Three new triterpenoid saponins from Albizia julibrissin

Three new triterpenoid saponins, julibrosides A₅–A₇ (1–3), together with five known saponins (4–8), were isolated from the stem bark of Albizia julibrissin. Their structures were elucidated on the basis of extensive spectroscopic data analysis of MS, 1D and 2D NMR, and chemical methods. Compounds 7 and 8 were isolated from the genus Albizia for the first time. The new compounds showed no cytotoxicity and anti-inflammatory activity.     

Quinolinedione nucleus as a novel scaffold for A1 and A2A adenosine receptor antagonists

In the last few years, much effort has been directed towards the synthesis of selective adenosine receptor (AR) antagonists since they are attractive tools for pharmacological intervention in many pathophysiological conditions. During our studies aimed at obtaining new nonclassical adenosine antagonists devoid of phosphodiesterase (PDE) inhibition, a series of 2-pyridones and 2,5-quinolinediones (3a–f, 5a–f, 6a,c–f) has been synthesized as potential AR ligands. Binding affinities of the new compounds were determined for bovine and human adenosine A₁, A_2A, and A₃ receptors. Compound 5f showed good affinity (K _i = 7.8 μM) towards human A₁AR but no selectivity (K _i = 7.0 μM) towards human A_2AAR, whereas compound 6f showed more affinity towards human A_2A (K _i = 16 μM) than A₁ receptor (percentage inhibition at 10 μM concentration = 11). In the 1–100 μM range, the new compounds did not inhibit cardiac PDE3 activity at all. Molecular modeling studies carried out on 5f and 6f support the pharmacological results and suggest 6f as a potential lead compound selective towards A_2AAR.     

Binding of the A1-selective adenosine antagonist 8-cyclopentyl-1,3-dipropylxanthine to rat brain membranes

Summary 8-Cyclopentyl-1,3-dipropylxanthine (PD 116,948) is a very potent, very A₁-selective adenosine antagonist, with a K _i of 0.46 nM in ³H-CHA binding to A₁ receptors in rat whole brain membranes and 340 nM in ³H-NECA binding to A₂ receptors in rat striatal membranes. Its 740-fold A₁-selectivity is the highest reported for an adenosine antagonist. ³H-PD 116,948 (117 Ci/mmol) was prepared by reduction of the diallyl analog. ³H-PD 116,948 bound to a single site in rat whole brain membranes, with a B _max of 46 pmol/g wet weight and K _d of 0.42 nM. Nonspecific binding was extremely low, amounting to about 3% of total binding under standard conditions and less than 1 % when higher tissue concentrations were used. Affinities of compounds for inhibition of ³H-PD 116,948 binding were highly consistent with an A₁ adenosine receptor. Antagonists were equally potent in ³H-PD 116,948 binding and in ³H-CHA binding, while agonists were consistently about 12-fold more potent in ³H-CHA binding. Hill coefficients were 1.0 for antagonists and about 0.65 for agonists. ³H-PD 116,948 should be a useful antagonist ligand for adenosine A₁ receptors. Send offprint request toR. F. Bruns at above address     

Discovery of Pyridone-Substituted Triazolopyrimidine Dual A2A/A1 AR Antagonists for the Treatment of Ischemic Stroke

Ischemic stroke is a complex systemic disease characterized by high morbidity, disability, and mortality. The activation of the presynaptic adenosine A_2A and A₁ receptors modifies a variety of brain insults from excitotoxicity to stroke. Therefore, the discovery of dual A_2A/A₁ adenosine receptor (AR)-targeting therapeutic compounds could be a strategy for the treatment of ischemic stroke. Inspired by two clinical phase III drugs, ASP-5854 (dual A_2A/A₁ AR antagonist) and preladenant (selective A_2A AR antagonist), and using the hybrid medicinal strategy, we characterized novel pyridone-substituted triazolopyrimidine scaffolds as dual A_2A/A₁ AR antagonists. Among them, compound 1a exerted excellent A_2A/A₁ AR binding affinity (K_i = 5.58/24.2 nM), an antagonistic effect (IC₅₀ = 5.72/25.9 nM), and good metabolic stability in human liver microsomes, rat liver microsomes, and dog liver microsomes. Importantly, compound 1a demonstrated a dose–effect relationship in the oxygen-glucose deprivation/reperfusion (OGD/R)-treated HT22 cell model. These findings support the development of dual A_2A/A₁ AR antagonists as a potential treatment for ischemic stroke.     

