P08.61 N cadherin upregulation mediates adaptive radioresistance in glioma stem cells

N cadherin upregulation mediates adaptive radioresistance in glioma stem cells Satoru Osuka,1 Oltea Sampetrean,2 Nobuyuki Onishi,2 Hideyuki Saya,2 Erwin G Van Meir,1, Laboratory of Molecular Neuro-Oncology, Departments of Neurosurgery and Hematology & Medical Oncology, School of Medicine and Winship Cancer Institute, Emory University, Atlanta, Georgia, USA. 2, Division of Gene Regulation, Institute for Advanced Medical Research, Keio University School of Medicine, Tokyo, Japan Glioblastoma (GBM) is composed of heterogeneous tumor cell populations including those with stem cell properties and slow growth, termed glioma stem cells (GSCs). GSCs display reduced sensitivity to chemo- and radiotherapy and play an important role in tumor recurrence, the main cause of patient demise. However, the molecular mechanisms underlying GSC’s therapeutic resistance are still unknown. To identify important mediators of radioresistance in GBM, we generated radioresistant GSCs by exposing mice GSCs derived from a mouse model of glioblastoma (Ink4A/Arf-/- neural stem cells with H-RasV12 overexpression) to repeated cycles of irradiation (5Gy x 12). The surviving subpopulation acquired strong radioresistance in vivo, accompanied by a reduction in cell proliferation and an increase in cell-cell adhesion. Because cell-cell adhesion is important for the maintenance of a quiescent stem-like state in normal stem cells, we analyzed the expression of related adhesion molecules, and found gradual increase in N-cadherin (N-cad) expression during repeated irradiation. To unravel the role of N-cad in radioresistance we modulated its expression in GSCs through overexpression vectors and CRISPR/Cas9-mediated knockout. Increased N-cad expression was associated with radio- and chemoresistance to temozolomide, reduced GSC proliferation and increased stemness properties, including suppression of neural differentiation. Conversely, radioresistant GSCs lost their acquired phenotypes (in vivo radioresistance, slow proliferation, cell-cell adhesion and stemness) upon CRISPR/Cas9-mediated knockout of N-cad. Mechanistically, we found that elevated N-cad expression led to the accumulation of β-catenin on the cell surface, resulting in the suppression of Wnt/ β-catenin signaling. To further validate our mice GSC findings in patients, we analyzed single cell RNA-seq data of human glioblastoma samples ({"type":"entrez-geo","attrs":{"text":"GSE57872","term_id":"57872"}}GSE57872) and found that cells with high N-cad expression had low expression of cell cycle genes and high expression of stemness-related