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K A Jacobson

Publications and source records attributed to K A Jacobson.

At least 55 records · Page 3Linked to original sources
PubMed1999

Selective A(3) adenosine receptor antagonists: water-soluble 3, 5-diacyl-1,2,4-trialkylpyridinium salts and their oxidative generation from dihydropyridine precursors.

A(3) adenosine receptor antagonists are sought for their potential antiinflammatory, antiasthmatic, and antiischemic properties. We have found that 3,5-diacyl-1,2,4-trialkyl-6-phenylpyridinium derivatives constitute a novel class of selective A(3) adenosine receptor antagonists. The structure-activity relationships of this class of antagonists, incorporating the 3-thioester, have been explored. The most potent analogue in this group was 2, 4-diethyl-1-methyl-3-(ethylsulfanylcarbonyl)-5-ethyloxycarbonyl -6-phe nylpyridinium iodide (11), which had an equilibrium inhibition constant (K(i)) value of 219 nM at human A(3) receptors (binding of [(125)I]AB-MECA (N(6)-(4-amino-3-iodobenzyl)-5'-N-methylcarbamoyladenosine)) expressed in Chinese hamster ovary (CHO) cells and >10 microM at rat brain A(1) and A(2A) receptors and at recombinant human A(2B) receptors. Compound 11 could be generated through oxidation of the corresponding 3,5-diacyl-1,2,4-trialkyl-6-phenyl-1,4-dihydropyridine, 24, with iodine or in the presence of rat brain homogenates. A 6-cyclopentyl analogue was shown to increase affinity at human A(3) receptors upon oxidation from the 1-methyl-1,4-dihydropyridine analogue, 25, to the corresponding pyridinium derivative, 23 (K(i) 695 nM), suggesting a prodrug scheme. Homologation of the N-methylpyridinium derivatives to N-ethyl and N-propyl at the 1-position caused a progressive reduction in the affinity at A(3) receptors. Modifications of the alkyl groups at the 2-, 3-, 4-, and 5-positions failed to improve potency in binding at A(3) receptors. The pyridinium antagonists are not as potent as other recently reported, selective A(3) receptor antagonists; however, they display uniquely high water solubility (43 mM for 11). Compound 11 antagonized the inhibition of adenylate cyclase elicited by IB-MECA in CHO cells expressing the human A(3) adenosine receptor, with a K(B) value of 399 nM, and did not act as an agonist, demonstrating that the pyridinium salts are pure antagonists.

Adenylyl Cyclase Inhibitors↗
PubMed1999

Chiral resolution and stereospecificity of 6-phenyl-4-phenylethynyl- 1,4-dihydropyridines as selective A(3) adenosine receptor antagonists.

Racemic 6-phenyl-4-phenylethynyl-1,4-dihydropyridine derivatives have been shown to be highly selective A(3) adenosine receptor antagonists (Jiang et al. J. Med. Chem. 1997, 40, 2596-2608). Methods for resolving the optical isomers at the C4 position, involving selective crystallization or chromatographic separation of diastereomeric ester derivatives, have been developed. Optically pure glycerol and threitol derivatives were used as chiral auxiliary groups for ester formation at the 3-position, resulting in diastereomeric mixtures of dihydropyridines. Esterification of a 6-phenyl-4-phenylethynyl-1,4-dihydropyridine derivative at the 3-position with a chiral, protected glycerol moiety, (S)-(+)-2, 2-dimethyl-1,3-dioxolane-4-methanol, allowed the selective crystallization of a pure diastereomer, 9. The (1)H NMR spectrum of 9 using the lanthanide shift reagent Eu(fod)(3) indicated optical purity, and the (4S,2'R)-configuration was assigned using X-ray crystallography. The noncrystalline (4R,2'R)-isomer 10 was also isolated and shown to be 3-fold more potent than the (4S,2'R)-isomer in binding to A(3) receptors. The 2,2-dimethyl-1,3-dioxolane moiety also served as a protected form of a diol, which showed selective reactivity versus a 5-ethyl ester in basic transesterification reactions. A racemic 5-carboxylic acid derivative could not be resolved through crystallization of diastereomeric salts. Enantiomers of 5-benzyl 3-ethyl 2-methyl-6-phenyl-4-phenylethynyl-1, 4-dihydropyridine-3,5-dicarboxylate (2) were obtained via an ester derived from (4R,5R)-(-)-2,3-O-isopropylidene-D-threitol at the 3-position, which was resolved using HPLC, and each diastereomer was subsequently deprotected in acidic conditions. The resulting diols were exchanged for ethyl ester groups by base-catalyzed transesterification. The binding of pure enantiomers of 2 at A(3) adenosine receptors indicated a 35-fold stereoselectivity for the (4S)-isomer 21. A receptor docking hypothesis, using a previously derived human A(3) receptor model, shows the bulkier of the two ester groups (5-Bn) of 21 oriented toward the exofacial side and the 4-position phenylethynyl group situated between transmembrane helical domain TM6 and TM7.

