Sphingosine kinase type 1 inhibition reveals rapid turnover of circulating sphingosine 1-phosphate

S1P (sphingosine 1-phosphate) is a signalling molecule involved in a host of cellular and physiological functions, most notably cell survival and migration. SIP, which signals via a set of five G-protein-coupled receptors (S1P1-S1P5), is formed by the action of two SphKs (sphingosine kinases) from Sph (sphingosine). Interfering RNA strategies and SphK1 (sphingosine kinase type 1)-null (Sphkl(-1-)) mouse studies implicate SphKl in multiple signalling cascades, yet there is a paucity of potent and selective SphK1 inhibitors necessary to evaluate the effects of rapid onset inhibition of this enzyme. We have identified a set of submicromolar amidine-based SphKl inhibitors and report using a pair of these compounds to probe the cellular and physiological functions of SphK1. In so doing, we demonstrate that our inhibitors effectively lower SIP levels in cell-based assays, but we have been unable to correlate SphK1 inhibition with changes in cell survival. However, SphK1 inhibition did diminish EGF (epidermal growth factor)-driven increases in SIP levels and Akt (also known as protein kinase B)/ERK (extracellular-signal-regulated kinase) phosphorylation. Finally, administration of the SphK1 inhibitor to wild-type, but not Sphkl(-/-), mice resulted in a rapid decrease in blood SIP levels indicating that circulating SIP is rapidly turned over.