Willem Soudijn

5-HT3 receptor antagonists and anxiety; a preclinical and clinical review

Abstract The present paper reviews the evidence for anxiolytic activity of 5-HT3 receptor antagonists in animal models of anxiety and in clinical trials in humans. Compared to the established anxiolytics (benzodiazepine receptor agonists and, to a lesser extent, 5-HT1A receptor agonists) 5-HT3 receptor antagonists display a different anxiolytic profile. They are anxiolytic in a limited number of animal anxiety models. If active, they often are very potent and display bell-shaped dose response curves, whereas the ratio between therapeutic activity and side effects appears remarkably large. 5-HT3 receptor antagonists remain active after chronic dosing and no indications for tolerance, dependence or rebound effects were found, which seems to make these drugs an attractive alternative to the benzodiazepines. However, the large body of animal data indicating a complete lack of psychotropic activity of 5-HT3 receptor antagonists weakens the prediction of anxiolytic activity in these drugs. Human data are also controversial; some investigators have reported positive effects in anxiety disorders (panic disorder, GAD), others did not. It can be concluded that 5-HT3 receptor antagonists do not represent a breakthrough in the treatment of various anxiety disorders, as initially suggested.

Serotonin, dopamine and norepinephrine transporters in the central nervous system and their inhibitors

An overview is presented on progress made in the research on neuronal transporters of serotonin, dopamine and norepinephrine in the central nervous system. Tools developed by molecular biology, such as expression of cloned transporters, their mutants and chimera in non-neuronal cells offered the opportunity to study the putative domains for binding of substrates and uptake inhibitors and discover factors in the regulation of the transporter function. The study of the distribution of monoamine transporters in human brain became possible by the development of selective radiolabelled transport inhibitors. The relationships between the chemical structure of the uptake inhibitors and the affinity for the monoamine transporters is reported, and the (potential) therapeutic applications of the compounds are discussed.

The 5-HT1A receptor and its ligands: structure and function

An overview is presented on progress made in research on 5-HT1A receptors and their ligands since their discovery in 1983. Molecular biology has offered new tools, for example cloned 5-HT1A receptors, their mutants and chimeras to study structure and function. Many compounds, belonging to different chemical classes, display high affinity and selectivity for 5-HT1A receptors. The majority of these compounds are agonists or partial agonists, full antagonists are still scarce. Agonists and partial agonists are active in various animal models of anxiety and depression. Partial receptor agonists have been proven to be effective in general anxiety disorder and depression in man. Potential therapeutic applications for 5-HT1A receptor antagonists are evaluated, for example, in cognition disorders.

Suramin analogs, divalent cations and ATPγS as inhibitors of ecto-ATPase

Ecto-nucleotidases are plasma membranebound enzymes that sequentially dephosphorylate extracellular nucleotides such as ATP. This breakdown of ATP and other nucleotides obscures the characterization and classification of P2 (nucleotide) receptors. We therefore studied suramin and several of its analogs, divalent cations and ATPγS for their ability to inhibit ecto-ATPase in human blood cells. Suramin itself and Ni2+ were the more potent, non-competitive inhibitors with micromolar affinity. ATPγS also displayed micromolar affinity and inhibited ecto-ATPase competitively. The data obtained with the divalent cations demonstrate that coordination of the phosphate chain but not the N7 of the adenine ring is required for the breakdown of ATP by ecto-ATPase. Divalent cations that coordinate both the phosphate chain and N7 inhibit ecto-ATPase in a non-competitive manner.

Interaction of amiloride and its analogues with adenosine A1 receptors in calf brain

Amiloride, a potassium sparing diuretic, is known to interact with a number of ion transport systems, receptors and enzymes. Here, we report on the interaction between this drug and the adenosine A1 receptor as present in calf brain membranes. Adenosine A1 receptors are characterized by a subnanomolar affinity for the antagonists [3H]8-cyclopentyl-1,3-dipropylxanthine ([3H]DPCPX) and the agonist [3H]N6-R-1-phenyl-2-propyladenosine ([3H]PIA). Amiloride displaces both agonist and antagonist binding with a Ki value in the low micromolar range. This inhibition is counteracted by NaCl and protons, in contrast to the binding of [3H]PIA and [3H]DPCPX. The results suggest that amiloride interacts with the adenosine A1 receptor at a site distinct from the ligand binding site. In order to elucidate the role of one of the ion transport systems known to be inhibited by amiloride, eight amiloride analogues with different sensitivities for these systems were tested. The potency and order of potency of these compounds towards adenosine A1 receptors excludes the involvement of the epithelial Na+ channel, Na+/H+ exchanger or Na+/Ca2+ exchanger.

