identified with IC values range from 1.7 to 7.9µM in competing with compound 9 in binding EPAC2. Furthermore, they expanded their screening to the Maybridge Hitfinder compound library with about 14,400 compounds. Seven compounds (compounds to , ) were discovered to completely inhibit EPAC2 GEF activities at 25µM in the presence of a same concentration of cAMP.