Synergistic Effect Of Notch-3-specific Inhibition And Paclitaxel In Non-small Cell Lung Cancer(NSCLC) Cells Via Activation Of The Intrinsic Apoptosis Pathway

Objective: Lung cancer is one of the high incidence of human cancer,including non-small cell lung cancer(NSCLC)is its main type.Although there were surgery,a variety of chemotherapy drugs and targeted therapy for its treatment,but its prognosis was still poor.Notch3 protein expression in NSCLC was significantly higher than that in normal lung tissue,and was closely related to the prognosis of NSCLC.Patients with lung cancer have a certain resistance to conventional chemical therapy due to individual differences,such as paclitaxel and gemcitabine.Moreover,Notch signaling is critical in the chemical resistance of lung cancer cells.Therefore,this study explored whether Notch3 protein is associated with paclitaxel sensitivity in NSCLC and explored whether Notch3 inhibitors enhance the chemosensitivity of paclitaxel by activating the apoptotic pathway.Materials and Methods: In this study,A549 and H1299 cells were used to detect the sensitivity of the two cells to paclitaxel.According to their sensitivity to paclitaxel,the cells were incubated with different concentrations of paclitaxel for a long time.Western blot was used to detect the two The expression of Notch3 protein in cells was compared with untreated cells.MTT assay was used to detect the effect of Notch3-siRNA and GSI synergistic paclitaxel on the proliferation of two cells compared with paclitaxel alone and control group.Then two kinds of inhibitors were used to detect the apoptosis of these two lung cancer cells by flow cytometry.And then continue to use the method of cloning to detect the two cells after different pretreatment cell cloning ability.At same time,the expression levels of Notch3 protein and NICD3(NIC6)and apoptosis-related proteins were detected by Western blot.Results: Notch3 protein was significantly overexpressed in both cell lines and increased Notch3 expression after paclitaxel administration,indicating activation of the Notch signaling pathway.After treatment with Notch3-siRNA,the IC50 of paclitaxel was significantly decreased compared with that of the control group.After treatment with GSI,the cell proliferation was significantly decreased at 24 h,48 h and 72 h after adding the paclitaxel alone.After the two inhibitors treated cells significantly increased the role of apoptosis and clonal formation capacity decreased significantly.At the same time,the two inhibitors significantly regulated the expression of apoptotic proteins and thus affected the effect of paclitaxel on cells.Notch3 siRNA and GSI significantly reduced Notch3 protein and NICD3 expression levels.GSI or siRNA was associated with down-regulation of Bcl-2 expression and up-regulation of Bax expression.Conclusion: These results indicated that synergistic effect of Notch-3-specific inhibition and paclitaxel by altering the intrinsic apoptosis pathway,which was involved in Notch-3-induced chemoresistance in NSCLC cells,and GSI inhibited Notch-3-induced chemoresistance in a concentration-dependent manner.This approach that combine Notch-3-specific inhibition and paclitaxel would be likely to apply in NSCLC.