Pharmacologic Inhibition Of EGFR And MTOR Signaling Pathway In C6and Its Glioma Stem Cells

Backgroud:Glioma drug resistance are still problems to be solved in glioma treatment, there is a small amount of glioma stem cells in glioma. Cancer stem cells play a key role in tumor migration and aggressive, is the root of glioma recurrence and treatment resistance. cancer stem cells for the research object has important value to elucidate the mechanism of brain development. Epidermal growth factor receptor (EGFR) is a tyrosine kinase transmembrane receptor, Mammals rapamycin target protein (mTOR) is a kind of silk cytoplasm/threonine protein kinase, EGFR and mTOR signaling pathway has important role of cell proliferation and cell cycle. Applies both glioma and the role of glioma stem cells rarely has been reported.Objective:Using Erlotinib and Rapamycin alone and combination Observe proliferation and cycle in glioma C6cells and cancer stem cells (GSCs) at a cellular level and gene and protein expression in EGFR and mTOR signaling pathways.Discuss and testthe possible mechanisms ofproliferation, cycle and metastasis of Erlotinib and Rapamycin combined inhibition of glioma cell forproviding a new research direction in gliomas targeted therapy.Methods:C6cells were cultured by serum free medium, and the tumor stem cells (GSCs) were obtained. Cell immunofluorescence staining was used for detection CD133and nestin which expressed by tumor stem cells; CCK-8. RT-PCR and Western blot were used to observe the relationship of tumor cell proliferation, cell cycle and EGFR,mTOR signal pathway related genes, and its expressions of protein (EGFR, Akt, p-Akt, mTOR, p-mTOR, rps6k, p-rps6k after intervention of Erlotinib and Rapamycin.Results:The expressions of CD133and Nestin were detected in glioma stem cells; Erlotinib and Rapamycin had the dose-time dependence antiproliferative effect in C6and C6GSCs (P<0.05). Erlotinib and Rapamycin can increase the ratios of cells in G1phase and reduce the ratios of cells in S phase, the sensitivity to Erlotinib and Rapamycin in GSCswas lower than in C6cells(P<0.05); Protein expressions of EGFR and mTOR pathway in treatment group were lower than those in the control group(P<0.05), the efficacy is significantly in combined treatment.Conclusion:Tumor stem cells exist in C6glioma cell lines; Erlotinib and Rapamycin can inhibit the growth and proliferation of C6and C6GSCs by means of blocking the EGFR and mTOR signaling pathway.