Putative role of the adenosine A3 receptor in the antiproliferative action of N6-(2-isopentenyl)adenosine

We tested a panel of naturally occurring nucleosides for their affinity towards adenosine receptors. Both N6-(2-isopentenyl)adenosine (IPA) and racemic zeatin riboside were shown to be selective human adenosine A3 receptor (hA3R) ligands with affinities in the high nanomolar range (Ki values of 159 and 649 nM, respectively). These values were comparable to the observed Ki value of adenosine on hA3R, which was 847 nM in the same radioligand binding assay. IPA also bound with micromolar affinity to the rat A3R. In a functional assay in Chinese hamster ovary cells transfected with hA3R, IPA and zeatin riboside inhibited forskolin-induced cAMP formation at micromolar potencies. The effect of IPA could be blocked by the A3R antagonist VUF5574. Both IPA and reference A3R agonist 2-chloro-N6-(3-iodobenzyl)adenosine-5′-N-methylcarboxamide (Cl-IB-MECA) have known antitumor effects. We demonstrated strong and highly similar antiproliferative effects of IPA and Cl-IB-MECA on human and rat tumor cell lines LNCaP and N1S1. Importantly, the antiproliferative effect of low concentrations of IPA on LNCaP cells could be fully blocked by the selective A3R antagonist MRS1523. At higher concentrations, IPA appeared to inhibit cell growth by an A3R-independent mechanism, as was previously reported for other A3R agonists. We used HPLC to investigate the presence of endogenous IPA in rat muscle tissue, but we could not detect the compound. In conclusion, the antiproliferative effects of the naturally occurring nucleoside IPA are at least in part mediated by the A3R

Time course of action of three adenosine A1 receptor agonists with differing lipophilicity in rats: comparison of pharmacokinetic, haemodynamic and EEG effects

In this study we investigated the relationship between the pharmacokinetics and the cardiovascular and electroencephalogram (EEG) effects of three adenosine agonists with differing lipophilicity. Conscious normotensive rats received either 600 microg/kg N6-(p-sulphophenyl) adenosine (SPA), 200 microg/kg N6-cyclopentyladenosine (CPA) or 600 microg/kg 1-deaza-2-chloro-N6-cyclopentyladenosine (DCCA) in a 5-min intravenous infusion. Changes in haemodynamics and EEG were monitored in conjunction with arterial blood sampling to determine blood concentrations of the compounds. The three adenosine agonists showed large differences in pharmacokinetic properties, resulting in terminal half-lives of 66 +/- 10, 8.2 +/- 0.4 and 24 +/- 1 min (mean +/- SEM) for SPA, CPA, and DCCA respectively. SPA had a significantly lower blood clearance relative to CPA and DCCA, whereas DCCA had the largest volume of distribution and degree of plasma protein binding. The relationship between concentration and heart rate could be described adequately by the sigmoidal Emax model. For SPA, CPA, and DCCA the EC50 values based on free drug concentrations were 423 +/- 92, 1.8 +/- 0.4 and 9.5 +/- 1.1 nM respectively. These in vivo values correlated closely with the affinity of the compounds for the adenosine A1 receptor as determined in radioligand binding studies, with corresponding Ki values of 1410 +/- 220, 4.7 +/- 0.6 and 102 +/- 74 nM (mean +/- SEM) respectively. In the EEG, only CPA produced a small decrease in the amplitude of beta waves. This study demonstrates that the three adenosine analogues have large differences in pharmacokinetics, which complicates comparison of their cardiovascular and central responses simply on the basis of dose. The application of an integrated PK/PD approach permits estimates of potency and activity which are independent of underlying dose and pharmacokinetic

Ribose-modified adenosine analogues as potential partial agonists for the adenosine receptor

We have adopted a practical three-step route for the synthesis of 2'- and 3'-deoxy analogues of N6-substituted adenosines: protection of the hydroxyl groups, replacement of the N6-amino by a better leaving group, and combined deprotection and N6-amination in the last step. This route was used to synthesize deoxy analogues of CPA, CHA, and R- and S-PIA. The compounds were tested on the adenosine A1 and A2a receptors in our search for partial agonists for these receptors. The GTP shift was used as an in vitro measure for the intrinsic activity of these compounds; the in vivo intrinsic activities of the deoxy analogues of CPA and R-PIA were determined in the rat cardiovascular system. Thus, it was shown that the hydroxyl groups are determinants for the affinity and intrinsic activity of these analogues. Removal of the 2'- and 3'-hydroxyl groups affects affinity and intrinsic activity, whereas removal of the 5'-hydroxyl group decreases only affinit