Involvement of endothelial thromboxane A2 in the vasoconstrictor response induced by 15-E2t-isoprostane in isolated human umbilical vein

The present study was undertaken to evaluate the contractile response of several E- and F-ring isoprostanes (IsoP) in human umbilical vein (HUV) and to investigate the role of the endothelium on the effect of 15-E_2t-IsoP, the most potent vasoconstrictor isoprostane, in human vessels. HUV rings with or without endothelium were suspended in an organ bath for recording the isometric tension in response to different agonists. The inhibitors to be evaluated were applied 30 min before the addition of the agonist. All of the compounds tested produced concentration-dependent contractions when tested on HUV rings with endothelium. Although these compounds were equieffective, significant differences were observed in their potency, with U46619 being the most potent followed by 15-E_2t-IsoP > 15-E_1t-IsoP = 15-F_2t-IsoP > 15-F_1t-IsoP = 9-epi-15-F_2t-IsoP in descending rank order of potency. 15-E_2t-IsoP was the most potent of the isoprostanes evaluated and, therefore, the one employed in the present study. When intact endothelium HUV rings were used, 15-E_2t-IsoP-induced contraction was unaffected by the endothelin-converting enzyme inhibitor, phosphoramidon (10 μM), suggesting that short-term endothelin-1 release is not involved in this response. However, the non-selective cyclooxygenase (COX) inhibitor, indomethacin (10 and 30 μM), and the COX-2 selective inhibitor, NS-398 (3, 10 and 30 μM) produced inhibitory effects on 15-E_2t-IsoP-induced contraction of HUV rings with endothelium. These results indicate that COX-derived contractile prostanoids are involved in this effect. Furthermore, the apparent pK _b values estimated for indomethacin (5.5) and NS-398 (5.4) suggest that the prostanoids involved are derived from the COX-2 isoenzyme pathway. On HUV rings with endothelium, the phospholipase A₂ inhibitor, oleyloxyethyl phosphorylcholine (30 and 100 μM), induced an inhibitory effect on 15-E_2t-IsoP-induced contraction, suggesting that the phospholipase A₂ pathway is also involved in this effect. In addition, the thromboxane A₂ synthase inhibitor furegrelate (10 and 30 μM) also inhibited 15-E_2t-IsoP-induced contraction of HUV rings with endothelium, indicating that thromboxane A₂ is one of the contractile prostanoids involved in this response. Endothelium denudation clearly diminished the vasoconstrictor potency of 15-E_2t-IsoP, demonstrating that the endothelium releases a vasoconstrictor factor in response to 15-E_2t-IsoP. The absence of an inhibitory effect at the highest concentration of furegrelate (30 μM) on 15-E_2t-IsoP-induced contraction of HUV rings without endothelium suggested that endothelium is the source of thromboxane A₂. We conclude that prostanoids derived from the COX-2 isoenzyme pathway participate in 15-E_2t-IsoP-induced vasoconstriction of isolated HUV rings. Our results also indicate that endothelial thromboxane A₂ is one of the prostanoids involved in this effect.     

A new sesquiterpene lactone glucoside with inhibitory effect on K562 cells from Ixeris sonchifolia (Bge) Hance

A new minor sesquiterpene lactone glucoside, ixerin Z_A (1), together with 16 known compounds, were isolated from the whole plants of Ixeris sonchifolia (Bge) Hance. The structure of 1 was elucidated as 1(10),3,11(13)-guaiatriene-12,6-olide-2-one-3-O-[6′-(p-metheoxyphenylacetyl)]-β-glucopyranoside on the basis of spectroscopic and chemical evidence. Compound 1 exhibited an inhibitory effect on K₅₆₂ cells.     

