title: Structure-activity relationships of pyridoxal phosphate derivatives as potent and selective antagonists of P2X1 receptors.
abstract: Novel analogues of the P2 receptor antagonist pyridoxal-5'-phosphate 6-azophenyl-2',5'-disulfonate (2) were synthesized and studied as antagonists in functional assays at recombinant rat P2X1, P2X2, and P2X3 receptors expressed in Xenopus oocytes (ion flux stimulation) and at turkey erythrocyte P2Y1 receptors (phospholipase C activation). Selected compounds were also evaluated as antagonists of ion flux and the opening of a large pore at the recombinant human P2X7 receptor. Modifications were made in the 4-aldehyde and 5'-phosphate groups of the pyridoxal moiety: i.e. a CH2OH group at the 4-position in pyridoxine was either condensed as a cyclic phosphate or phosphorylated separately to form a bisphosphate, which reduced potency at P2 receptors. 5-Methylphosphonate substitution, anticipated to increase stability to hydrolysis, preserved P2 receptor potency. At the 6-position, halo, carboxylate, sulfonate, and phosphonate variations made on the phenylazo ring modulated potency at P2 receptors. The p-carboxyphenylazo analogue, 4, of phosphate 2 displayed an IC50 value of 9 nM at recombinant P2X1 receptors and was 1300-, 16-, and > 10,000-fold selective for P2X1 versus P2X2, P2X3, and P2Y1 subtypes, respectively. The corresponding 5-methylphosphonate was equipotent at P2X1 receptors. The 5-methylphosphonate analogue containing a 6-[3,5-bis(methylphosphonate)]phenylazo moiety, 9, had IC50 values of 11 and 25 nM at recombinant P2X1 and P2X3 receptors, respectively. The analogue containing a phenylazo 4-phosphonate group, 11, was also very potent at both P2X1 and P2X3 receptors. However, the corresponding 2,5-disulfonate analogue, 10, was 28-fold selective for P2X1 versus P2X3 receptors. None of the analogues were more potent at P2X7 and P2Y1 receptors than 2, which acted in the micromolar range at these two subtypes.
authors.affiliation: Laboratory of Bioorganic Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, NIH, Bethesda, Maryland 20892-0810, USA.
authors.forename: Y C
authors.initials: YC
authors.lastname: Kim
authors.forename: S G
authors.initials: SG
authors.lastname: Brown
authors.forename: T K
authors.initials: TK
authors.lastname: Harden
authors.forename: J L
authors.initials: JL
authors.lastname: Boyer
authors.forename: G
authors.initials: G
authors.lastname: Dubyak
authors.forename: B F
authors.initials: BF
authors.lastname: King
authors.forename: G
authors.initials: G
authors.lastname: Burnstock
authors.forename: K A
authors.initials: KA
authors.lastname: Jacobson
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D066298:In Vitro Techniques; D009865:Oocytes / Q000378:metabolism; D018408:Patch-Clamp Techniques; D058919:Purinergic P2 Receptor Antagonists; D011732:Pyridoxal Phosphate / Q000031:analogs & derivatives* / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology; D051381:Rats; D018048:Receptors, Purinergic P2 / Q000502:physiology; D058469:Receptors, Purinergic P2X; D058476:Receptors, Purinergic P2X2; D058477:Receptors, Purinergic P2X3; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors; D013329:Structure-Activity Relationship; D014981:Xenopus
chemical_list: C546401:P2RX2 protein, human; C546407:P2RX3 protein, human; C546403:P2rx2 protein, rat; C546410:P2rx3 protein, rat; D058919:Purinergic P2 Receptor Antagonists; D018048:Receptors, Purinergic P2; D058469:Receptors, Purinergic P2X; D058476:Receptors, Purinergic P2X2; D058477:Receptors, Purinergic P2X3; D011994:Recombinant Proteins; D011732:Pyridoxal Phosphate; D000255:Adenosine Triphosphate