Animals↗
PubMed1999

Characterization of "mini-nucleotides" as P2X receptor agonists in rat cardiomyocyte cultures. An integrated synthetic, biochemical, and theoretical study.

The design and synthesis of "mini-nucleotides", based on a xanthine-alkyl phosphate scaffold, are described. The physiological effects of the new compounds were evaluated in rat cardiac cell culture regarding Ca(2+) elevation and contractility. The results indicate biochemical and physiological profiles similar to those of ATP, although at higher concentrations. The biological target molecules of these "mini-nucleotides" were identified by using selective P2-R and A(1)-R antagonists and P2-R subtype selective agonists. On the basis of these results and of experiments in Ca(2+) free medium, in which [Ca(2+)](i) elevation was not observed, we concluded that interaction of the analogues is likely with P2X receptor subtypes, which causes Ca(2+) influx. Theoretical calculations analyzing electronic effects within the series of xanthine-alkyl phosphates were performed on reduced models at quantum mechanical levels. Calculated dipole moment vectors, electrostatic potential maps, and volume parameters suggest an explanation for the activity or inactivity of the synthesized derivatives and predict a putative binding site environment for the active agonists. Xanthine-alkyl phosphate analogues proved to be selective agents for activation of P2X-R subtypes, whereas ATP activated all P2-R subtypes in cardiac cells. Therefore, these analogues may serve as prototypes of selective drugs aiming at cardiac disorders mediated through P2X receptors.

Animals↗
PubMed1999

Additive effects of late preconditioning produced by monophosphoryl lipid A and the early preconditioning mediated by adenosine receptors and KATP channel.

BACKGROUND: The cardioprotective effect of preconditioning can be exerted within 1 to 2 hours after initial ischemia, termed classical or early preconditioning, or can reappear 24 hours later as second window or late preconditioning. The objective of this study was to study the interaction between late and early preconditioning and to determine the potential underlying mechanism. METHODS AND RESULTS: Adenosine receptor agonists and a KATP channel opener were used to achieve early preconditioning, and Monophosphoryl lipid A (MLA) was used to induce late preconditioning. Cultured chick ventricular myocytes were used as a myocyte model of simulated ischemia and preconditioning. Prior treatment of the myocyte with MLA caused a dose-dependent decrease in the ischemia-induced myocyte injury 24 hours later, consistent with a late preconditioning effect. L-NMMA, glibenclamide, or 5-hydroxydecanoic acid administered during the ischemia blocked the MLA effect. Twenty four hours after MLA treatment, a 5-minute exposure to ischemia, adenosine, adenosine A1 agonist CCPA, or A3 agonist resulted in less myocyte injury during the subsequent prolonged ischemia, as compared with cells pretreated with the vehicle and subsequently exposed to the same early preconditioning stimuli. In addition to its ability to enhance the early preconditioning effect by A1 and A3 agonists, MLA pretreatment also increased the phorbol ester- and pinacidil-mediated early preconditioning effect. CONCLUSIONS: This study defined a novel interaction in which the cardioprotective effect of early preconditioning is additive to that of late preconditioning and raised the possibility that both agents can be used as combined therapy in the treatment of ischemic heart disease.

