Triptolide Inhibits The Proliferation Of MV411 Cell And PI3K-AKT-mTOR Pathway Mechanism Research

Objectives:Acute myeloid leukemia(AML)is a malignant clonal disease,which is originated in the bone marrow hematopoietic stem and progenitor cells.FLT3 / ITD mutation is one of the important genetic mutations in AML.It is also an important sign of poor prognosis.This kind of patients with poor efficacy of conventional chemotherapy.At present,targeted drug therapy of patients with FLT3/ITD mutation with AML has become a hot research topic,and has achieved some effect in clinical application.However,drug resistance and relapse after targeted therapy has become a major problem in the treatment of patients with AML associated with FLT3/ITD mutation,so it is necessary to seek new therapeutic drugs and new targets.In recent years,traditional Chinese medicine Tripterygium wilfordii has attracted much attention from scholars both at home and abroad for its anti-inflammatory,immunosuppressive and anti-tumor effects.Triptolide(TP)is one of the main active ingredients of Tripterygium wilfordii,related research reports show that TP for acute myeloid leukemia HL60 cells and chronic myeloid leukemia K562 and K562G01 cells has significantly inhibited the proliferation.So does TP have an inhibitory effect on FLT3/ITD mutant MV411 cells? What is the possible mechanism? In this study,we investigated the effect of TP on proliferation inhibition and apoptosis of MV411 cells and the expression of PI3K-AKT-mTOR pathway,FLT3,PTEN,PI3 K,AKT and mTOR by in vitro method.To explore the possible anti-tumor mechanism and provide theoretical basis for the treatment of patients with AML FLT3 / ITD mutation.Methods:1.MTT was used to determine the proliferation inhibition of TP cells with different concentrations of24 h,48h,72 h in MV411 cells,and the IC50 values were calculated.2.The apoptosis rate of 48 h and 72 h was measured by flow cytometry.3.Real time quantitative PCR was used to detect the expression levels of PI3K-AKT-mTOR pathway related genes FLT3,PTEN,PI3 K,AKT,mTOR.Results:1.The results of MTT showed that TP significantly inhibited the proliferation of MV411 cells,and with the increase of drug concentration and treatment time increases,showed time dose dependent,choose 48 hours for the TP IC50 value is 16.92nmol/L.2.Flow cytometry results showed: the 48 hour time point 0,10,20nmol/L TP corresponds to the average apoptosis rate were(3.30 + 0.20)%,(17.10 + 0.36)%,(35.67 + 0.61)%;the 72 hour time point 0,10,20nmol/L TP corresponds to the average apoptosis rate respectively(7.37 + 0.32)%,(49.33 + 0.40)%,(68.92 + 0.11)%.The apoptosis rate was found to increase with the increase of concentration.3.Real time fluorescence quantitative PCR detection showed that the choice of 0,5,10,20nmol/L TP in MV411 cells after 48 h by real-time fluorescence quantitative PCR assay,PI3 K,AKT,FLT3,mTOR m RNA decreased,PTEN m RNA increased,and showed a dose dependent.Conclusions:1.TP inhibited the proliferation of MV411 cells in a time-dose-dependent manner.2.TP can induce apoptosis of MV411 cells.3.TP acted on MV411 cells,and found that FLT3,PI3 K,AKT,mTOR levels of m RNA decreased,and the PTEN level of m RNA increased.TP may induce apoptosis of tumor cells by affecting the expression of related genes in PI3K-AKT-mTOR pathway.