Synthesis of pyridoxal phosphate derivatives with antagonist activity at the P2Y13 receptor |
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Authors: | Kim Yong-Chul Lee Jung-Sun Sak Katrin Marteau Frederic Mamedova Liaman Boeynaems Jean-Marie Jacobson Kenneth A |
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Institution: | Laboratory of Drug Discovery, Department of Life Science, Gwangju Institute of Science and Technology, 1 Oryong-dong, Buk-gu, Gwangju 500-712, Republic of Korea. |
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Abstract: | We have synthesized a series of derivatives of the known P2 receptor antagonist PPADS (pyridoxal-5'-phosphate-6-azo-phenyl-2,4-disulfonate) and examined their ability to inhibit functional activity of the recombinant human P2Y13 nucleotide receptor expressed in 1321N1 human astrocytoma cells co-expressing G(alpha)16 protein (AG32). Analogues of PPADS modified through substitution of the phenylazo ring, including halo and nitro substitution, and 5'-alkyl phosphonate analogues were synthesized and tested. A 6-benzyl-5'-methyl phosphonate analogue was prepared to examine the effect of stable replacement of the azo linkage. The highest antagonistic potency was observed for 6-(3-nitrophenylazo) derivatives of pyridoxal-5'-phosphate. The 2-chloro-5-nitro analogue (MRS 2211) and 4-chloro-3-nitro analogue (MRS 2603) inhibited ADP (100 nM)-induced inositol trisphosphate (IP3) formation with pIC50 values of 5.97 and 6.18, respectively, being 45- and 74-fold more potent than PPADS. The antagonism of MRS 2211 was competitive with a pA2 value of 6.3. MRS2211 and MRS2603 inhibited phospholipase C (PLC) responses to 30 nM 2-methylthio-ADP in human P2Y1 receptor-mediated 1321N1 astrocytoma cells with IC50 values of >10 and 0.245 microM, respectively. Both analogues were inactive (IC50>10 microM) as antagonists of human P2Y12 receptor-mediated PLC responses in 1321N1 astrocytoma cells. Thus, MRS2211 displayed >20-fold selectivity as antagonist of the P2Y13 receptor in comparison to P2Y1 and P2Y12 receptors, while MRS2603 antagonized both P2Y1 and P2Y13 receptors. |
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Keywords: | AG32 human astrocytoma 1321N1 cells co-expressing Gα16 protein AR-C67085MX 2-(propylthio)-β γ-dichloromethylene-ATP CHO Chinese Hamster ovary DMEM Dulbecco's Modified Eagle's Medium FBS fetal bovine serum HRMS high-resolution mass spectrometry IP3 inositol trisphosphate KRH Krebs-Ringer-HEPES MRS 2211 pyridoxal-5′-phosphate-6-azo-(2-chloro-5-nitrophenyl)-2 4-disulfonate MRS 2603 pyridoxal-5′-phosphate-6-azo-(4-chloro-3-nitrophenyl)-2 4-disulfonate PLC phospholipase C PPADS pyridoxal-5′-phosphate-6-azo-phenyl-2 4-disulfonate |
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