Adenosine↗
PubMed1999

The role of amino acids in extracellular loops of the human P2Y1 receptor in surface expression and activation processes.

The P2Y1 receptor is a membrane-bound G protein-coupled receptor stimulated by adenine nucleotides. Using alanine scanning mutagenesis, the role in receptor activation of charged amino acids (Asp, Glu, Lys, and Arg) and cysteines in the extracellular loops (EL) of the human P2Y1 receptor has been investigated. The mutant receptors were expressed in COS-7 cells and measured for stimulation of phospholipase C induced by the potent agonist 2-methylthioadenosine-5'-diphosphate (2-MeSADP). In addition to single point mutations, all receptors carried the hemagglutinin epitope at the N- terminus for detection of cell-surface expression. The C124A and C202A mutations, located near the exofacial end of transmembrane helix 3 and in EL2, respectively, ablated phospholipase C stimulation by </=100 microM 2-MeSADP. Surface enzyme-linked immunosorbent assay detection of both mutant receptors showed <10% expression, suggesting that a critical disulfide bridge between EL2 and the upper part of transmembrane 3, as found in many other G protein-coupled receptors, is required for proper trafficking of the P2Y1 receptor to the cell surface. In contrast, the C42A and C296A mutant receptors (located in the N-terminal domain and EL3) were activated by 2-MeSADP, but the EC50 values were >1000-fold greater than for the wild-type receptor. The double mutant receptor C42A/C296A exhibited no additive shift in the concentration-response curve for 2-MeSADP. These data suggest that Cys42 and Cys296 form another disulfide bridge in the extracellular region, which is critical for activation. Replacement of charged amino acids produced only minor changes in receptor activation, with two remarkable exceptions. The E209A mutant receptor (EL2) exhibited a >1000-fold shift in EC50. However, if Glu209 were substituted with amino acids capable of hydrogen bonding (Asp, Gln, or Arg), the mutant receptors responded like the wild-type receptor. Arg287 in EL3 was impaired similarly to Glu209 when substituted by alanine. Substitution of Arg287 by lysine, another positively charged residue, failed to fully restore wild-type activity.

Amino Acid Sequence↗
PubMed1999

Structure-activity relationships of bisphosphate nucleotide derivatives as P2Y1 receptor antagonists and partial agonists.

The P2Y1 receptor is present in the heart, in skeletal and various smooth muscles, and in platelets, where its activation is linked to aggregation. Adenosine 3',5'- and 2',5'-bisphosphates have been identified as selective antagonists at the P2Y1 receptor (Boyer et al. Mol. Pharmacol. 1996, 50, 1323-1329) and have been modified structurally to increase receptor affinity (Camaioni et al. J. Med. Chem. 1998, 41, 183-190). We have extended the structure-activity relationships to a new series of deoxyadenosine bisphosphates with substitutions in the adenine base, ribose moiety, and phosphate groups. The activity of each analogue at P2Y1 receptors was determined by measuring its capacity to stimulate phospholipase C in turkey erythrocyte membranes (agonist effect) and to inhibit phospholipase C stimulation elicited by 10 nM 2-(methylthio)adenosine 5'-diphosphate (antagonist effect). 2'-Deoxyadenosine bisphosphate analogues containing halo, amino, and thioether groups at the 2-position of the adenine ring were more potent P2Y1 receptor antagonists than analogues containing various heteroatom substitutions at the 8-position. An N6-methyl-2-chloro analogue, 6, was a full antagonist and displayed an IC50 of 206 nM. Similarly, N6-methyl-2-alkylthio derivatives 10, 14, and 15 were nearly full antagonists of IC50 < 0.5 microM. On the ribose moiety, 2'-hydroxy, 4'-thio, carbocyclic, and six-membered anhydrohexitol ring modifications have been prepared and resulted in enhanced agonist properties. The 1,5-anhydrohexitol analogue 36 was a pure agonist with an EC50 of 3 microM, i.e., similar in potency to ATP. 5'-Phosphate groups have been modified in the form of triphosphate, methyl phosphate, and cyclic 3',5'-diphosphate derivatives. The carbocyclic analogue had enhanced agonist efficacy, and the 5'-O-phosphonylmethyl modification was tolerated, suggesting that deviations from the nucleotide structure may result in improved utility as pharmacological probes. The N6-methoxy modification eliminated receptor affinity. Pyrimidine nucleoside 3', 5'-bisphosphate derivatives were inactive as agonists or antagonists at P2Y receptor subtypes.

