Receptor agonist, antagonist and enzyme inhibitors for studying lysolipid signaling

Lysophosphatidic acid (LPA, 1- or 2-radyl-sn-glycerol 3-phosphate) is an important phospholipid, which elicits growth-factor-like effects in a number of cell types through cell surface G protein-coupled receptors (GPCRs). Extracellularly, the sole source of LPA is from the hydrolysis of lysophosphatidylcholine (LPC) by a secreted enzyme Autotaxin (ATX) through its lysophospholipase D activity. Due to exhibition of common features of the phosphodiesterase family, ATX was included as the second member of the nucleotide pyrophosphatase/phosphodiesterase (NPP) family. The emergence of ATX-LPA receptor axis has encouraged researchers to develop novel therapies for the prevention and treatment of cancer and other diseases. In order to study the precise physiological roles of each LPA receptors and ATX, I synthesized a set of LPA and LPC analogues as either ATX inhibitors or LPA receptor agonists/antagonists. Specific modifications in this thesis include optimization of the polar phosphonate head of LPA to phosphorothioate, restriction of glycerol backbone of LPA to phenyl ring and change of phosphonate head to vinyl sulfone in LPC molecule. All these analogues were tested in either in vitro or in vivo assays through external collaborations. One interesting result includes the identification of sn-2 radyl phosphorothioate analogues of LPA (sn-2 OMPT) as the most potent LPA5 receptor agonists. Furthermore, aromatic phosphonate analogues of LPA displayed very potent ATX inhibition. An in vivo study revealed that one of aromatic phosphonate analogues significantly reduced pulmonary fibrosis induced by bleomycin. In addition, fluorophosphonate and vinyl sulfone analogues of LPC were designed as activity-based probe (ABP) for selected enzymes of the NPP family. The study indicated that fluorophosphonate analogues cannot serve as ABP due to its high reactivity. In contrast, vinyl sulfone ABP analogues of LPC showed good inhibition of ATX. Meanwhile, the kinetics and dialysis suggested that vinyl sulfone analogues were irreversible inhibitors of ATX. We are still working on finding further evidence for irreversible inhibition.