N-cadherin upregulation mediates adaptive radioresistance in glioblastoma
Satoru Osuka, … , Christopher D. Willey, Erwin G. Van Meir
Satoru Osuka, … , Christopher D. Willey, Erwin G. Van Meir
Published March 15, 2021
Citation Information: J Clin Invest. 2021;131(6):e136098. https://doi.org/10.1172/JCI136098.
View: Text | PDF
Research Article Oncology

N-cadherin upregulation mediates adaptive radioresistance in glioblastoma

Abstract

Glioblastoma (GBM) is composed of heterogeneous tumor cell populations, including those with stem cell properties, termed glioma stem cells (GSCs). GSCs are innately less radiation sensitive than the tumor bulk and are believed to drive GBM formation and recurrence after repeated irradiation. However, it is unclear how GSCs adapt to escape the toxicity of repeated irradiation used in clinical practice. To identify important mediators of adaptive radioresistance in GBM, we generated radioresistant human and mouse GSCs by exposing them to repeat cycles of irradiation. Surviving subpopulations acquired strong radioresistance in vivo, which was accompanied by a reduction in cell proliferation and an increase in cell-cell adhesion and N-cadherin expression. Increasing N-cadherin expression rendered parental GSCs radioresistant, reduced their proliferation, and increased their stemness and intercellular adhesive properties. Conversely, radioresistant GSCs lost their acquired phenotypes upon CRISPR/Cas9-mediated knockout of N-cadherin. Mechanistically, elevated N-cadherin expression resulted in the accumulation of β-catenin at the cell surface, which suppressed Wnt/β-catenin proliferative signaling, reduced neural differentiation, and protected against apoptosis through Clusterin secretion. N-cadherin upregulation was induced by radiation-induced IGF1 secretion, and the radiation resistance phenotype could be reverted with picropodophyllin, a clinically applicable blood-brain-barrier permeable IGF1 receptor inhibitor, supporting clinical translation.

Authors

Satoru Osuka, Dan Zhu, Zhaobin Zhang, Chaoxi Li, Christian T. Stackhouse, Oltea Sampetrean, Jeffrey J. Olson, G. Yancey Gillespie, Hideyuki Saya, Christopher D. Willey, Erwin G. Van Meir

×

Figure 2

Fractionated irradiation increases N-cad expression in GSCs and N-cad drives the radioresistance phenotype.

Options: View larger image (or click on image) Download as PowerPoint
Fractionated irradiation increases N-cad expression in GSCs and N-cad dr...
(A) Western blot showing expression of cell-cell adhesion molecules following 6 to 12 cycles of 5 Gy irradiation in mGS cells. (B) Fluorescence microscopy shows that N-cad expression is increased on the cell surface of mGSRR cells (green). mGS and mGSRR cells are stably expressing mCherry (red). Nuclei were counterstained with Hoechst 33342 (blue). Scale bars: 25 μm. (C) Western blot showing expression of N-cad following 3 cycles of irradiation (1–2 Gy) in human GSCs, MGG4. (D) Western blot showing expression of N-cad, Tuj1, and Olig2 in human GSCs (MGG4) transfected with either control (Ctrl) or human N-cad expression vectors (OE). (E) Cell proliferation analysis for MGG4-Ctrl and MGG4 N-cad OE cells. (F) Clonogenic survival assay showing the surviving fraction of MGG4+/– N-cad cells after radiation doses of 1, 3, or 5 Gy. (G) Schematic showing experimental design to establish radioresistant PDX models. Subcutaneous tumors were exposed to repeated irradiation (2 Gy × 6 = 12 Gy total over 2 weeks). (H) Western blot showing expression of N-cad, Tuj1, and Olig2 in primary (P) or adapted to radiation therapy (RT) tumors of 2 PDX models. (I) Kaplan-Meier curve shows that increased N-cad mRNA expression is correlated with reduced survival in the TCGA-GBM data set. Log-rank test. High and low are defined as the top and bottom 15%. All blots show representative images (n = 3 or more). *P < 0.05, **P < 0.01, 2-tailed Student’s t tests unless otherwise indicated. The intensity of the immunoreactive bands was quantified in 3 independent experiments and the average is indicated below the blot.