A Preliminary Study Of Nasopharyngeal Carcinoma EBV-LMP1 By AKT/mTOR Signaling Pathway In The Regulation Of WWOX

ObjectiveOur previous studies found the expression of tumor suppressor gene WWOX was reduced in nasopharyngeal carcinoma, and WWOX expression gradually declined with the progress and lymph node metastasis in patients. These suggested that WWOX was related with the development of nasopharyngeal carcinoma. Nasopharyngeal is closely related to infection of EB virus (EBV). AKT/mTOR signaling pathway is considered the primary pathway of cancer cell survival. The study mainly used oncogene LMP1 of EBV as a starting point to explore whether LMP1 regulated WWOX expression through AKT/mTOR signaling pathway.Methods1. Correlation between LMP1 and AKT/mTOR signaling pathway, WWOX gene expressionWestern blot and qPCR were used to detect the expression of AKT/mTOR pathway (Akt, p-Akt, p70S6K and p-p70S6K) and WWOX in nasopharyngeal carcinoma cell lines CNE1 and CNE1-LMP1, and accessed relationship of LMP1 with AKT/mTOR and WWOX.2. Studies of preliminary validation that LMP1 regulated WWOX expression by AKT/mTOR pathwayWith targeted inhibitors (rapamycin) of AKT/mTOR signaling pathway to inhibit CNE1-LMP1 cell line, Western blot and qPCR were used to detect the expression of AKT/mTOR pathway (Akt, p-Akt, p70S6K and p-p70S6K) and WWOX in the CNE1-LMP1 cell line, and verified that LMP1 regulated WWOX expression via AKT/mTOR pathway.Results1. Correlation between LMP1 and AKT/mTOR signaling pathway, WWOX gene expressionResearch of correlation between LMP1 and WWOX gene, protein expression:in CNE1-LMP1 cells, WWOX gene and protein levels were decreased compared with CNE1 cells (P=0.025,P=0.042, respectively), which was significantly different from each other, and suggested that LMP1 expression correlated with WWOX.Research of correlation between LMP1 and AKT/mTOR signaling pathway: when nasopharyngeal carcinoma cell line CNE1-LMP1 was compared with CNE1 in AKT/mTOR pathway key protein of Akt, p-Akt, p70S6K and p-p70S6K expression, P values were 0.075,0.008,0.124,0.034, respectively, and expression of p-Akt, p-p70S6K in CNE1-LMP1 were higher than CNE1, which were significantly different from each other. It suggested AKT/mTOR pathway was regulated by LMP1.2. Studies of preliminary validation that LMP1 regulated WWOX expression by AKT/mTOR pathwayAfter using targeted pathway inhibitor (rapamycin) inhibits CNE1-LMP1 cell lines, expression levels of WWOX gene and protein were elevated, P values were 0.036,0.044, respectively. It suggested that AKT/mTOR signaling pathway regulated expression of WWOX gene. In addition, after using targeted pathway inhibitor (rapamycin) inhibits, all P values of AKT/mTOR signaling pathway key proteins (Akt, p-Akt, p70S6K p-p70S6K) were 0.136,0.354,0.103,0.045, respectively. Protein expression of Akt, p-Akt, p70S6K was no statistically significant difference respectively, while expression level of p-p70S6K in CNE1-LMP1 cells was lower after drug inhibition. The difference was statistically significant. It indicated that the drug inhibited protein expression of AKT/mTOR pathway.Conclusions1. WWOX gene expression in CNE1-LMP1 cell lines was lower than CNE1 cell lines, which might be regulated by the EBV-LMP1 oncogene.2. LMP1 could regulate AKT/mTOR signaling pathway and the relevant expression of downstream genes.3. Reduced expression of WWOX might related with the regulation by LMP1 via AKT/mTOR signaling pathway.