A1 adenosine receptor agonists and their potential therapeutic applications

Background: The challenges in developing any A₁ adenosine receptor (A₁-AdoR) agonist involve having the desired effect on target tissue while avoiding side effects due to activation of A₁-AdoR on other tissues. A₁-AdoR de-sensitization leading to tachyphylaxis is also another challenge. Objectives: The major goal of this review is twofold: to highlight the structure affinity relationships (SAR) of A₁-AdoR agonists, starting with initial lead compounds that were the genesis for second-generation compounds with high selectivity, affinity, and partial agonism; and to give an overview of the A₁-AdoR agonists under development for various indications. Results: Intense efforts by many pharmaceutical companies and academicians in the A₁-AdoR agonist field have led to the discovery of clinical candidates for the following conditions: atrial arrhythmias – Tecadenoson, Selodenoson and PJ-875; type 2 diabetes (T2D) and insulin-sensitizing agents – GR79236, ARA, and CVT-3619; pain management – SDZ WAG 994, GW493838; and angina – BAY-68-4986. For the i.v. antiarrhythmic agents that act as ventricular rate control agents, a selective response can be accomplished by careful dosing paradigms. The treatment of T2D using A₁-AdoR agonists has been met by limited success due to cardiovascular side effects and well-defined desensitization of full agonists in both animal models and human trials (GR79236 and ARA). However, new partial A₁-AdoR agonists are in development, including CVT-3619 (hA₁-AdoR K_i = 55 nm, selectivity A_2A > 200; A_2B > 1000; A₃ > 20, CV Therapeutics), that have the potential to provide enhanced insulin sensitivity without cardiovascular side effects or tachyphylaxis. The A₁-AdoR agonists GW493838 and GR792363 are under evaluation for pain management. The non-nucleosidic A₁-AdoR agonist, BAY-68-4986 (Capadenoson), represents a unique approach to angina wherein both animal studies and early human studies are promising. Conclusion: The challenges associated with developing an A₁-AdoR agonist for therapeutic intervention are now well defined in humans. Significant progress has been made in identifying agents for the treatment of atrial arrhythmias, T2D, and angina.     

Effects of selective A1 and A2 adenosine receptor agonists on cardiovascular tissues

Summary We investigated the negative chronotropic and vasodilating properties of new selective A₁ and A₂ adenosine agonists such as 2-chloro-N⁶-cyclopentyladenosine (CCPA) and 2-hexynyl-5-N-ethyl-carboxamidoadenosine (2-hexynyl-NECA) as compared with reference adenosine analogues. The potency of these compounds on heart rate was assessed in the rat atrial preparation and their activity on the vascular tone was determined in both rat aorta and bovine coronary artery. CCPA was found to be the most potent At agonist of those currently available in producing negative chronotropic effects (EC₅₀ = 8.2 nM). The A1 antagonist 8-cyclopentyl-1,3-dipropyl-xanthine (DPCPX) blocked CCPA activity in a dose-dependent manner. There was also a significant correlation between its biological effect and the affinity for A₁ receptors as measured in the rat brain by [³H]-N⁶-cyclohexyladenosine (³[H]-CHA) binding. The A₂ selective agonist 2-hexynyl-NECA showed vasodilating properties comparable with those observed with the reference compounds, CGS 21680 and NECA. EC₅₀ values were 596 and 569 nM in rat aorta and bovine coronary artery, respectively. Moreover, the rank order of potency was similar in the two vascular districts examined, suggesting that the rat aorta is a useful model for studying the effects of adenosine derivatives on vascular tone. In addition, the potency of the compounds in inducing vasodilation was found to be correlated with their affinity for A₂ receptors as measured in the rat striatum by ³[H]-CGS 21680 binding.These data further support that A₁ receptors are involved in depressing cardiac activity and A₂ receptors in inducing vasorelaxation.Correspondence to A. Conti at the above address     

A detailed behavioral analysis of the acute motor effects of caffeine in the rat: involvement of adenosine A<Subscript>1</Subscript> and A<Subscript>2A</Subscript> receptors

Rationale There is no consensus on the contribution of adenosine A₁ and A_2A receptor blockade to motor-activating effects of caffeine.Objective Our aim was to use a detailed and continuous observational method to compare the motor effects induced by caffeine with those induced by selective A₁ and A_2A receptor antagonists.Methods The behavioral repertoire induced by systemic administration of caffeine (3, 10, and 30 mg/kg), A₁ receptor antagonist 8-cyclopentyl-1,3-dimethylxanthine (CPT; 1.2, 4.8 and 7.2 mg/kg), and A_2A receptor antagonist 3-(3-hydroxypropyl)-8-(m-methoxystyryl)-7-methyl-1-propargylxanthine phosphate disodium salt (MSX-3; 1, 3, and 10 mg/kg) was analyzed. The effects of pretreatment with the selective A₁ receptor agonist N ⁶-cyclopentyladenosine (CPA; 0.1 mg/g) and the selective A_2A receptor agonist 2-p-(2-carboxyethyl)phenethylamino-5′-N-ethylcarboxyamidoadenosine (CGS 21680; 0.2 mg/kg) on the pattern of motor activation induced by caffeine, CPT, or MSX-3 were also examined.Results The pattern of behavioral activation induced by caffeine was better mimicked by CPT than by MSX-3. Coadministration of CPT and MSX-3 gave different results depending on the dose and the type of behavioral response. CPA was more effective at decreasing the activating effects of caffeine and CPT than those of CGS 21680. On the other hand, CGS 21680 was more effective at decreasing the activating effects of MSX-3 than those of caffeine or CPT. Factor analysis revealed a complex three-dimensional behavioral profile for caffeine that was similar to the profile for CPT and was different from the profile for MSX-3.Conclusions The results indicate a predominant role for A₁ receptors in the motor-activating effects of acutely administered caffeine.     

