Current Pharmaceutical Design - Volume 8, Issue 26, 2002

The synthesis and the pharmacological activity of alkynyl derivatives of adenosine (Ado) and Nethylcarboxamidoadenosine (NECA), that have been tested on adenosine receptors from different sources, have been reviewed. Most of compounds have been characterized in the last ten years by using radioligand binding assays on rat brain membranes and functional studies on different animal models. More recently, the four human adenosine receptor subtypes have been stably transfected into Chinese hamster ovary (CHO) cells allowing for comparative studies in a similar cellular background, utilizing radioligand binding studies (A1, A2A, A3) or adenylate cyclase activity assays (A2B). From the whole pattern of studies the following structure-activity relationships have been drown:The activities of 2-alkynylAdos resulted slightly higher at A1 and lower at A3 and A2B subtypes than the corresponding NECA derivatives, whereas the affinities at A2A subtype are similar for the two series of nucleosides. -The presence of a methyl group on N6 of the 2-alkynyladenosines, inducing a contemporary increase in affinity at the human A3 receptor and a decrease at the other subtypes, resulted in a relevant increase in A3 selectivity. In particular, 2-phenylethynyl-N6-methylAdo showed an A3 affinity in the low nanomolar range (Ki A3 = 3.4 nM), and about 500 fold A1 / A3 and about 2500 fold A2A / A3 selectivity.The presence of a hydroxyl group in some alkynyl side chains led to potent inhibitors of platelet aggegation induced by ADP. -Introduction of particular substituents, such as the racemic 2-phenylhydroxypropynyl group, both in adenosine and in NECA analogues, led to highly potent, non selective agonists at all the four subtypes.For the potency at A2B receptor it seems to be very important the type of alkynyl chain in 2-position and the presence of the carboxyamido group on the sugar, in fact, the (S)-2-phenylhydroxypropynylNECA [(S)- PHPNECA, EC50 A2B = 220 nM] proved to be one of the most potent A2B agonist reported so far.-The introduction of alkynyl chain in 8-position of adenosine led to very selective ligands for the A3 receptor subtype. These nucleosides behave as adenosine antagonists, since they do not stimulate basal [35S]GTPγS binding, but inhibit NECA-stimulated binding.

Adenosine, a widely distributed modulator, regulates many physiological functions through specific cell membrane G-protein-coupled receptors classified as A1, A2A, A2B and A3. An intense medicinal chemistry effort made over the last 20 years has led to a variety of selective adenosine receptor agonists and antagonists. In particular, the pyrazolo-triazolo-pyrimidine nucleus has been strongly investigated in the last years by our group. All the modifications performed and a tentative of structure-activity-relationship is reported. In fact, the combination of different substitutions at the N7, N8 and N5 positions afford compounds which showed good affinity and selectivity for the different adenosine receptor subtypes. The data herein summarized, permit to speculate on the use of this nucleus as possible template for the adenosine receptor subtypes.

Novel phenomena, such as constitutive activity and inverse agonism, have led to a ligand (re)classification along an agonists-neutral antagonist-inverse agonist continuum. This review focuses on adenosine A1 receptor ligands and their intrinsic activity (α). The intrinsic activity of a ligand depends both on the chemical characteristics of the compound itself and on the experimental conditions in which it is assayed. Consequently, due to this tissue-dependency determination of a ligand's intrinsic activity is not always easily performed. Meanwhile, this feature may also be used in a profitable manner, namely to separate desired and unwanted effects of the drug. Briefly, this review provides possible screening methods to distinguish the different classes of ligands. It also deals with the structural elements and functional groups in the adenosine A1 receptor ligands that determine their intrinsic activity.

