Evidence that cyclic guanosine monophosphate (cGMP) mediates endothelium-dependent relaxation

The mechanism of action of endothelium-derived relaxant factor (EDRF) was studied using aortic strip preparations of the rabbit and a bioassay system of a rabbit coronary artery perfused in series with an intact aorta. Methylene blue (an inhibitor of guanylate cyclase) inhibited, and 2-O-propoxyphenyl-8-azapurine-6-one (MB22948, an inhibitor of cGMP phosphodiesterase) potentiated the vascular effects of EDRF whether these were due to its basal or to stimulated release. Infusion of these agents at different sites in the bioassay indicated that they act pharmacologically at the smooth muscle level and not on release of EDRF or by chemical interaction with EDRF. The data are consistent with the hypothesis that EDRF-induced relaxation is mediated by elevation of smooth muscle cGMP levels.     

Nipecotic acid directly activates GABA(A)-like ion channels.

The GABA-related compound nipecotic acid is commonly used to inhibit GABA uptake. This report shows that nipecotic acid can also directly activate GABA(A)-like chloride channels. When applied to outside-out patches of paraventricular neurones, nipecotic acid (1 mM) activated inward unitary currents (approximately 3 pA at a holding potential of -60 mV, E(Cl)+44 mV). The EC(50) for ion channel activation was approximately 300 microM, 3 fold greater than that found for GABA itself in this preparation. The nipecotic acid activated channels had similar conductance and kinetic properties to those of GABA activated channels in the same patches, reversed near E(Cl) and were inhibited by bicuculline (3 microM). This study indicates that for experiments in which relatively high concentrations of nipecotic acid are used, possible direct GABA(A) receptor agonist properties should be considered.     

Study of A(2A) adenosine receptor gene deficient mice reveals that adenosine analogue CGS 21680 possesses no A(2A) receptor-unrelated lymphotoxicity

Cell surface A(2A) adenosine receptor (A(2A)R) mediated signalling affects a variety of important processes and adenosine analogues possess promising pharmacological properties. Demonstrating the receptor specificity of potentially lymphotoxic adenosine-based drugs facilitates their development for clinical applications. To distinguish between the receptor-dependent and -independent lymphotoxicity and apoptotic activity of adenosine and its analogues we used lymphocytes from A(2A)R-deficient mice. Comparison of A(2A)R-expressing (+/+) and A(2A)R-deficient (-/-) cells in cyclic AMP accumulation assays confirmed that the A(2A)R agonist CGS 21680 is indeed selective for A(2A) receptors in T-lymphocytes. Incubation of A(2A)R-expressing thymocytes with extracellular adenosine or CGS 21680 in vitro results in the death of about 7-15% of thymocytes. In contrast, no death was induced in parallel assays in cells from A(2A)R-deficient mice, providing genetic evidence that CGS 21680 does not display adenosine receptor-independent intracellular cytotoxicity. The A(2A) receptor-specific lymphotoxicity of CGS 21680 is also demonstrated in a long-term (6-day) in vitro model of thymocyte positive selection where addition of A(2A)R antagonist ZM 241,385 did block the effects of CGS 21680, allowing the survival of T cells. The use of cells from adenosine receptor-deficient animals is proposed as a part of the screening process for potential adenosine-based drugs for their receptor-independent cytotoxicity and lymphotoxicity.     

2-Fluoro-beta-alanine, a previously unrecognized substrate for bile acid coenzyme A:amino acid:N-acyltransferase from human liver

Our laboratory has demonstrated recently that conjugates of 2-fluoro-beta-alanine (FBAL) and bile acids are the major biliary metabolites of 5-fluorouracil (FUra) in cancer patients. Bile acids are normally conjugated with glycine or taurine, and therefore the identification of the FBAL-bile acid conjugates suggested that FBAL may also be a substrate for the bile acid conjugating enzyme, bile acid CoA:amino acid:N-acyltransferase. Enzyme activity detected using glycine and taurine as substrates was purified 8-fold from human liver cytosol using a DEAE-cellulose column. This preparation when tested for its activity towards beta-alanine and FBAL using cholyl CoA as the bile acid substrate only catalyzed the formation of FBAL-cholate. beta-Alanine was not a substrate. Confirmation of FBAL-cholate as the enzymatic product was demonstrated by (1) coelution of the product of this reaction on HPLC with authentic FBAL-cholate, (2) specific hydrolysis of this product by cholylglycine hydrolase, and (3) molecular weight of the product (497) being identical to that of the authentic FBAL-cholate. Kinetic experiments demonstrated that the enzyme had an affinity for FBAL (Km 1.45 mM) comparable to taurine (Km 1.32 mM), but greater than glycine (Km 6.45 mM). Formation of FBAL-cholate was inhibited competitively by taurine (Ki 1.27 mM) and glycine (Ki 4.47 mM), suggesting that a single enzyme is responsible for conjugation of glycine, taurine and FBAL with bile acids. These data indicate that the formation of the FBAL-bile acid conjugates in patients receiving FUra results from high affinity of the bile acid conjugating enzyme for FBAL.     