Therapeutic potential of adenosine receptor antagonists and agonists

The adenosine receptors (A₁, A_2A, A_2B and A₃) are important and ubiquitous mediators of cellular signalling, which play vital roles in protecting tissues and organs from damage. Launched drugs include the adenosine receptor antagonists theophylline and doxofylline (both used as bronchodilators in respiratory disorders such as asthma), while several compounds are presently in clinical trials for a range of indications, including heart failure, Parkinson's disease, rheumatoid arthritis, cancer, pain and chronic obstructive pulmonary disease. A host of companies and institutions are addressing the huge potential for the development of selective adenosine receptor agonists and antagonists, so that it appears we are on the verge of a new wave of compounds approaching the market for many unmet medical needs. This review presents an analysis of the patenting activity in the area for 2006 and an interpretation and reflection on the developments that we can expect in the future.     

The quest for the treatment of cognitive impairment: α7 nicotinic and α5 GABAA receptor modulators

Age, age-related pathologies, certain psychiatric disorders, head traumas and other conditions are characterised by an impairment of cognitive functions. Cognition is a complex process involving a large number of neurotransmitters that can modulate, positively or negatively, learning and memory; therefore, their receptors may represent suitable targets to develop cognition-enhancing drugs. Among others, the α₇ nicotinic cholinergic receptor and the α₅ GABA_A receptor are emerging as attractive targets for developing therapeutics in this field. The important role of α₇ nicotinic receptors has been proven thanks to the discovery of α₇-selective agonists, such as GTS-21 and AR-R17779, which has stimulated the synthesis of a large number of new compounds, some of which are in clinical trials. The observation that the classical tranquilliser benzodiazepines (agonists that potentiate GABA_A receptor functions), are amnesic, while inverse agonists (that attenuate the functions of the same receptor) improve cognitive tasks, stimulated the search for modulators mainly directed toward the α₅-containing GABA_A receptor, which seems at present the most important GABA_A receptor subtype involved in cognitive processes. This article reviews the patents on modulators of α₇ nicotinic acetylcholine and GABA_A receptors disclosed during the period 2000 – 2006.     

3-氨基苯甲酰胺增强平阳霉素的抗瘤作用

3 AB是聚(ADP-R)合成酶抑制剂,能在体外和体内协同地增加平阳霉素对S₁₈₀的抑制作用,而本身不具有抗瘤与细胞毒作用。这种加强作用与3 AB的剂量有关,剂量加强因子为4。在体内实验中加用3 AB后,平阳霉素对小鼠S₁₈₀的抑制率由原来的32.3%,41.4%,37.4%和66.0%分别增至60.1%,72.4%,68.3%和77.8%,我们还在体内探讨了量效关系及给药方式的影响。     

Adenosine receptor subtype-selective antagonists in inflammation and hyperalgesia

In this study, we examined the effects of systemic and local administration of the subtype-selective adenosine receptor antagonists PSB-36, PSB-1115, MSX-3, and PSB-10 on inflammation and inflammatory hyperalgesia. Pharmacological blockade of adenosine receptor subtypes after systemic application of antagonists generally led to a decreased edema formation after formalin injection and, with the exception of A₃ receptor antagonism, also after the carrageenan injection. The selective A_2B receptor antagonist PSB-1115 showed a biphasic, dose-dependent effect in the carrageenan test, increasing edema formation at lower doses and reducing it at a high dose. A₁ and A_2B antagonists diminished pain-related behaviors in the first phase of the formalin test, while the second, inflammatory phase was attenuated by A_2B and A₃ antagonists. The A_2B antagonist was particularly potent in reducing inflammatory pain dose-dependently reaching the maximum effect at a low dose of 3 mg/kg. Inflammatory hyperalgesia was totally eliminated by the A_2A antagonist MSX-3 at a dose of 10 mg/kg. In contrast to the A₁ antagonist, the selective antagonists of A_2A, A_2B, and A₃ receptors were also active upon local administration. Our results demonstrate that the blockade of adenosine receptor subtypes can decrease the magnitude of inflammatory responses. Selective A_2A antagonists may be useful for the treatment of inflammatory hyperalgesia, while A_2B antagonists have potential as analgesic drugs for the treatment of inflammatory pain.     