Animals↗
PubMed1999

Protection against ischemic damage by adenosine amine congener, a potent and selective adenosine A1 receptor agonist.

Although the selectivity and potency of adenosine amine congener (ADAC) at adenosine A1 receptors are similar to other highly selective agonists at this receptor type, the chemical structure of the N6 substituent is completely different. We now demonstrate that the characteristics of the therapeutic profile of ADAC are distinct from those observed during our previous studies of adenosine A1 receptor agonist-mediated neuroprotection. Most significantly, chronic treatment with low microgram doses of ADAC (25-100 microg/kg) protects against both mortality and neuronal damage induced by 10 min bilateral carotid occlusion in gerbils. At higher chronic doses, the statistical significance of the protective effect is lost. Acute preischemic administration of the drug at 75-200 microg/kg also results in a statistically significant reduction of postischemic mortality and morbidity. These data indicate that, contrary to other adenosine A1 receptor agonists whose chronic administration enhances postocclusive brain damage, ADAC may be a promising agent in treatment of both acute (e.g., cerebral ischemia) and chronic (seizures) disorders of the central nervous system in which adenosine A receptors appear to be involved.

Adenosine↗
PubMed1999

Role of the extracellular loops of G protein-coupled receptors in ligand recognition: a molecular modeling study of the human P2Y1 receptor.

The P2Y1 receptor is a G protein-coupled receptor (GPCR) and is stimulated by extracellular ADP and ATP. Site-directed mutagenesis of the three extracellular loops (ELs) of the human P2Y1 receptor indicates the existence of two essential disulfide bridges (Cys124 in EL1 and Cys202 in EL2; Cys42 in the N-terminal segment and Cys296 in EL3) and several specific ionic and H-bonding interactions (involving Glu209 and Arg287). Through molecular modeling and molecular dynamics simulations, an energetically sound conformational hypothesis for the receptor has been calculated that includes transmembrane (TM) domains (using the electron density map of rhodopsin as a template), extracellular loops, and a truncated N-terminal region. ATP may be docked in the receptor, both within the previously defined TM cleft and within two other regions of the receptor, termed meta-binding sites, defined by the extracellular loops. The first meta-binding site is located outside of the TM bundle, between EL2 and EL3, and the second higher energy site is positioned immediately underneath EL2. Binding at both the principal TM binding site and the lower energy meta-binding sites potentially affects the observed ligand potency. In meta-binding site I, the side chain of Glu209 (EL2) is within hydrogen-bonding distance (2.8 A) of the ribose O3', and Arg287 (EL3) coordinates both alpha- and beta-phosphates of the triphosphate chain, consistent with the insensitivity in potency of the 5'-monophosphate agonist, HT-AMP, to mutation of Arg287 to Lys. Moreover, the selective reduction in potency of 3'NH2-ATP in activating the E209R mutant receptor is consistent with the hypothesis of direct contact between EL2 and nucleotide ligands. Our findings support ATP binding to at least two distinct domains of the P2Y1 receptor, both outside and within the TM core. The two disulfide bridges present in the human P2Y1 receptor play a major role in the structure and stability of the receptor, to constrain the loops within the receptor, specifically stretching the EL2 over the opening of the TM cleft and thus defining the path of access to the binding site.

