ki16425 and lysophosphatidic-acid

Lysophosphatidic acid (LPA) is a potent activator of human platelets in vitro. Recently, the G protein-coupled receptor LPA5/GPR92 has been identified to be the relevant LPA receptor responsible for the activation of human platelets by LPA. In a high-throughput screening campaign we identified a diphenyl pyrazole carboxylic acid as a small-molecule inhibitor for LPA5. Confirmation for the specificity of this small molecule was achieved in human platelets as the relevant cellular in vitro model. We could confirm using antagonists for alternative LPA receptors that we identified in our work the first non-lipid, small-molecule inhibitor for LPA5/GPR92 specifically inhibiting LPA-mediated platelet activation in vitro.

Topics: Binding Sites; Blood Platelets; Computer Simulation; Humans; Lysophospholipids; Platelet Activation; Protein Isoforms; Protein Structure, Tertiary; Receptors, Lysophosphatidic Acid

In CHO cells stably expressing the GPR87 fused with a G16alpha protein, lysophosphatidic acid (LPA) evoked an intracellular Ca(2+) increase in a high affinity manner. The Ca(2+) increase was reversibly blocked by the LPA receptor antagonists and inhibited by pretreatment of the cells with GPR87-specific siRNAs. GPR87 was shown to be closer to the P2Y and P2Y-related receptors than LPA receptors by ClustalW analyses. However, none of nucleotides and their derivatives activated GPR87. The human gpr87 is located on the chromosome 3q25 in a cluster containing p2y12,13,14. RT-PCR analysis showed that the mouse GPR87 was expressed in placenta, ovary, testis, prostate, brain, and skeletal muscle. The 3D model of GPR87-LPA complex indicated that the ligand interacted with R115 and K296 of GPR87, which are well conserved in the P2Y receptors. These results suggest that the GPR87 is a LPA receptor which evolved from a common ancestor of P2Y receptors.

Topics: Animals; Binding Sites; Calcium; CHO Cells; Cricetinae; Cricetulus; Electrophoresis, Polyacrylamide Gel; Female; Gene Expression Profiling; GTP-Binding Protein alpha Subunits; Humans; Intracellular Fluid; Isoxazoles; Lysophospholipids; Male; Mice; Mice, Inbred BALB C; Models, Molecular; Propionates; Protein Structure, Tertiary; Receptors, G-Protein-Coupled; Receptors, Lysophosphatidic Acid; Recombinant Fusion Proteins; Reverse Transcriptase Polymerase Chain Reaction; RNA Interference; Uridine Triphosphate

A series of isoxazole derivatives were synthesized and their antagonistic activities against LPA stimulation on both LPA(1)/CHO cells and rHSC cells were evaluated. Among them, 3-(4-[4-[1-(2-chloro-cyclopent-1-enyl)-ethoxycarbonylamino]-isoxazol-3- y]]-benzylsulfanyl)-propionic acid (34) showed the most potent activities.

Topics: Animals; Cell Proliferation; Chemistry, Pharmaceutical; CHO Cells; Cricetinae; Cricetulus; Drug Design; Fibrosis; Humans; Inhibitory Concentration 50; Isoxazoles; Liver; Lysophospholipids; Propionates; Rats