Anti-Tumor Activity of a Novel Immunosuppressant FTY720 in Multiple Myeloma.

Sphingosine and its metabolites are bioactive sphingolipids involved in lipid biosynthesis, signal transduction and apoptosis. FTY720, a synthetic sphingosine analogue of myriocine derived from culture filtrates of Isaria sinclairii, has been reported to interact with the sphingosine-1-phosphate specific G protein-linked receptors (S1P1, 3, 4 and 5) (Mandala S et al. Science, 2002) and alter the migration and homing of lymphocytes, thereby inhibiting the immune response (Matloubian M et al. Nature, 2004). Recent studies have also shown that FTY720 induces growth inhibition and/or apoptosis in human cancer cells in vitro as well as in vivo murine model (Azuma H et al. Cancer Research, 2002). To date, however, the biologic sequelae of inhibiting sphingosine-1-phosphate activity on multiple myeloma (MM) cells have not been demonstrated. In the present study, we examined whether FTY720 triggers anti MM activity. FTY720 induced potent cytotoxicity against MM cell lines including MM.1S, U266, RPMI8226, with IC₅₀ at 24 h of 3.0 – 7.0 mM, assessed by trypan-blue exclusion and MTT assays. FTY720 also inhibited growth of doxorubicin (Dox)-resistant RPMI8226-Dox40 and dexamethasone (Dex)-resistant MM.1R cell lines, with IC₅₀ values similar to the parental drug-sensitive cell lines. In contrast, no cytotoxicity of FTY720 was recognized against human peripheral blood mononuclear cells from normal healthy donors. The combination of Dex with FTY720 demonstrated enhanced cytotoxicity compared to either agent alone. Importantly, neither interleukin-6 (IL-6) nor insulin like growth factor-I (IGF-I), which induces MM cell growth and protection against Dex-induced apoptosis, protected against FTY720-induced growth inhibition. The anti-MM mechanisms of action of FTY720 were next studied, and FTY720 induced caspase-dependent apoptosis in MM cell lines: FTY720 triggers caspase−8, −9 and −3 cleavage, followed by PARP cleavage and DNA fragmentation, as confirmed by Western blotting and agarose gel electrophoresis, respectively. Moreover, FTY720 abrogated both IL-6 mediated phosphorylation of Akt-1, STAT3 and p42/44MAPK, and IGF-I mediated Akt-1 phosphorylation. Importantly, paracrine MM cell growth with bone marrow stromal cells was strongly inhibited by FTY720. These results suggest that FTY720 overcomes drug resistance in MM cells and, providing the rationale for its clinical evaluation to improve patient outcome in MM.