Amino Acid Sequence↗
PubMed1999

Synthesis, CoMFA analysis, and receptor docking of 3,5-diacyl-2, 4-dialkylpyridine derivatives as selective A3 adenosine receptor antagonists.

3,5-Diacyl-2,4-dialkyl-6-phenylpyridine derivatives have been found to be selective antagonists at both human and rat A3 adenosine receptors (Li et al. J. Med. Chem. 1998, 41, 3186-3201). In the present study, ring-constrained, fluoro, hydroxy, and other derivatives in this series have been synthesized and tested for affinity at adenosine receptors in radioligand binding assays. Ki values at recombinant human and rat A3 adenosine receptors were determined using [125I]AB-MECA (N6-(4-amino-3-iodobenzyl)-5'-N-methylcarbamoyladenosine). Selectivity for A3 adenosine receptors was determined vs radioligand binding at rat brain A1 and A2A receptors, and structure-activity relationships at various positions of the pyridine ring (the 3- and 5-acyl substituents and the 2- and 4-alkyl substituents) were probed. At the 5-position inclusion of a beta-fluoroethyl (7) or a gamma-fluoropropyl ester (26) was favorable for human A3 receptor affinity, resulting in Ki values of 4.2 and 9.7 nM, respectively, while the pentafluoropropyl analogue was clearly less potent at human A3 receptors. At the 2-, 3-, and 4-positions, fluoro or hydroxy substitution failed to enhance potency and selectivity at human A3 receptors. Several analogues were nearly equipotent at rat and human A3 receptors. To further define the pharmacophore conformationally, a lactam, a lactone, and thiolactones were tested in adenosine receptor binding. The most potent analogue in this group was compound 34, in which a thiolactone was formed between 3- and 4-positions and which had a Ki value of 248 nM at human A3 receptors. Using affinity data and a general pharmacophore model for A3 adenosine receptor antagonists recently proposed, we applied comparative molecular field analysis (CoMFA) to obtain a three-dimensional quantitative structure-activity relationship for pyridine derivatives, having good predictability (r2pred = 0.873) for compounds in the test set. A rhodopsin-based model of the human A3 receptor was built, and the pyridine reference ligand 2,3,4, 5-tetraethyl-6-phenyl-pyridine-3-thiocarboxylate-5-carboxylate (MRS 1476) was docked in the putative ligand binding site. Interactions between receptor transmembrane domains and the steric and the electrostatic contour plots obtained from the CoMFA analysis were analyzed.

Animals↗
PubMed1999

Chronic administration of adenosine A3 receptor agonist and cerebral ischemia: neuronal and glial effects.

We have previously shown that chronic administration of the selective A3 receptor agonist N6-(3-iodobenzyl)-5'-N-methylcarboxoamidoadenosine (IB-MECA) leads to a significant improvement of postocclusive cerebral blood flow, and protects against neuronal damage and mortality induced by severe forebrain ischemia in gerbils. Using immunocytochemical methods we now show that chronic with IB-MECA results in a significant preservation of ischemia-sensitive microtubule associated protein 2 (MAP-2), enhancement of the expression of glial fibrillary acidic protein (GFAP), and a very intense depression of nitric oxide synthase in the brain of postischemic gerbils. These changes demonstrate that the cerebroprotective actions of chronically administered IB-MECA involve both neurons and glial cells, and indicate the possibility of distinct mechanisms that are affected in the course of chronic administration of the drug.

Adenosine↗
PubMed1999

Adenosine-induced cell death: evidence for receptor-mediated signalling.

