STEM-16. EXPLORING THE ROLE OF HYPOXIA AND MACROPHAGES ON GLIOBLASTOMA QUIESCENCE AND STEMNESS AS MEDIATORS OF RADIORESISTANCE

Abstract Our ultimate goal is to identify, characterize and therapeutically target the glioma cell population that is responsible for treatment resistance and recurrence following chemoradiation. Two cell types that may have overlapping phenotypes and are known to influence radioresistance are 1) quiescent cells and 2) glioblastoma (GBM) stem cells (GSCs). The extent of overlap between these populations is unknown as are the influence of hypoxia and immune cells on their radioresistant properties. We have shown that hypoxia and co-culturing with THP-1 macrophages dramatically increases the percentage of GSCs. Using the Fluorescent Ubiquitination-based Cell Cycle Indicator (FUCCI) system expressed in patient-derived gliomas, we used flow cytometry to distinguish cells based on their cell cycle status (G1/G0 and G2/M) and classified them as quiescent (high p27, low Ki67) or GSC (high CD133/CD44 co-expression). We found that the fraction of cells in G0 expressed high p27 and low Ki67, indicating cell cycle suppression and low proliferation, respectively. G0 glioma cells also contained a high percentage of GSCs, but GSCs were also distributed in G1 and G2/M. Following exposure to hypoxia, flow cytometry of FUCCI GBM cells showed an increase of cells in G0. After 3 Gy radiation under normoxic conditions, GBM cells demonstrated a shift to increased levels of G2/M cells compared to non-irradiated cells. In hypoxia, however, GBM cells sustained their population of cells in G0 and did not exhibit such cell cycle shifts. Our results indicate that both hypoxia and macrophages cause GSC enrichment and that hypoxia increases the proportion of quiescent cells in G0. Additional study is necessary to determine the extent of overlap between the GSCs and quiescent cells and to identify the intrinsic cell properties that drive tumor recurrence.