title: Methanocarba modification of uracil and adenine nucleotides: high potency of Northern ring conformation at P2Y1, P2Y2, P2Y4, and P2Y11 but not P2Y6 receptors.
abstract: The potency of nucleotide antagonists at P2Y1 receptors was enhanced by replacing the ribose moiety with a constrained carbocyclic ring (Nandanan, et al. J. Med. Chem. 2000, 43, 829-842). We have now synthesized ring-constrained methanocarba analogues (in which a fused cyclopropane moiety constrains the pseudosugar ring) of adenine and uracil nucleotides, the endogenous activators of P2Y receptors. Methanocarba-adenosine 5'-triphosphate (ATP) was fixed in either a Northern (N) or a Southern (S) conformation, as defined in the pseudorotational cycle. (N)-Methanocarba-uridine was prepared from the 1-amino-pseudosugar ring by treatment with beta-ethoxyacryloyl cyanate and cyclization to form the uracil ring. Phosphorylation was carried out at the 5'-hydroxyl group through a multistep process: Reaction with phosphoramidite followed by oxidation provided the 5'-monophosphates, which then were treated with 1,1'-carbonyldiimidazole for condensation with additional phosphate groups. The ability of the analogues to stimulate phospholipase C through activation of turkey P2Y1 or human P2Y1, P2Y2, P2Y4, P2Y6, and P2Y11 receptors stably expressed in astrocytoma cells was measured. At recombinant human P2Y1 and P2Y2 receptors, (N)-methanocarba-ATP was 138- and 41-fold, respectively, more potent than racemic (S)-methanocarba-ATP as an agonist. (N)-methanocarba-ATP activated P2Y11 receptors with a potency similar to ATP. (N)-Methanocarba-uridine 5'-triphosphate (UTP) was equipotent to UTP as an agonist at human P2Y2 receptors and also activated P2Y4 receptors with an EC(50) of 85 nM. (N)-Methanocarba-uridine 5'-diphosphate (UDP) was inactive at the hP2Y6 receptor. The vascular effects of (N)-methanocarba-UTP and (N)-methanocarba-UDP were studied in a model of the rat mesenteric artery. The triphosphate was more potent than UTP in inducing a dilatory P2Y4 response (pEC(50) = 6.1 +/- 0.2), while the diphosphate was inactive as either an agonist or antagonist in a P2Y6 receptor-mediated contractile response. Our results suggest that new nucleotide agonists may be designed on the basis of the (N) conformation that favors selectivity for P2Y1, P2Y2, P2Y4, and P2Y11 receptors.
authors.affiliation: Molecular Recognition Section, Laboratory of Bioorganic Chemistry, National Institute of Diabetes, Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892-0810, USA.
authors.forename: Hak Sung
authors.initials: HS
authors.lastname: Kim
authors.forename: R Gnana
authors.initials: RG
authors.lastname: Ravi
authors.forename: Victor E
authors.initials: VE
authors.lastname: Marquez
authors.forename: Savitri
authors.initials: S
authors.lastname: Maddileti
authors.forename: Anna-Karin
authors.initials: AK
authors.lastname: Wihlborg
authors.forename: David
authors.initials: D
authors.lastname: Erlinge
authors.forename: Malin
authors.initials: M
authors.lastname: Malmsjö
authors.forename: José L
authors.initials: JL
authors.lastname: Boyer
authors.forename: T Kendall
authors.initials: TK
authors.lastname: Harden
authors.forename: Kenneth A
authors.initials: KA
authors.lastname: Jacobson
mesh_terms: D000227:Adenine Nucleotides / Q000737:chemistry*; D000818:Animals; D001643:Bridged Bicyclo Compounds / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology; D004910:Erythrocyte Membrane / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007295:Inositol Phosphates / Q000378:metabolism; D008968:Molecular Conformation; D058910:Purinergic P2 Receptor Agonists; D058919:Purinergic P2 Receptor Antagonists; D051381:Rats; D018048:Receptors, Purinergic P2; D058472:Receptors, Purinergic P2Y1; D058473:Receptors, Purinergic P2Y2; D013237:Stereoisomerism; D013329:Structure-Activity Relationship; D014407:Tumor Cells, Cultured; D014421:Turkey; D014500:Uracil Nucleotides / Q000737:chemistry*
chemical_list: D000227:Adenine Nucleotides; D001643:Bridged Bicyclo Compounds; D007295:Inositol Phosphates; C546382:P2RY1 protein, human; C109734:P2RY11 protein, human; C546385:P2RY2 protein, human; C546384:P2ry1 protein, rat; C546387:P2ry2 protein, rat; D058910:Purinergic P2 Receptor Agonists; D058919:Purinergic P2 Receptor Antagonists; D018048:Receptors, Purinergic P2; D058472:Receptors, Purinergic P2Y1; D058473:Receptors, Purinergic P2Y2; D014500:Uracil Nucleotides; C104749:purinoceptor P2Y4; C099845:purinoceptor P2Y6