Sphingosine Kinase 1 Is Involved in Proliferation and Survival of Chronic Myeloid Leukemia Cells.

Sphingosine Kinase 1 (SK1) is a key enzyme of the sphingolipid metabolism. By phosphorylating sphingosine (Sph), SK1 induces accumulation of sphingosine 1 phosphate (S1P). As a second messenger, S1P stimulates proliferation and survival and protects cells against ceramide-induced apoptosis. Although oncogenic activity of SK1 has been demonstrated in a variety of solid tumors, its role in leukemic cells growth remains unclear. Recently, SK1 overexpression has been reported to represent an oncogenic event during erythroleukemia progression (Lescolan E. et al, Blood 2005). We focused on Chronic Myeloid Leukemia (CML), a hematopoietic disorder characterized by the presence of the Philadelphia (Ph) chromosome that leads to the expression of the chimeric protein Bcr/Abl. Through its constitutive tyrosine kinase activity, Bcr/Abl is recognized as the pathogenetic event that causes CML and thus the basis for the development of specific inhibitors. Among these, Imatinib Mesylate (IM) represents the more successful drug for CML treatment. However, most patients who are treated in advanced stages of the disease relapse after an initial response to IM. Therefore, the existence of a functional link between Bcr/Abl and SK1 would provide new strategies to overcome acquired resistance to IM in CML. To test the hypothesis that SK1 contributes to the growth and survival of leukemic cells, we first verified that the kinase transcript is expressed in a panel of CML cell lines [AR230 (p230^Bcr/Abl+), K562 and RWLeu4 (both p210^Bcr/Abl+)], although at different levels. Then the effect of pharmacological inhibition of SK1 was assessed by using a competitive inhibitor (SKI, SK Inhibitor, Calbiochem) of the ATP-binding site of SK1. Cells were treated with increasing doses (1 to 50 μM) of SKI for up to 72 hrs and the WST-1 assay was performed. A dose- and time-dependent anti-proliferative and cytotoxic effect was observed (IC₅₀: 7.02 μM for AR230, 7.06 μM for K562 and 35.8 μM for RWLeu4 cells). Surprisingly, up-regulation of SK1 transcript was detected. Finally, when the Eosinophilic Leukemia cell line Eol-1 (FIP1L1-PDGFRα⁺) was exposed to SKI under the same experimental conditions of CML cells, an even more potent anti-proliferative and cytotoxic effect was observed, indicating a higher sensitivity of these cells to SKI (IC₅₀: 0.43 μM). Altogether our results suggest involvement of SK1 in the control of growth and survival of myeloid leukemia cells of different origin. To address the question whether a functional link between SK1 and Bcr/Abl occurs, we are now investigating if Bcr/Abl expression and phosphorylation status are modulated following treatment with SKI and conversely if SK1 expression and activity are affected by treatment with IM.