Next Generation Sequencing from control and CD97 knockdown of glioblastoma stem cells

Glioblastoma (GBM) stem cells (GSCs) are largely associated with a poor prognosis of GBM. Although the importance of identifying and validating molecular markers in GBM has been emphasized, single markers covering all GSCs are lacking. Through in vitro antibody screening, in this study, we demonstrated that CD97 identifies GSCs as a novel surface marker. Compared to conventional well-known GSC markers, such as CD133, CD44 and CD15, only CD97 was notably detected in all tested patient-derived GSCs. GBM patients expressing high levels of CD97 experienced worse clinical outcomes, and were predictive of poor overall survival in the public database. CD97 is required for GSC proliferation and self-renewal and was shown to reduce mouse survival with aggressive tumor progression in an orthotopic xenograft model. A comparative transcriptomic analysis revealed that CD97 triggers mTORC2 signaling through Akt (S473) phosphorylation, enhancing the expression of downstream genes, including ARHGAP1, BZW1, and BZW2. Moreover, inhibition of mTORC2 signaling with the pharmacological inhibitor JR-AP2-011 suppressed GSC proliferation, self-renewal, and downstream gene expression. Furthermore, sorting of CD97-enriched cells from patient-derived GSCs revealed that CD97 promotes GSC self-renewal and tumorigenicity via mTORC2/AKT signaling. Thus, our findings suggest that CD97 may be a general GSC enrichment marker in GBM and that the CD97-related pathway might serve as a therapeutic target for GBM.