Alkoxymethylenephosphonate analogues of (Lyso)phosphatidic acid stimulate signaling networks coupled to the LPA₂ receptor

An efficient stereocontrolled synthesis afforded alkoxymethylene- phosphonate (MP) analogues of lysophosphatidic acid (LPA) and phosphatidic acid (PA). The pharmacological properties of MP-LPA and MP-PA analogues were characterized for LPA receptor subtype-specific agonist and antagonist activity using Ca²⁺-mobilization assays in RH7777 cells expressing the individual LPA₁-LPA₃ receptors and CHO cells expressing LPA₄. In addition, activation of a PPARγ reporter gene construct expressed in CV-1 cells was assessed. These metabolically stabilized LPA analogues exhibited an unexpected pattern of partial agonist/antagonist activity for the LPA G-protein-coupled receptor family and the intracellular LPA receptor PPARγ. Analogues were compared with 18:1 LPA for activation of downstream signaling in HT-29colon cancer cells, which exclusively express LPA₂, and both SKOV3 and OVCAR3 ovarian cancer cells, which express LPA₁, LPA₂, and LPA₃. Unexpectedly, reverse phase protein arrays showed that four MP-LPA and MP-PA analogues selectively activated downstream signaling in HT-29 cells with greater potency than LPA. In particular, the oleoyl MP-LPA analogue strongly promoted phosphorylation and activation of AKT, MEK, and pS6 in HT-29cells in a concentration-dependent manner. In contrast, the four MP-LPA and MP-PA analogues were equipotent with LPA for pathway activation in the SKOV3 and OVCAR3 cells. Taken together, these results suggest that the MP analogues may selectively activate signaling via the LPA₂ receptor subtype, while simultaneously suppressing signaling through the LPA₁ and LPA₃ subtypes.