A(3) adenosine receptor: a plausible therapeutic target for cardio-protection in diabetes

Diabetes mellitus categorized as type I and II, is a disease of pancreatic insulin, affecting blood glucose level in the body. Recent evidence suggests that cardiac diseases such as hypertension, coronary artery disease, congestive heart failure, and diabetic cardiomyopathy are associated with diabetes and hyperglycemia. The adenosine receptors (AR) have been reported to play an important role in the regulation of these diseases. Four adenosine receptors have been cloned and characterized from several different mammalian species. The receptors are named adenosine A(1), A(2A), A(2B), and A(3). The A(2A) and A(2B) receptors preferably interact with members of the Gs family of G proteins and the A(1) and A(3) receptors with Gi/o proteins. The ubiquitous levels of adenosine are found in each cell in normal conditions but in disease conditions its level has been shown to increase and activate G-protein mediated signaling pathway leading to artery constriction in cardiovascular diseases and diabetes. Various studies have demonstrated that A(3)AR is a potent cardioprotectant during myocardial ischemeia/ischemic reperfusion. Role of A(3)AR receptor as a possible cardioprotectant in diabetes is under investigation and studies have verified the involvement of cyclooxygenases (COXs) and NADPH oxidase pathways. This review summarizes the possible role of A(3)AR in cardiovascular disease and discusses advancement in the development of therapeutic agents targeting cardioprotection with discussion on recent patents on A(3) agonists that are being utilized in the clinical setting. We anticipate that detailed pharmacological studies of adenosine A(3) receptors could help in understanding the link between cardiovascular disease and diabetes and this can be utilized to develop newer therapies that selectively target A(3) receptor to overcome cardiac challenges.     

Evidence that (-)-7-hydroxy-4'-dimethylheptyl-cannabidiol activates a non-CB(1), non-CB(2), non-TRPV1 target in the mouse vas deferens

Previous experiments showed that R-(+)-WIN55212-induced inhibition of electrically-evoked contractions of mouse vasa deferentia could be antagonized by cannabidiol in a manner that appeared to be competitive but not to involve direct competition for established cannabinoid receptors. We have now discovered that (−)-7-hydroxy-4′-dimethylheptyl-cannabidiol (7-OH-DMH-CBD) inhibits electrically-evoked contractions of the vas deferens (EC₅₀ = 13.3 nM). This it appeared to do by acting on prejunctional neurones as 100 nM 7-OH-DMH-CBD did not attenuate contractile responses to phenylephrine or β,γ-methylene-ATP. Although 7-OH-DMH-CBD was antagonized by SR141716A, it was less susceptible to antagonism by this CB₁ receptor antagonist than R-(+)-WIN55212. 7-OH-DMH-CBD was also antagonized by cannabidiol (1 μM; apparent K_B = 222.2 nM) but not by the CB₂ receptor antagonist, SR144528 (32 nM), or by naloxone (300 nM), ruthenium red (1 μM) or capsazepine (10 μM). Yohimbine (100 nM) enhanced the ability of 7-OH-DMH-CBD to inhibit electrically-evoked contractions. R-(+)-WIN55212 was also potentiated by 100 nM yohimbine, possibly reflecting ongoing sequestration of G_i/o proteins from CB₁ receptors by ₂-adrenoceptors. Our results suggest that 7-OH-DMH-CBD may activate a neuronal target in the vas deferens that is not a CB₁, CB₂, TRPV1, opioid or ₂-adrenergic receptor but do not exclude the possibility that it also activates CB₁ receptors.     

