ABSTRACT
Lysophosphatidic acid (LPA) is a bioactive compound that has gained attention due to its role in neoplastic diseases. Our group has developed a potent dual LPA1/LPA3 receptor antagonist, VPC51098 (LPA1 IC(50) = 84 nM, LPA1 IC(50) = 48 nM) that contained a labile phosphate head group. This lability has impaired our evaluation of our scaffold of LPA receptor antagonists in vivo. We wished to replace the phosphate with a potentially more stable head group while retaining potency at both LPA1 and LPA3 to facilitate future in vivo studies. We tested in vitro potency of all head groups including α-methylene, α-fluoromethylene, α-hydroxymethylene; vinyl phosphonates; α-fluoro vinyl phosphonates. The most potent compound was found to be a low micromolar inhibitor VPC51299 that contained a vinyl phosphonate and possessed a half-life of approximately 90 min in rats when dosed intravenously. Herein, we describe the synthesis and initial biological evaluation of these compounds.
ABSTRACT
Phosphatidic acid and lysophosphatidic acid are minor but important anionic bioactive lipids involved in a number of key cellular processes, yet these molecules have a simple phosphate headgroup. To find out what is so special about these lipids, we determined the ionization behavior of phosphatidic acid (PA) and lysophosphatidic acid (LPA) in extended (flat) mixed lipid bilayers using magic angle spinning 31P NMR. Our data show two surprising results. First, despite identical phosphomonoester headgroups, LPA carries more negative charge than PA when present in a phosphatidylcholine bilayer. Dehydroxy-LPA [1-oleoyl-3-(phosphoryl)propanediol] behaves in a manner identical to that of PA, indicating that the difference in negative charge between LPA and PA is caused by the hydroxyl on the glycerol backbone of LPA and its interaction with the phosphomonoester headgroup. Second, deprotonation of phosphatidic acid and lysophosphatidic acid was found to be strongly stimulated by the inclusion of phosphatidylethanolamine in the bilayer, indicating that lipid headgroup charge depends on local lipid composition and will vary between the different subcellular locations of (L)PA. Our findings can be understood in terms of a hydrogen bond formed within the phosphomonoester headgroup of (L)PA and its destabilization by competing intra- or intermolecular hydrogen bonds. We propose that this hydrogen bonding property of (L)PA is involved in the various cellular functions of these lipids.
Subject(s)
Lipid Bilayers/chemistry , Lysophospholipids/chemistry , Membranes, Artificial , Models, Molecular , Phosphatidic Acids/chemistry , Cell Membrane/chemistry , Endoplasmic Reticulum/chemistry , Hydrogen Bonding , Hydrogen-Ion Concentration , Intracellular Membranes/chemistry , Ions/chemistry , Least-Squares Analysis , Magnetic Resonance Spectroscopy , Molecular Structure , Phosphatidylcholines/chemistry , Phosphatidylethanolamines/chemistry , Protons , TitrimetryABSTRACT
A recently reported dual LPA(1)/LPA(3) receptor antagonist (1) has been modified so as to modulate the basicity, sterics, and dipole moment of the 2-pyridyl moiety. Additionally, the implications of installing nonhydrolyzable phosphate head group isosteres with regard to antagonist potency and selectivity at LPA receptors is described. This study has resulted in the development of the first nonhydrolyzable and presumably phosphatase-resistant LPA(3)-selective antagonist reported to date.
Subject(s)
Pyridines/pharmacology , Receptors, Lysophosphatidic Acid/antagonists & inhibitors , Animals , Fibroblasts/drug effects , Fibroblasts/physiology , Guanosine 5'-O-(3-Thiotriphosphate)/metabolism , Molecular Conformation , Molecular Structure , Pyridines/chemical synthesis , Pyridines/chemistry , Structure-Activity RelationshipABSTRACT
Using an N-oleoyl ethanolamide scaffold, a series of phosphate polar head group analogues of LPA comprised of various alpha-substituted phosphonates and thiophosphates was prepared. In a broken cell GTP[gamma35S] binding assay, agonist activity was evaluated at the three LPA receptors of the endothelial differentiation gene (Edg) family. This study has resulted in the discovery of a nonhydrolyzable LPA1-selective agonist (11). Additionally, thiophosphate 19 bears an isosteric phosphate mimetic that confers agonism at the LPA1 receptor but not LPA2.
