Structure−Activity Relationships of Fluorinated Lysophosphatidic Acid Analogues

Lysophosphatidic acid (LPA, 1- or 2-acyl-sn-glycerol 3-phosphate) displays an intriguing cell biology that is mediated via interactions with seven-transmembrane G-protein-coupled receptors (GPCRs) and the nuclear hormone receptor PPARγ. To identify receptor-selective LPA analogues, we describe a series of fluorinated LPA analogues in which either the sn-1 or sn-2 hydroxyl group was replaced by a fluoro or fluoromethyl substituent. We also describe stabilized phosphonate analogues in which the bridging oxygen of the monophosphate was replaced by an α-monofluoromethylene (−CHF−) or α-difluoromethylene (−CF<INF>2</INF>−) moiety. The sn-2- and sn-1-fluoro-LPA analogues were unable to undergo acyl migration, effectively “freezing” them in the sn-1-O-acyl or sn-2-O-acyl forms, respectively. We first tested these LPA analogues on insect Sf9 cells induced to express human LPA<INF>1</INF>, LPA<INF>2</INF>, and LPA<INF>3</INF> receptors. While none of the analogues were found to be more potent than 1-oleoyl-LPA at LPA<INF>1</INF> and LPA<INF>2</INF>, several LPA analogues were potent LPA<INF>3</INF>-selective agonists. In contrast, 1-oleoyl-LPA had similar activity at all three receptors. The α-fluoromethylene phosphonate analogue <BO>15</BO> activated calcium release in LPA<INF>3</INF>-transfected insect Sf9 cells at a concentration 100-fold lower than that of 1-oleoyl-LPA. This activation was enantioselective, with the (2S)-enantiomer showing 1000-fold more activity than the (2R)-enantiomer. Similar results were found for calcium release in HT-29 and OVCAR8 cells. Analogue <BO>15</BO> was also more effective than 1-oleoyl-LPA in activating MAPK and AKT in cells expressing high levels of LPA<INF>3</INF>. The α-fluoromethylene phosphonate moiety greatly increased the half-life of <BO>15</BO> in cell culture. Thus, α-fluoromethylene LPA analogues are unique new phosphatase-resistant ligands that provide enantiospecific and receptor-specific biological readouts.