Partial agonists for A(3) adenosine receptors

Selective agonists for A(3) adenosine receptors (ARs) could potentially be therapeutic agents for a variety of disorders, including brain and heart ischemic conditions, while partial agonists may have advantages over full agonists as a result of an increased selectivity of action. A number of structural determinants for A(3)AR activation have recently been identified, including the N(6)-benzyl group, methanocarba substitution of ribose, 2-chloro and 2-fluoro substituents, various 2'- and 3'-substitutions and 4'-thio substitution of oxygen. The 2-chloro substitution of CPA and R-PIA led to A(3) antagonism (CCPA) and partial agonism (Cl-R-PIA). 2-Chloroadenosine was a full agonist, while 2-fluoroadenosine was a partial agonist. Both 2'- and 3'- substitutions have a pronounced effect on its efficacy, although the effect of 2'-substitution was more dramatic. The 4-thio substitution of oxygen may also diminish efficacy, depending on other substitutions. Both N(6)-methyl and N(6)-benzyl groups may contribute to the A(3) affinity and selectivity; however, an N(6)-benzyl group but not an N(6)-methyl group diminishes A(3)AR efficacy. N(6)-benzyl substituted adenosine derivatives have similar potency for human and rat A(3)ARs while N(6)-methyl substitution was preferable for the human A(3)AR. The combination of 2-chloro and N(6)-benzyl substitutions appeared to reduce efficacy further than either modification alone. The A(2A)AR agonist DPMA was shown to be an antagonist for the human A(3)AR. Thus, the efficacy of adenosine derivatives at the A(3)AR appears to be more sensitive to small structural changes than at other subtypes. Potent and selective partial agonists for the A(3)AR could be identified by screening known adenosine derivatives and by modifying adenosine and the adenosine derivatives.     

Amino acid and fatty acid composition of an aqueous extract of Channa striatus (Haruan) that exhibits antinociceptive activity

1. The present study was performed in order to determine the amino acid and fatty acid composition of an aqueous extract of the freshwater fish Channa striatus, obtained by soaking (1:2, w/v) fresh fillets overnight in a chloroform:methanol (2:1, v/v) solvent, to elucidate the mechanism responsible for its antinociceptive activity and to clarify the relationship between the presence of the amino and fatty acids and the expected activity. 2. The aqueous extract was found to contain all amino acids with the major amino acids glycine, alanine, lysine, aspartic acid and proline making up 35.77 +/- 0.58, 10.19 +/- 1.27, 9.44 +/- 0.56, 8.53 +/- 1.15 and 6.86 +/- 0.78% of the total protein, respectively. 3. In addition, the aqueous extract was found to have a high palmitic acid (C16:0) content, which contributed approximately 35.93 +/- 0.63% to total fatty acids. The other major fatty acids in the aqueous extract were oleic acid (C18:1), stearic acid (C18:0), linoleic acid (C18:2) and arachidonic acid (C20:4), contributing 22.96 +/- 0.40, 15.31 +/- 0.33, 11.45 +/- 0.31 and 7.44 +/- 0.83% of total fatty acids, respectively. 4. Furthermore, the aqueous extract was demonstrated to possess concentration-dependent antinociceptive activity, as expected, when assessed using the abdominal constriction test in mice. 5. It is concluded that the aqueous extract of C. striatus contains all the important amino acids, but only some of the important fatty acids, which are suggested to play a key role in the observed antinociceptive activity of the extract, as well as in the traditionally claimed wound healing properties of the extract.     

Studies on the relation of γ-hydroxybutyric acid (GHB) to γ-aminobutyric acid (GABA): Evidence that GABA is not the sole source for GHB in rat brain

The effects of γ-aminobutyric acid (GABA)-α-oxoglutarate aminotransferase (GABA-T) inhibitors, l-glutamic acid decarboxylase (GAD) inhibitors, and antipetit mal anticonvulsants on γ-hydroxybutyric acid (GHB) and GABA were studied. Treatment with anticonvulsants and GABA-T inhibitors resulted in an increase in steady-state brain levels of both GHB and GABA. GAD inhibitors produced markedly decreased levels of brain GABA but no change in GHB concentrations. Studies of GHB derived exclusively from GABA showed that GABA-T inhibitors which produced an elevation of steady-state levels of GHB in brain also resulted in a decrease in GABA-derived GHB. Intracere-broventricular (i.c.v.) administration of GABA, putrescine, and 1,4-butanediol all produced significant elevations in brain GHB, but GABA-T inhibitors blocked this effect of GABA and putrescine. These data suggest that there may be another source for GHB in brain in addition to GABA and raise the possibility that 1,4-butanediol may be that source.     

