Effects of Small-Molecule Inhibitors on Motility and Proliferation of Differentiated and Undifferentiated Glioblastoma Stem Cells

• Emily Borell, Biological Sciences, University of Delaware

• Deni Galileo, Biological Sciences, University of Delaware

Glioblastoma (GBM) is an aggressive and incurable form of brain cancer. Glioblastoma treatment is difficult due to the minor population of glioblastoma stem cells (GSCs), which invade the surrounding brain tissue to escape surgical resection and resist radiation and chemotherapy. Our lab previously found evidence that the L1CAM cell adhesion protein stimulates the motility and proliferation of GBM cells through the integrin, focal adhesion kinase (FAK), and fibroblast growth factor receptor (FGFR) signaling pathways. Decreased motility and proliferation of GBM cells from established (non-stem) glioblastoma cell lines and GSC lines was observed in previous experiments when using small molecule inhibitors FGFR inhibitor PD173074, FAK inhibitors PF431396 and Y15, and αvβ3/αvβ5 integrin inhibitor cilengitide. These results were expanded to test the hypothesis that motility and proliferation of patient-derived GSCs and “differentiated” GSCs similarly would be inhibited by the small molecule inhibitors. GSCs were cultured in high serum media for several weeks to induce differentiation. Some of these differentiated GSCs were later cultured in stem cell media to determine if the cells were able to return to a stem-like phenotype.  Our “Super Scratch” time-lapse microscopy assay was performed to measure motility after exposing the cells to the inhibitors. Cell proliferation was assessed by staining cells for DNA content and flow cytometry analysis to determine the percentage of cells in the S phase of the cell cycle. Both GSCs and differentiated GSCs exhibited similar decreases in motility when treated with inhibitors. However, the differentiated GSCs exhibited slower overall motility, but differentiated GSCs put back into stem-cell media exhibited similar mobility as regular GSCs. Analysis of the proliferation assay is ongoing. These results suggest that the small molecule inhibitors tested may be similarly effective against both GSCs and differentiated GBM cells within a tumor and provide evidence that GSCs are able to enter in and out of a “differentiated” state depending on the extracellular environment.