Extracellular sphingosine-1-phosphate contributes to survival properties of glioblastoma stem cells

Extracellular sphingosine-1-phosphate contributes to survival properties of glioblastoma stem cells Riccitelli E., Giussani P., Di Vito C., Condomitti G., Tringali C., Galli R., Viani P., Riboni L. Department of Medical Biotechnology and Translational Medicine, University of Milan, LITA Segrate, Via Fratelli Cervi 93, 20090 Segrate, Milan, Italy (elena.riccitelli@unimi.it) Sphingosine-1-phosphate (S1P) is a potent bioactive lipid formed from sphingosine by sphingosine kinases (SKs) action. S1P is considered as an onco-promoter molecule, favouring growth, invasion, and therapy-resistance of different tumours, including glioblastomas, the most frequent and aggressive human intracranial cancers. Despite the introduction of the alkylating agent temozolomide in glioblastoma therapy has improved patient survival, drug resistance limits its benefits. Accumulating literature indicates that glioblastoma stem-like cells (GSCs), a subpopulation of cells with the exclusive ability to self-renew and maintain the tumor, might contribute to glioblastoma aggressiveness and resistance to therapy. This study investigated S1P secretion by GSCs, and its possible role in cell survival. To this purpose we used GSCs isolated from the human U87 glioblastoma cell line (U-SC) and GSCs derived from a primary culture of human glioblastoma (L0627). We found that both GSC models efficiently form neurospheres in mitogen-defined medium, and express high levels of recognized neural-stem cell markers. Moreover, GSCs exhibited resistance to temozolomide, despite not expressing the DNA repair protein MGMT, a major contributor to temozolomide-resistance. Further analyses revealed the presence of S1P not only inside the cells, but also in the culture medium from both GSCs and U87. Notably the extracellular S1P level was found much higher in GSC models than in U87 cells, and the ratio between extracellular and intracellular S1P was 1:10 and 1:1 in U87 and GSCs, respectively. Enzyme activity assays excluded SKs presence in GSC medium, implicating an efficient secretion of S1P in GSCs. Intriguingly, concomitant treatment with temozolomide and a SKs inhibitor made GSCs sensitive to drug toxicity. Furthermore, S1P administration promoted cell survival after this co-treatment. Altogether our data implicate GSCs as an important source of extracellular S1P, which might act as an autocrine signal contributing to the survival properties of GSCs.