Anales RANF

P.02 P2X7 RECEPTOR ACTIVATION INCREASESMESENCHYMAL TRANSDIFFERENTIATION AND INVASIVENESS OF GLIOBLASTOMA-DERIVED STEM-LIKE CELLS S. Ziberi 1,2 , M. Carluccio 1,2 , M. Zuccarini 1 , L. Ricci-Vitiani 3 , R. Pallini 4 , P. Giuliani 1 , F. Caciagli 1 , P. Di Iorio 1 , R. Ciccarelli 1,2 . 1 University of Chieti-Pescara and 2 StemTeCh Group.Chieti, Italy. 3 Istituto Superiore di Sanità and 4 Università Cattolica del Sacro Cuore. Rome. Italy . Stem-like cells (GSCs) with high self-renewal, resistance to radio/chemotherapy and metastatic potential contributes to glioblastoma(GBM)unfavorable prognosis. We previously showed high expression levels of P2X7 receptors (P2X7R) in GSCs isolated from human primary GBM. Here, using the same GSCs, we investigated the effects of P2X7R modulation on genes associated to epithelial-to-mesenchymal transition (EMT), a process likely contributing to GBM malignancy. The P2X7R agonist2'(3')-O-(4- benzoylbenzoyl)-ATP (BzATP, 50- 200μM)or the known EMT inducer,Transforming Growth Factor- β1 (TGFβ1, 5 -10ng/ml)up-regulated mRNAs and increased the protein content of selected EMT markers during 24-72h following drug administration to GSCs. Both agents also enhanced GSC migration and invasiveness, evaluated by scratch and trans-well migration assays. BzATP effects were mediated by increased phosphorylation of SMAD2, a downstream TGFβ signalingeffector. A8301, an inhibitor of this pathway,abrogated them, suggesting TGFβ involvement in P2X7R -mediated effects in GSCs. BzATPalso decreased lactate dehydrogenase and caspase-3/7 activity in GSC medium. The P2X7R antagonist A438079mostly counteracted BzATP effects. Finally, i) GSCs expressed two main human P2X7R splicing variants, the full length P2X7AR and the truncated P2X7BR, lacking the carboxylic tail, which have different functional properties; ii)BzATP up-regulated their expression, thus possibly favoring A/B subunit assembly into a heterotrimeric P2X7R with greater sensitivity towards agonists/support to cell energy. These results are in line with the increased expression of EMT markers, cell migration/invasion and GSC survival induced by P2X7R stimulation. Since in GBM microenvironment extracellular ATP levels may activate P2X7R, our data also suggest a role in GBM recurrence/invasiveness.