Extracellular adenosine deprivation induces epithelial differentiation of HT29 cells: evidence for a concomitant adenosine A(1)/A(2) receptor balance regulation

HT29 cells display an undifferentiated phenotype in culture. However, numerous treatments are able to induce both epithelial differentiation and cell growth inhibition. We have previously demonstrated that adenosine and its analogues act through specific adenosine receptors to modulate cell proliferation in HT29 and other human colon adenocarcinoma cell lines. Among the treatments tested, the most potent inhibition of HT29 cell growth was induced by deprivation of extracellular adenosine using adenosine deaminase. Here, we investigated the capacity of adenosine deaminase to initiate epithelial differentiation. After 1 month of daily addition of 10 U/ml adenosine deaminase to the culture medium, HT29 cells were cloned by limited dilution. Among the clones obtained, we focused our attention on clone 13. Microscopic visualization and proliferation studies indicated that cells from this clone grew very slowly and in a pseudo-monolayer, in marked contrast with the situation observed in the mother HT29 cell line. In addition, clone 13 cells displayed epithelial features that mimic the enterocytic differentiation of Caco-2 cells. These modifications were accompanied by dramatic changes in the activity of adenosine receptors, as demonstrated by pharmacological studies. In contrast to the original HT29 cells, clone 13 as well as Caco-2 cells displayed (i) a very low number of adenosine A(1) receptors, and (ii) increases in intracellular cAMP levels when challenged with adenosine analogues. It is hypothesized that a loss of adenosine A(1) receptors, with no change or a concomitant increase in adenosine A(2) receptors, results in the emergence of adenosine A(2) receptor-mediated differentiation and inhibition of proliferation, through a cAMP-dependent pathway.     

Evidence for the presence of A(1) adenosine receptors in the aorta of spontaneously hypertensive rats.

1. Isolated aortic rings (endothelium-intact and -denuded) from spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) rats were used in this study to examine the vasoactive effects of various adenosine analogues. 2. In phenylephrine contracted aortic rings, concentration-response curves were constructed by cumulative additions (10(-11) - 10(-5) M) of (2S)-N(6)-[2-endo-Norbornyl] adenosine (ENBA), N(6)-cyclopentyladenosine (CPA), R-N(6)-(2-phenylisopropyl) adenosine (R-PIA), 2-p-(-2-carboxyethyl) phenethylamino-5'-N-thylcarboxamido adenosine (CGS-21680). 3. A non-specific adenosine receptor agonist 2-chloroadenosine (CAD) resulted in biphasic response with a small contraction at lower concentrations (10(-9) - 10(-8) M) followed by a significant relaxation at higher concentration in endothelium-intact SHR tissues, suggesting presence of both A(1) and A(2) adenosine receptors in SHR aorta. However, only relaxation was observed in WKY. 4. Contractile response in SHR had the following rank order of potency: ENBA>CPA>R-PIA>CAD. The relaxation response in SHR and WKY had the following rank order of potency: CGS 21680>CAD>R-PIA>CPA>ENBA. 5. Removal of endothelium abolished the adenosine analogue induced contractions in SHR aorta and attenuated the vasorelaxation responses in the WKY and SHR. 6. The contractile response in SHR was abolished by A(1) adenosine receptor antagonist N(6)-endonorbornan-2-yl-9-methyladenine (N-0861). A(2) adenosine receptor antagonist, 3,7-dimethyl-1-proparglyxanthine (DMPX) did not affect the contraction response of adenosine analogues. 7. Endothelium-dependent contractions elicited by A(1) receptor agonists were blocked by indomethacin and by free radical scavengers. 8. These data suggest that the contractile response to adenosine analogues in SHR aorta is probably mediated by free radicals which are generated through the increased cyclo-oxygenase activity occurring in the vascular endothelium of SHR but not the WKY rats.     