Partial agonism of theophylline-7-riboside on adenosine receptors

Theophylline-7-riboside was evaluated as a partial agonist for rat adenosine receptors. Radioligand binding experiments were performed on both A1 and A2a adenosine receptors, using several methodologies to discriminate between agonists and antagonists. Mainly from thermodynamic data it was concluded that on A1 receptors theophylline-7-riboside had characteristics intermediate between full agonists, such as N6-cyclopentyladenosine, and full antagonists, such as the xanthines. The partial agonistic behaviour of theophylline-7-riboside was further explored in second messenger studies in intact cells. In FRTL-5 rat thyroid cells theophylline-7-riboside behaved as a partial agonist for A1 receptors, slightly inhibiting forskolin-stimulated cyclic AMP levels. The implications of these biochemical findings were further analysed in in vivo pharmacology. The infusion of theophylline-7-riboside in conscious, normotensive rats led to marked changes in cardiovascular parameters, although less outspoken than observed with full agonists for either A1 or A2a receptors. The concomitant determination of the blood concentrations of theophylline-7-riboside and its metabolite theophylline allowed the estimation of in vivo pharmacokinetic and pharmacodynamic parameters. Thus, the EC50 value of theophylline-7-riboside for lowering the mean arterial pressure was 47 +/- 12 micrograms/ml blood. The short duration of action of theophylline-7-riboside makes it improbable that its metabolite theophylline interferes with its effects. In conclusion, theophylline-7-riboside is one of the first partial agonists for adenosine receptors. It may serve as a tool in further investigations of adenosine receptor partial agonis

8-substituted adenosine and theophylline-7-riboside analogues as potential partial agonists for the adenosine A1 receptor

A series of 8-substituted adenosine and theophylline-7-riboside analogues (28 and 9 compounds, respectively) was tested on adenosine A1 and A2A receptors as an extensive exploration of the adenosine C8-region. Alkylamino substituents at the 8-position cause an affinity decrease for adenosine analogues, but an affinity increase for theophylline-7-riboside derivatives. The affinity decrease is probably due to a direct steric hindrance between the C8-substituent and the binding site as well as to electronic effects, not to a steric influence on the ribose moiety to adopt the anti conformation. The 8-substituents increase the affinity of theophylline-7-riboside analogues probably by binding to a lipophilic binding site. The intrinsic activity was tested in vitro for some 8-substituted adenosine analogues, by determining the GTP shift in receptor binding studies and the inhibition of adenylate cyclase in a culture of rat thyroid FRTL-5 cells, and in vivo in the rat cardiovascular system for 8-butylaminoadenosine. Thus, it was shown that 8-ethyl-, 8-butyl-, and 8-pentylamino substituted analogues of adenosine may be partial agonists in vitro, and that 8-butylaminoadenosine is a partial agonist for the rat cardiovascular A1 receptor in viv

Interleukin-6 upregulates neuronal adenosine A(1) receptors: Implications for neuromodulation and neuroprotection

The immunological response in the brain is crucial to overcome neuropathological events. Some inflammatory mediators, such as the immunoregulatory cytokine interleukin-6 (IL-6) affect neuromodulation and may also play protective roles against various noxious conditions. However, the fundamental mechanisms underlying the long-term effects of IL-6 in the brain remain unclear. We now report that IL-6 increases the expression and function of the neuronal adenosine A(1) receptor, with relevant consequences to synaptic transmission and neuroprotection. IL-6-induced amplification of A(1) receptor function enhances the responses to readily released adenosine during hypoxia, enables neuronal rescue from glutamate-induced death, and protects animals from chemically induced convulsing seizures. Taken together, these results suggest that IL-6 minimizes the consequences of excitotoxic episodes on brain function through the enhancement of endogenous adenosinergic signaling