Serotonin receptors represent highly favorable molecular targets for cognitive enhancement in schizophrenia and other disorders

Rationale Current treatments for schizophrenia adequately treat the positive symptoms of schizophrenia but only modestly improve cognitive deficits. This review provides evidence for and against the use of selective 5-HT receptor drugs as cognition enhancing agents for schizophrenia and other disorders.Methods Pre-clinical and clinical literature concerned with the role of the serotonergic system in cognition and memory as it relates to schizophrenia is reviewed. Individual 5-HT receptor subtypes for which selective drugs are available that are likely to improve cognition are reviewed. Recommendations for clinical testing are proposed.Results and conclusions Four 5-HT receptor systems (5-HT_1A, 5-HT_2A, 5-HT₄, 5-HT₆) are highlighted as suitable targets for enhancing cognition and memory. Because many clinically available antipsychotic drugs already target 5-HT_1A, 5-HT_2A and 5-HT₆ receptors, design of clinical trials will need to take into account the serotonergic pharmacology of concurrently administered antipsychotic medications. 5-HT_1A partial agonists and 5-HT_2A antagonists have shown modest effectiveness in improving cognition in schizophrenia. 5-HT₆-selective compounds for cognition enhancement are in late-stage clinical trials, while 5-HT₄ compounds have not yet been tested in humans for cognition enhancement.Recommendations For stand-alone therapy for enhancing cognition, 5-HT_1A partial agonists, 5-HT_2A antagonists, 5-HT₄ partial agonists and 5-HT₆ antagonists are all likely to induce at least modest improvement in cognition in schizophrenia. If add-on therapy is contemplated, antipsychotic drugs with weak affinities for serotonin receptors should be used to avoid confounds. It is likely that serotonergic drugs will soon be available as cognition enhancing medications for disorders other than schizophrenia (e.g. dementia).     

Investigational A3 adenosine receptor targeting agents

Introduction : Adenosine is an endogenous nucleoside that accumulates in the extracellular space in response to metabolic stress and cell damage. Extracellular adenosine is a signaling molecule that signals by activating four GPCRs: the A₁, A_2A, A_2B and A₃ receptors. Since the discovery of A₃ adenosine receptors, accumulating evidence has identified these receptors as potential targets for therapeutic intervention.

Areas covered : A₃ adenosine receptors are expressed on the surface of most immune cell types, including neutrophils, macrophages, dendritic cells, lymphocytes and mast cells. A₃ adenosine receptor activation on immune cells governs a broad array of immune cell functions, which include cytokine production, degranulation, chemotaxis, cytotoxicity, apoptosis and proliferation. In accordance with their multitudinous immunoregulatory actions, targeting A₃ adenosine receptors has been shown to impact the course of a wide spectrum of immune-related diseases, such as asthma, rheumatoid arthritis, cancer, ischemia and inflammatory disorders.

Expert opinion : Given the existence of both preclinical and early clinical data supporting the utility of A₃ adenosine receptor ligands in treating immune-related diseases, further development of A₃ adenosine receptor ligands is anticipated.     

Characterization of a basic phospholipase A2-homologue myotoxin isolated from the venom of the snake Bothrops neuwiedii (yarará chica) from Argentina

A basic protein was isolated by CM-Sephadex C-25 chromatography from the venom of Bothrops neuwiedii from Argentina, and named B. neuwiedii myotoxin I. This protein exerted local myotoxic and edema-forming effects in mice, with potencies comparable to other myotoxins isolated from Bothrops spp. venoms. When injected by i.v. route at doses up to 4.7 mg/kg of body weight, the toxin was not lethal. In vitro, the toxin had no detectable phospholipase A₂ activity on egg yolk phospholipids. B. neuwiedii myotoxin I appeared as a homodimer in sodium dodecylsulphate–polyacrylamide gel electrophoresis, with a subunit molecular weight of 15 kD. Gel immunodiffusion revealed a pattern of partial antigenic identity between the newly isolated myotoxin and myotoxin II from Bothrops asper venom. The sequence of B. neuwiedii myotoxin I was determined for the first 40 amino acid residues, showing high homology to several class II phospholipase A₂ myotoxins of the Lys-49 family from crotalids. Altogether, results suggest that this toxin is a new member of the Lys-49 phospholipase A₂-homologues with myotoxic, cytolytic, and edema-inducing activities.     