Adenosine modulates the proliferation, survival and apoptosis of many different cell types, ranging from epithelial, endothelial and smooth muscle cells, to cells of the immune and neural lineages. In this review, we critically discuss the available in vitro and in vivo data which support a role for adenosine in both development-associated apoptosis, and in diseases characterized by either pathologically increased cell death (e.g., ischemia, trauma and aging-associated neurodegeneration) or abnormally reduced spontaneous apoptosis (e.g., cancer). Particular emphasis is given to the possible role of extracellular adenosine receptors, since these may represent novel and attractive molecular targets for the pharmacological modulation of apoptosis. In some instances, adenosine-induced cell death has been demonstrated to require entry of the nucleoside inside cells; however, in many other cases, activation of specific adenosine extracellular receptors has been demonstrated. Of the four G protein-coupled adenosine receptors so far identified, the A2A and the A3 receptors have been specifically implicated in modulation of cell death. For the A3 receptor, results obtained by exposing both cardiomyocytes and brain astrocytes to graded concentrations of selective agonists suggest induction of both cell protection and cell death. Such opposite effects, which likely depend on the degree of receptor activation, may have important therapeutic implications in the pharmacological modulation of cardiac and brain ischemia. For the A2A receptor, recent intriguing data suggest a specific role in immune cell death and immunosuppression, which may be relevant to both adenosine-deaminase-immunodeficiency syndrome (a pathology characterized by accumulation of adenosine to toxic levels) and in tumors where induction of apoptosis via activation of specific extracellular receptors may be desirable. Finally, preliminary data suggest that, in a similar way to the adenosine-deaminase-immunodeficiency syndrome, the abnormal accumulation of adenosine in degenerative muscular diseases may contribute to muscle cell death. Although the role of adenosine receptors in this effect still remains to be determined, these data suggest that adenosine-induced apoptosis may also represent a novel pathogenic pathway in muscular dystrophies.

Journal Article↗
PubMed1999

Uptake of glucose analogs reflects the rate of contraction of cultured myocytes.

The present study demonstrates that: a) adenosine and R-N6-(2-phenylisopropyl)-adenosine (R-PIA, A1 and A3 adenosine receptor agonist) inhibited [3H]deoxyglucose uptake or [3H]3-O-methyl-D-glucose uptake; b) sugar uptake reflects the rate of contraction in cardiac cultures; c) [3H]deoxyglucose uptake or [3H]3-O-methyl-D-glucose uptake are useful quantitative probes for beating rate evaluation. A 25-40% decrease in [3H]deoxyglucose uptake (p < 0.01) was obtained following 13-21 min treatment with 100 microM adenosine together with 1 microM dipyridamole or with 10 microM R-PIA, which inhibited spontaneous contractions. Adenosine (10 microM) attenuated spontaneous beating rate and inhibited approximately 55% of the [3H]deoxyglucose uptake following 22 h treatment (p < 0.01). 1 microM R-PIA also attenuated beating rate following either a short (1 min) or long (24 h) application and decreased [3H]deoxyglucose uptake by 20-30% (p < 0.01) during 0.5-24 h of treatment. A 157 +/- 9% and 205 +/- 11% increase (p < 0.01) in [3H]deoxyglucose uptake was obtained at 27 and 37 degrees C, respectively, compared with the uptake at 17 degrees C, which completely inhibited spontaneous contractions. Similar results [33 +/- 6% (p < 0.01) and 21 +/- 8% (p < 0.05) inhibition in [3H]deoxyglucose uptake] were obtained following 2 and 22 h of carbamylcholine treatment, respectively. This treatment also reduced spontaneous contractions. [3H] 3-O-Methyl-D-glucose uptake also decreased by 31 +/- 12% (p < 0.05) as a result of the arrest of contractions by adenosine. Elevations of 90 +/- 13% and 34 +/- 11% (p < 0.01) in [3H]deoxyglucose uptake were obtained following treatment with isoprenaline after 2 and 22 h application, respectively. It is concluded that adenosine and R-PIA inhibited [3H]deoxyglucose uptake or [3H] 3-O-methyl-D-glucose uptake in rat heart culture and that there is a linkage between the rate of cardiac contractions in culture and sugar uptake.