Structure-based design of a new series of D-glutamic acid based inhibitors of bacterial UDP-N-acetylmuramoyl-L-alanine:D-glutamate ligase (MurD)

MurD ligase is one of the key enzymes participating in the intracellular steps of peptidoglycan biosynthesis and constitutes a viable target in the search for novel antibacterial drugs to combat bacterial drug-resistance. We have designed, synthesized, and evaluated a new series of D-glutamic acid-based Escherichia coli MurD inhibitors incorporating the 5-benzylidenethiazolidin-4-one scaffold. The crystal structure of 16 in the MurD active site has provided a good starting point for the design of structurally optimized inhibitors 73-75 endowed with improved MurD inhibitory potency (IC(50) between 3 and 7 μM). Inhibitors 74 and 75 showed weak activity against Gram-positive Staphylococcus aureus and Enterococcus faecalis. Compounds 73-75, with IC(50) values in the low micromolar range, represent the most potent D-Glu-based MurD inhibitors reported to date.     

3-m-bromoacetylamino benzoic acid ethyl ester: a new cancericidal agent that activates the apoptotic pathway through caspase-9

The mechanism underlying the cancericidal activity of 3-m-bromoacetylamino benzoic acid ethyl ester (3-BAABE) was investigated. 3-BAABE exerted a strong cancericidal effect on human leukemia and lymphoma cells (IC(50) < 0.2 microgram/mL) and on cell lines of prostate, colon, ductal, and kidney cancer (IC(50) 0.8 to 0.88 microgram/mL). Multiple drug resistance (MDR) had no effect on the susceptibility of human lymphoma cells to 3-BAABE, since Daudi/MDR(20) and wild-type Daudi cells had a similar susceptibility to the cytotoxic effect of 3-BAABE. The cancericidal effect of 3-BAABE, which was not associated with changes in the cell cycle, was mediated by apoptosis. Thus, cells exposed to 3-BAABE displayed the DNA fragmentation ladder characteristic for apoptosis, associated with a marked increase of the activity of apoptosis effector caspases-3 and -6, which was followed by proteolytic cleavage of DNA fragmentation factor (DFF) and poly(ADP-ribose) polymerase (PARP). Exposure of tumor cells to 3-BAABE increased the activity of apical caspase-9, but had no effect on caspase-8. Complete inhibition of 3-BAABE-induced apoptosis was exerted by LEHD-FMK, a caspase-9 inhibitor. DEVD-FMK, a caspase-3 inhibitor, and VEID-FMK, a caspase-6 inhibitor, partially inhibited 3-BAABE-induced apoptosis, whereas exposure to IETD-FMK, a caspase-8 inhibitor, had no effect. The fragmentation and elevated activity of caspase-9 in 3-BAABE-treated cells and the fact that only an inhibitor of caspase-9 abrogated 3-BAABE-induced apoptosis indicate that 3-BAABE is a distinctive compound that elicits apoptosis through a pathway that is limited specifically to activation of apical caspase-9.     

Adrenomedullin induces direct (endothelium-independent) vasorelaxations and cyclic adenosine monophosphate elevations that are synergistically enhanced by brain natriuretic peptide in isolated rings of rat thoracic aorta

Our laboratory previously demonstrated that nitric oxide and natriuretic peptides can synergistically enhance cAMP elevations and vasorelaxations in rat aortic rings induced by calcitonin gene-related peptide, likely involving cyclic guanosine monophosphate (cGMP)-mediated inhibition of type-3 phosphodiesterase (PDE3). It was predicted that this cellular mechanism may also serve as a point of synergism between adrenomedullin (ADM) and brain natriuretic peptide (BNP) in aortic smooth muscle cells. The current study shows that ADM (100 nM)-induced vasorelaxations in isolated aortic rings of Sprague-Dawley rats are dependent on endothelium (34.1 +/- 4.2% relaxation with endothelium versus 3.0 +/- 0.6% relaxation without endothelium; P < 0.001). To determine interactions between ADM and BNP in smooth muscle cells without interference from endothelium-derived factors, further studies used aortic rings denuded of endothelium. Pretreatment with BNP (1 nM), which elevated cGMP levels 1.6 fold, uncovered direct vasorelaxant effects of ADM in endothelium-denuded rings, showing 5.6 +/- 1.8%, 20.9 +/- 6.1%, and 55 +/- 9.4% relaxations with ADM at 1, 10, and 100 nM, respectively (n = 6). ADM (100 nM) significantly (P < 0.05) increased cyclic adenosine monophosphate (cAMP) levels in denuded aortic rings pretreated with BNP (1 nM), but not in denuded rings without BNP. Quazinone (20 microM), a PDE3 inhibitor, caused similar enhancement of direct cAMP elevations to ADM (100 nM). The data indicate vasodilatory synergism between ADM and BNP in aorta, likely mediated by enhanced accumulation of cAMP in smooth muscle cells resulting from BNP/cGMP-induced inhibition of PDE3. This synergistic mechanism may be especially important in subjects with dysfunctional endothelium, in which BNP may uncover direct vasorelaxant effects of ADM in arteries that normally require healthy (nitric oxide-releasing) endothelium for ADM-induced vasorelaxations to occur.     