Therapeutic potential of A1 adenosine receptor ligands: a survey of recent patent literature

Background: In recent years, a number of companies, universities and other institutions have invested in research on new molecules acting as agonists, antagonists and enhancers on A₁ adenosine receptors (ARs) and in the evaluation of their new therapeutic applications. Objective: To review recent patenting activity on this topic. Methods: The first part of this article describes the compounds patented, subdivided by functional activity, and the second part the most relevant therapeutic applications or new uses for A₁ AR ligands, with a focus on compounds in or soon to be in clinical trials. Conclusion: Although a number of potential therapeutic applications are proposed in the recent literature, at present, in the authors' opinion, very few A₁ ligands are close to coming on the market, probably due to limited selectivity towards the target tissue or organ.     

α-Subunit selective modulators of GABAA receptor function as CNS therapeutics

γ-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter within the CNS. Many drugs, including the benzodiazepines, exert their effects by modulating the GABA_A receptor complex, but side effects are common and reflect, in part, poor subunit selectivity. The subunits are arranged into heteropentamers and this produces a rich diversity of GABA_A receptor subtypes, which are potential targets for treating a variety of CNS disorders including epilepsy, anxiety and insomnia, as well as for ameliorating deficiencies arising from neurodegeneration, such as cognitive impairment. The identification of new ligands with improved subunit selectivity should reduce or abolish some of the side effects observed with current drugs, such as tolerance, dependence and withdrawal. This article focuses on new ligands that are reported to selectively recognise particular α-subunits of GABA_A receptors and may thereby offer improved treatments for CNS disorders. Only patents and literature since 1998 are necessarily included, although some earlier reports and reviews are also cited. Several publications and patent applications since 1998 disclose new compounds that modulate GABA_A receptor function without mentioning whether they have α-subunit selectivity; these compounds, like those known to interact with sites on other GABA_A subunits (particularly β), are not within the scope of this review.     

Binding of [H]MSX-2 (3-(3-hydroxypropyl)-7-methyl-8-(m-methoxystyryl)-1-propargylxanthine) to rat striatal membranes — a new, selective antagonist radioligand for A2A adenosine receptors

The present study describes the preparation and binding properties of a new, potent, and selective A_2A adenosine receptor (AR) antagonist radioligand, [³H]3-(3-hydroxypropyl)-7-methyl-8-(m-methoxystyryl)-1-propargylxanthine ([³H]MSX-2). [³H]MSX-2 binding to rat striatal membranes was saturable and reversible. Saturation experiments showed that [³H]MSX-2 labeled a single class of binding sites with high affinity (K_d=8.0 nM) and limited capacity (B_max=1.16 fmol·mg⁻¹ of protein). The presence of 100 μM GTP, or 10 mM magnesium chloride, respectively, had no effect on [³H]MSX-2 binding. AR agonists competed with the binding of 1 nM [³H]MSX-2 with the following order of potency: 5′-N-ethylcarboxamidoadenosine (NECA)>2-[4-(carboxyethyl)phenylethylamino]-5′-N-ethylcarboxamidoadenosine (CGS-21680)>2-chloroadenosine (2-CADO)>N⁶-cyclopentyladenosine (CPA). AR antagonists showed the following order of potency: 8-(m-bromostyryl)-3,7-dimethyl-1-propargylxanthine (BS-DMPX)>1,3-dipropyl-8-cyclopentylxanthine (DPCPX)>(R)-5,6-dimethyl-7-(1-phenylethyl)-2-(4-pyridyl)-7H-pyrrolo[2,3-d]pyrimidine-4-amine (SH-128)>3,7-dimethyl-1-propargylxanthine (DMPX)>caffeine. The K_i values for antagonists were in accordance with data from binding studies with the agonist radioligand [³H]CGS21680, while agonist affinities were 3–7-fold lower. [³H]MSX-2 is a highly selective A_2A AR antagonist radioligand exhibiting a selectivity of at least two orders of magnitude versus all other AR subtypes. The new radioligand shows high specific radioactivity (85 Ci/mmol, 3150 GBq/mmol) and acceptable nonspecific binding at rat striatal membranes of 20–30%, at 1 nM.