3-O-Methylglucose↗
PubMed1999

Adenosine and ischemic preconditioning.

Adenosine is released in large amounts during myocardial ischemia and is capable of exerting potent cardioprotective effects in the heart. Although these observations on adenosine have been known for a long time, how adenosine acts to achieve its anti-ischemic effect remains incompletely understood. However, recent advances on the chemistry and pharmacology of adenosine receptor ligands have provided important and novel information on the function of adenosine receptor subtypes in the cardiovascular system. The development of model systems for the cardiac actions of adenosine has yielded important insights into its mechanism of action and have begun to elucidate the sequence of signalling events from receptor activation to the actual exertion of its cardioprotective effect. The present review will focus on the adenosine receptors that mediate the potent anti-ischemic effect of adenosine, new ligands at the receptors, potential molecular signalling mechanisms downstream of the receptor, mediators for cardioprotection, and possible clinical applications in cardiovascular disorders.

Adenosine↗
PubMed1999

Use of the triazolotriazine [3H]ZM 241385 as a radioligand at recombinant human A2B adenosine receptors.

Radiolabeled ZM 241385 (4-(2-[7-amino-2- ¿furyl¿¿1,2,4¿triazolo¿2,3-a¿¿1,3,5¿triazin-5-ylaminoethyl)p henol), has previously been used as a high affinity radioligand for the labeling of A2A adenosine receptors in cell membranes. Another subtype, the A2B receptor, is the least well-defined subtype of adenosine receptors and lacks selective pharmacological probes. In the present study, we have used [3H]ZM 241385 as a radioligand to label recombinant human A2B adenosine receptors in HEK-293 cell membranes, that do not express A2A adenosine receptors, and found that the pharmacological profile is consistent with the SAR of A2B receptors. Saturable, specific binding (Kd 33.6 nM, Bmax 4.48 pmol/mg protein) that was best described by a one-site model was found, and specific binding was approximately 75% of total binding. [3H]ZM 241385 binding was displaceable by a large number of compounds known to interact with A2B receptors; thus, this method has promise as a tool in the search for agonists and antagonists selective for this subtype. Xanthine analogs, which are antagonists, proved to be the most potent displacers. The Ki of XAC, xanthine amine congener, was 12.3 nM, while CPX (8-cyclopentyl-1,3-dipropylxanthine) was less potent. The non-selective triazoloquinazoline antagonist CGS 15943 (Ki 16.4 nM), which is similar in structure to ZM 241385, was slightly less potent than XAC. The non-xanthine A2B-antagonist alloxazine displaced [3H]ZM 241385-binding with a Ki of 462 nM, similar to its affinity in functional assays. Adenosine derivatives known to activate this receptor subtype, such as NECA (5'-N-ethylcarboxamidoadenosine) and R-PIA (N6-phenylisopropyladenosine), were considerably less potent than the 8-substituted xanthines examined.

Animals↗
PubMed1998

Induction of apoptosis in cardiac myocytes by an A3 adenosine receptor agonist.