Deoxyribose analogues of N6-cyclopentyladenosine (CPA): partial agonists at the adenosine A1 receptor in vivo

1 The purpose of the present study was to quantify the cardiovascular effects of the 2′‐, 3′‐ and 5′‐deoxyribose analogues of the selective adenosine A1 receptor agonist, N6‐cyclopentyladenosine (CPA) in vivo. The blood concentration‐effect relationships of the compounds were assessed in individual rats and correlated to their receptor binding characteristics. 2 The pharmacokinetics and pharmacodynamics of the compounds were determined after a single intravenous infusion of 0.80 mg kg−1 (2.5 μmol kg−1) of 5′dCPA, 1.2 mg kg−1 (3.8 μmol kg−1) of 3′dCPA or 20 mg kg−1 (63 μmol kg−1) of 2′ dCPA. The heart rate (HR) and mean arterial blood pressure (MAP) were monitored continuously during the experiment and serial arterial blood samples were taken for analysis of drug concentration. 3 The relationship between blood concentrations and the reductions in both heart rate and blood pressure were described according to the sigmoidal Emax model. For the bradycardiac effect, the potencies based on free drug concentrations (EC50, u) of 5′dCPA, 3′dCPA and 2′dCPA in blood were 5.9 ± 1.7, 18 ± 4 and 260 ± 70 ng ml−1 (19 ± 6, 56 ± 11 and 830 ± 210 nM), respectively, and correlated well with the adenosine A1 receptor affinity in vitro. The Emax value of 2′dCPA was significantly less than those of the other compounds, suggesting that this compound may be regarded as a partial agonist when compared to the other analogues. The rank order of the maximal reduction in heart rate of the compounds corresponded well with the order of the GTP‐shifts, as determined in vitro. 4 It is concluded that deoxyribose derivatives of CPA may be partial agonists for the adenosine A1 receptor and may serve as tools for further investigation of adenosine receptor partial agonism in vivo.

Xanthine-7-Ribosides as Adenosine Al Receptor Antagonists: Further Evidence for Adenosine's Anti Mode of Binding

Abstract The synthesis and A1 adenosine receptor affinity of some xanthine-7-ribosides is described. It appears that these compounds are A, receptor antagonists. The orientation of the ribose moiety, as determined by ′H-NMR spectroscopy and theoretical chemical calculations, is compared with the orientation of the ribose in the agonist adenosine. Implications for the syn vs anti modes of binding to the receptor are discussed.

Influence of the molecular structure of N6-(ω-aminoalkyl)adenosines on adenosine receptor affinity and intrinsic activity

The affinities of a series of N6-(omega-aminoalkyl)adenosines as probes for A1 and A2 adenosine receptors were determined in various radioligand binding assays and the intrinsic activities were measured in adenylate cyclase assays. Clear species differences were noticed for A1 receptor affinity of these adenosine receptor agonists, the compounds being more active in calf than in rat brain tissue. The affinity profile within the series was, however, rather similar in both membrane preparations, with N6-9-aminononyladenosine displaying highest affinity. The A2 receptor affinities were comparable to values measured for the A1 receptor in its low affinity state, as assessed with a radiolabelled antagonist in the presence of 1 mM GTP. Calculation of the intrinsic activities of the adenosine analogues from their modulating action on adenylate cyclase showed almost all the compounds to be equally effective to (-)-N6-(R-phenylisopropyl) adenosine, on either A1 or A2 adenosine receptors. N6-3-Aminopropyl- and N6-12-aminododecyladenosine, however, proved to be partial agonists, the first on A1 and the second on A2 adenosine receptors. The data are used as the basis for a discussion of adenosine receptor subtype selectivity and intrinsic activity in general.

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 vivo.

Mapping the N6-region of the adenosine A1 receptor with computer graphics

N6-substituted adenosine derivatives can be very potent and selective agonists at A1 receptors. The N6-region is usually described by a general model introduced by Kusachi et al. (J. Med. Chem. 1985, 28, 1636). Structure-activity relationships for this region have recently been reported from a detailed study by Daly et al. (Biochem. Pharmacol. 1986, 35, 2467). The N6-region of the A1 receptor was now investigated in further detail with the aid of computer graphics. A map of the N6-region was constructed on the basis of a selected set of data from Daly et al. This map allows Kusachis model to be extended with two new subregions: a C subregion, accommodating cycloalkyl substituents and a B subregion, which is filled by bulky substituents, such as a norbornanyl moiety. Furthermore, three regions can be distinguished, where occupation leads to diminished receptor affinity, so-called forbidden areas. The extended model permits the contribution of each subregion to be determined quantitatively, allowing accurate prediction of affinities of compounds that were not used to construct the map.