Microsomal oxidation of dodecylthioacetic acid (a 3-thia fatty acid) in rat liver.

[1-14C]Dodecylthioacetic acid (DTA), a 3-thia fatty acid, is omega (omega-1)-hydroxylated and sulfur oxygenated at about equal rates in rat liver microsomes. In prolonged incubations DTA is converted to omega-hydroxydodecylsulfoxyacetic acid. omega-Hydroxylation of DTA is catalysed by cytochrome P450IVA1 (or a very closely related isoenzyme in the same gene family), the fatty acid omega-hydroxylating enzyme. It is absolutely dependent on NADPH and inhibited by CO, and lauric acid is a competing substrate. omega-Hydroxylation of DTA is increased by feeding tetradecylthioacetic acid (TTA), a 3-thia fatty acid, for 4 days to rats. omega-Hydroxylation of [1-14C]lauric acid is also induced by TTA and other 3-thia carboxylic acids. A close relationship was observed between induction of microsomal omega-hydroxylation of fatty acid and palmitoyl-CoA hydrolase activity. DTA is omega-hydroxylated at about the same rate as the physiological substrate lauric acid. The sulfur oxygenation of DTA is catalysed by liver microsomal flavin-containing monooxygenase (FMO) (EC 1.14.13.8). It is dependent on either NADH or NADPH. The Km value for NADH was approx. five times larger than the Km value for NADPH. It is inhibited by methimazole and not affected by CO. It is not induced by TTA.     

2,5'-Disubstituted adenosine derivatives: evaluation of selectivity and efficacy for the adenosine A(1), A(2A), and A(3) receptor.

Novel 2,5'-disubstituted adenosine derivatives were synthesized in good overall yields starting from commercially available guanosine. Binding affinities were determined for rat adenosine A(1) and A(2A) receptors and human A(3) receptors. E(max) values were determined for the stimulation or inhibition of cAMP production in CHO cells expressing human adenosine A(2A) (EC(50) values as well) or A(3) receptors, respectively. The compounds displayed affinities in the nanomolar range for both the adenosine A(2A) and A(3) receptor, without substantial preference for either receptor. The derivatives with a 2-(1-hexynyl) group had the highest affinities for both receptors; compound 4 (2-(1-hexynyl)adenosine) had the highest affinity for the adenosine A(2A) receptor with a K(i) value of 6 nM (A(3)/A(2A) selectivity ratio of approximately 3), whereas compound 37 (2-(1-hexynyl)-5'-S-methyl-5'-thioadenosine) had the highest affinity for the adenosine A(3) receptor with a K(i) value of 15 nM (A(2A)/A(3) selectivity ratio of 4). In general, compounds with a relatively small 5'-S-alkyl-5'-thio substituent (methyl-5'-thio) displayed the highest affinities for both the adenosine A(2A) and A(3) receptor; the larger ones (n- or i-propyl-5'-thio) increased the selectivity for the adenosine A(3) receptor. The novel compounds were also evaluated in cAMP assays for their (partial) agonistic behavior. Overall, the disubstituted derivatives behaved as partial agonists for both the adenosine A(2A) and A(3) receptor. The compounds showed somewhat higher intrinsic activities on the adenosine A(2A) receptor than on the A(3) receptor. Compounds 37, 40 and 45, 48, with either a 5'-S-methyl-5'-thio or a 5'-S-i-propyl-5'-thio substituent had the lowest intrinsic activities on the adenosine A(2A) receptor. For the A(3) receptor, compounds 34, 35, 38, 39, and 46, 47, with a 5'-S-ethyl-5'-thio or a 5'-S-n-propyl-5'-thio substituent had the lowest intrinsic activities.