Effect Of Cell Growth Inhibition Through Targeting MTOR-mediated Signaling Pathway In Nasopharyngeal Carcinoma

Background: Nasopharyngeal carcinoma (NPC) is one of the most common malignant tumors in southern China. Lymphaden is abound innasopharynx and anatomic site is deep, so surgical operation is hardly applied on NPC. Radiotherapy is the main means to cure nasopharyngeal carcinoma, but five year survival rate of patients were not improved obviously although radiotherapy technique advanced continuously. Accordingly, exploring new therapeutics to enhance the effect of radiation is very important. The mammalian target rapamycin (mTOR) is a potential target for anti-cancer therapy as it is a key molecule in the PTEN/PI3K/AKT/mTOR signaling pathway and plays a critical role in controlling cell proliferation and survival. Accumulating studies indicated mTOR signaling pathway was always activated in human cancers. Thus, potential applications of mTOR inhibitor rapamycin in treatment of various cancers have been actively investigated in both preclinical and clinical stages. However, the application of rapamycin in treatment of NPC has not been investigated so far.Objective: My study was designed to investigate the growth inhibition effect of rapamycin in human NPC cell lines CNE-1 and CNE-2, and the significance of p70S6K and p4EBPl in this action.Methods: Human NPC cell lines CNE-1 and CNE-2 were treated with different concentrations (0,5,10,20,40,80,160nmol/L) of rapamycin. In the situations of pre- and post-treatment of rapamycin, variation of cell morphology was observed by microscope ; Cell proliferation of the CNE-1 and CNE-2 cell lines in response to rapamycin was assessed by CCK-8 assay; Cell cycle and apoptosis analyses were assayed by flow cytometry; mRNA levels of p70S6K and p4EBPl were detected using RT-PCR pre- and post-treatment of rapamycin.Results:(1) The cell growth was inhibited by rapamycin , and such effect showed a concentration-dependent manner in a certain range (P< 0.05) . 48h after treatment of rapamycin , the CNE-1 cell density was depressed and morphology of cell became smaller and rounder , the pedes spurii became shorter and less, but the morphological alteration in CNE-2 cell was not obvious. (2)Treatment of rapamycin was shown to increase in the proportion of cells in the G1 phase and decrease in the proportion of cells in the S phase in CNE-1 and CNE-2 cell lines compared to untreated cells, such effect showed a concentration-dependent manner in a certain range also (P<0.05) . But treatment of rapamycin didn't induce apoptosis significantly (p > 0.05). (3)Rapamycin significantly inhibited mRNA expression of mTOR and decreased the mRNA levels of phosphorylated p70S6K and 4EBP1 in CNE-2 cell lines (P<0.01) , such effect presented as a concentration-dependent manner in a certain range too. Conclusions :(1) Rapamycin can inhibit growth and proliferation of the CNE-1 and CNE-2 cell lines. (2) Rapamycin inhibit cell growth and proliferation mainly through the mechanism of effectively arrests CNE-1and CNE-2 cells in the G1 phase of the cell cycle, but does not induce apoptosis. (3) Rapamycin can down-regulate mTOR activity and inhibit the phosphorylation of p70S6K and 4EBP1, and then suppress 5'TOP mRNA translation initiation and protein synthesis, which finally inhibit growth of the CNE-2 cell.(4) Therefore, our finding provides evidences that rapamycin, a specific inhibitor of mTOR can be thought of as a potential target for NPC molecule targeted therapy.