Sphingosine Kinase Inhibitors Effectively Inhibit Growth of Mast Cells with D816V-KIT Mutations

Abstract Sphingosine-1-phosphate (S1P) is a lipid mediator that promotes cell proliferation and prevents apoptosis in a variety of cells and its increased synthesis by Sphingosine Kinase (SphK) has been associated with oncogenesis and cancer prognosis. Mastocytosis is a disorder characterized by excessive mast cell (MC) proliferation and accumulation in tissues. Treatment therapies for aggressive mastocytosis have to date been partially effective, and thus the search of new pharmacological targets is warranted. We thus investigated the involvement of SphK isoforms (SphK1 and SphK2) and generation of S1P in the regulation of abnormal MC growth. The specific SphK1 and SphK2 inhibitors, SKI-178 and ABC294640, respectively, were used in this study. The effect of SphK1 and SphK2 inhibition on human (LAD2, HMC1.1 and HMC1.2) and murine (P815) neoplastic MC proliferation and survival was evaluated. SphK2 inhibition retarded entry into the cell cycle in all MC lines with no significant effect on cell survival. However, SphK1 inhibition led to cell cycle arrest in G2/M and apoptosis predominantly in MCs carrying D816V-KIT mutation. The effect of SKI-178 was associated with an alteration in the damage response cascade. This included activation of checkpoint kinase 2 (chk2) combined with a depletion of chk1, leading respectively to p53- and caspase2-induced apoptosis and cell cycle arrest mediated by cdc25c depletion. Both SphK inhibitors reduced MC accumulation in an in vivo mouse model of aggressive mastocytosis and the number of cultured bone marrow MCs and MC progenitors from a patient with systemic mastocytosis. These results support further consideration for SphK inhibitors as a cytoreductive strategy in proliferative MC disorders.