The effects of the selective adenosine (ADO) A3 receptor agonist IB-MECA (N6-(3-iodobenzyl)adenosine-5'-N-methylcarboxamide) on cultured newborn rat cardiomyocytes were examined in comparison with ADO, the ADO A1 receptor-selective agonist R-PIA (N6-R-phenylisopropyladenosine), or the ADO A3 selective antagonist MRS 1191 (3-ethyl-5-benzyl-2-methyl-6-phenyl-4-phenylethynyl-1, 4-(+/-)-dihydropyridine-3,5 dicarboxylate), using digital image analysis of Feulgen-stained nuclei. At high concentration, IB-MECA (>/=10 microM ) and ADO (200 microM) induced apoptosis; however, R-PIA or MRS 1191 did not have any detectable effects on cardiac cells. In addition, DNA breaks in cardiomyocytes undergoing apoptosis following treatment by IB-MECA were identified in situ using the nick end labeling of DNA ("TUNEL"-like) assay. In the presence of >/=10 microM IB-MECA, disorder in the contraction waves appeared, and a decrease in the frequency of beats was observed. Analysis with light microscopy revealed that the number of contracting cells decreased in a concentration-dependent manner. The A3 receptor agonist IB-MECA caused an increase in intracellular free calcium concentration ([Ca2+]i). The drug produced a rapid rise followed by a sustained increase in [Ca2+]i, which lasted for 40-60 s. Finally, cessation of beating and Ca2+ transients were observed. Full recovery of contractile activity and rhythmical Ca2+ transients were observed 15-20 min after IB-MECA treatment. The induction of apoptosis in the cardiocytes by IB-MECA led to the appearance of features of apoptotic nuclei: the onset of condensation, compacting, and margination of nuclear chromatin. These effects were accompanied by the disintegration of the structural framework of the nucleus and nuclear breakdown. The results suggest that activation of the A3 adenosine receptor may participate in the process of apoptosis in cardiomyocytes.

Adenosine↗
PubMed1998

Structure-activity relationships and molecular modeling of 3, 5-diacyl-2,4-dialkylpyridine derivatives as selective A3 adenosine receptor antagonists.

The structure-activity relationships of 6-phenyl-1,4-dihydropyridine derivatives as selective antagonists at human A3 adenosine receptors have been explored (Jiang et al. J. Med. Chem. 1997, 39, 4667-4675). In the present study, related pyridine derivatives have been synthesized and tested for affinity at adenosine receptors in radioligand binding assays. Ki values in the nanomolar range were observed for certain 3,5-diacyl-2,4-dialkyl-6-phenylpyridine derivatives in displacement of [125I]AB-MECA (N6-(4-amino-3-iodobenzyl)-5'-N-methylcarbamoyladenosine) at recombinant human A3 adenosine receptors. Selectivity for A3 adenosine receptors was determined vs radioligand binding at rat brain A1 and A2A receptors. Structure-activity relationships at various positions of the pyridine ring (the 3- and 5-acyl substituents and the 2- and 4-alkyl substituents) were probed. A 4-phenylethynyl group did not enhance A3 selectivity of pyridine derivatives, as it did for the 4-substituted dihydropyridines. At the 2- and 4-positions ethyl was favored over methyl. Also, unlike the dihydropyridines, a thioester group at the 3-position was favored over an ester for affinity at A3 adenosine receptors, and a 5-position benzyl ester decreased affinity. Small cycloalkyl groups at the 6-position of 4-phenylethynyl-1,4-dihydropyridines were favorable for high affinity at human A3 adenosine receptors, while in the pyridine series a 6-cyclopentyl group decreased affinity. 5-Ethyl 2, 4-diethyl-3-(ethylsulfanylcarbonyl)-6-phenylpyridine-5-carboxylate , 38, was highly potent at human A3 receptors, with a Ki value of 20 nM. A 4-propyl derivative, 39b, was selective and highly potent at both human and rat A3 receptors, with Ki values of 18.9 and 113 nM, respectively. A 6-(3-chlorophenyl) derivative, 44, displayed a Ki value of 7.94 nM at human A3 receptors and selectivity of 5200-fold. Molecular modeling, based on the steric and electrostatic alignment (SEAL) method, defined common pharmacophore elements for pyridine and dihydropyridine structures, e.g., the two ester groups and the 6-phenyl group. Moreover, a relationship between affinity and hydrophobicity was found for the pyridines.

Adenosine↗