In the last decade the field of purinergic pharmacology has continued to grow as the complexity of the receptor families and the various enzymes involved in purine metabolism have been defined in molecular terms. Adenosine receptors (ARs) are currently divided into the four subclasses A1-, A2A-, A2B- and A3AR. The most intensively studied subtypes are the high-affinity A1 and A2A receptors, which are activated by adenosine in nano- to submicromolar concentrations. The clinical importance of the A1 adenosine receptor (A1AR) and the A2A adenosine receptor (A2AAR) makes them attractive targets for radionuclide in vivo imaging. Positron Emission Tomography (PET) is an imaging modality which can determine biochemical and physiological processes in vivo in a quantitative way by using radiopharmaceuticals labeled with positron emitting radionuclides as 11C, 13N, 15O and 18F and by measuring the annihilation radiation using a coincidence technique. This includes also measurement of the pharmacokinetics of labeled drugs and the assessment of the effects of drugs on metabolism. In the present article we review the radioligands which are currently available for visualisation and quantification of ARs using PET with a special focus on the A1AR and A2AAR.

Pyrimidine nucleotides, including UTP, UDP and UDP-glucose, are important signaling molecules which activate G protein-coupled membrane receptors of the P2Y family. Four distinct pyrimidine nucleotide-sensitive P2Y receptor subtypes have been cloned, P2Y2, P2Y4, P2Y6 and P2Y14. Pharmacological experiments indicate that further subtypes may exist. P2Y2 and P2Y4 receptors are activated by UTP (the P2Y2 and the rat, but not the human P2Y4 receptor are also activated by ATP), the P2Y6 receptor is activated by UDP, and the P2Y14 receptor by UDPglucose. Derivatives and analogs of the physiological nucleotides UTP, UDP and ATP have been synthesized and evaluated in order to obtain enzymatically stable, subtype-selective agonists. P2Y2 agonists are currently in clinical development for cystic fibrosis and dry eye syndrome. Selective antagonists for pyrimidinergic P2Y receptors are still lacking.

Extracellular adenine and uracil 5'-nucleotides are important signalling molecules that exert a great variety of effects in numerous tissues and cell types through the activation of P2 receptors. In the past eight years, an extended series of P2 receptors (P2X1-7, ionotropic subunits, P2Y1,2,4,6,11,12, metabotropic receptors) has been cloned from vertebrate tissues. In this rapidly expanding field, one of the main current challenges is to relate the cloned P2 receptor subtypes to the diverse physiological responses mediated by the pharmacological phenotypes of native P2 receptors. Unfortunately, subtype-selective P2 ligands, especially potent and selective antagonists, have been only slowly forthcoming, and this acts as a considerable impediment to progress. However, a number of new P2 receptor antagonists have recently been described which to some degree are more potent and more selective than earlier antagonists like suramin or pyridoxal-5'- phosphate-6-azophenyl-2',4'-disulfonic acid (PPADS). This work moves us closer to the ideal goal of classifying the recombinant and native P2 receptor subtypes on the basis of antagonist profiles. This review begins with a brief account of the current status of P2 receptors and their ligands. It then focuses on structureactivity relationships of PPADS and suramin analogues and will finish with a brief discussion of some related therapeutic possibilities.

Nucleotides are emerging as an ubiquitous family of extracellular signaling molecules. These effects are mediated through a specific class of plasma membrane receptors called P2 receptors that, according to the molecular structure, are further subdivided into two subfamilies: P2Y and P2X. Specifically, P2X-receptors are ligand-gated ion channels, whereas P2Y-receptors belong to the superfamily of G-protein-coupled receptors. In this review, we focus our attention to GPCRs molecular architecture, with the special emphasis on our work on the human P2Y1 receptor. In fact, despite an enormous amount of research on the structure and function of these receptors, fundamental understanding of the molecular details of ligand / GPCR interactions remains very rudimentary. How agonist binding transforms a resting GPCR into its active form and the microscopic basis of binding site blockade by an antagonist are generally still unclear. In the absence of high-resolution structural knowledge of GPCRs, such questions only can be addressed by building models, which are tested through pharmacological and biochemical studies. In this review, we underline how different molecular modeling approaches can help the investigation of both receptor architecture and ligand / receptor molecular recognition.