Differential Expression of P-Element–Induced Wimpy Testis (PIWI)–Interacting RNAs in Glioblastoma Stem Cells Affects Their Biological Features: Implications for Tumor Progression and Patient Survival

Glioblastoma isocitrate dehydrogenase-wild-type (GBM) is the most lethal and, concurrently, the most common malignant primary brain tumor. Its aggressive behavior is associated with the presence of a subpopulation of glioblastoma stem cells (GSCs) that drive resistance to therapy and early relapse. P-element-induced wimpy testis-interacting RNAs (piRNAs), which are characteristically expressed in undifferentiated cells, are thought to play an important role in GSC biology. To identify piRNAs associated with GSCs, we selected 24 paired GSC and non-GSC cultures from patients with GBM based on the expression of stem cell markers CD133 and Sox2, ability of GSCs to form spheres in culture, and to differentiate and replicate tumors in immunodeficient mice. Global piRNA profiling using next-generation sequencing revealed 98 significantly differentially expressed piRNAs in GSCs compared with non-GSCs, including piR-9491, whose dysregulation in GBM has been previously described. Downregulation of piR-9491 in GSCs led to decreased viability, growth, invasion, and increased apoptosis. Significance of piRNA pathway for GBM pathology was further highlighted by identification of 34 piRNAs connected to patients' survival. These molecules were further used for the establishment of a piRNA signature predicting survival of patients with GBM. Our results not only confirm the important role of piR-9491 in GSCs but also indicate a potential significant role of piRNAs in the biology of GBM.