Study Of Ibrutinib In CCRF-CEM Cell Lins Proliferation And Induction Of Apoptosis

Objective:To detect the effect of Ibrutinib on the proliferation of CCRF-CEM and its apoptosis in acute T-lymphocytic leukemia cell line,and to explore the role of PI3 K pathway in leukemia cell apoptosis induced by Ibrutinib.To provide new therapeutic drugs for clinical treatment of acute T-lymphocytic leukemia and to expand the therapeutic range of ibrutinib.Methods1.Cell proliferation inhibition rate test.Using different concentrations(0,5,10,15,20,25,30,35,40,45,50,75,100μmol/L)of Ibrutinib on CCRF-CEM cell line,respectively After 24 h,48h,72 hours,CCK-8 method was used to detect the inhibition rate of cell proliferation and observe the effect of drugs on the proliferation of CCRF-CEM cells.2.Inverted microscope observation of cell morphology changes at different drug concentrations3.After the cells were treated with different concentrations of Ibrutinib,the apoptotic rate was measured by Annexin V-FITC/PI double staining and the effect of Ibrutinib on the apoptosis of CCRF-CEM was observed.Observation of Cellular Morphological Changes by Fluorescence Microscopy4.Real-time fluorescent quantitative PCR was used to detect CCRT-CEM cells treated with different concentrations of Ibrutinib.The expression of PTEN and BCL-2mRNA was detected and the effects of ibrutinib on these genes were observed.5.After the cells were treated with Ibrutinib,the expression of PTEN and BCL-2protein was detected by Western blotting,and the changes of PTEN and BCL-2 protein expression levels were observed by Ibrutinib.Results1.After CCRF-CEM cells were treated with different concentrations of Ibrutinib,theinhibition rate of cell proliferation was significantly increased(P<0.01)and the inhibition rate increased with the increase of drug concentration.After the cells were treated with Ibrutinib at the same concentration,the inhibition rate also showed significant changes(P<0.01).The inhibition time was time-dependent,but at low concentrations,the 72 h cell proliferation inhibition rate was lower than that of 48 h.2.After Ibrutinib treated CCRF-CEM cells,the number of abnormal cells increased,cells shrank,ruptured,nuclear fragmentation and pyknosis3.After Ibrutinib treatment of cells,the apoptosis rate was significantly increased(P<0.01),and was drug-dependent.4.After Ibrutinib was used to treat CCRF-CEM cells,the mRNA expression of PENT and BCL-2 was significantly increased(P<0.01),and there was concentration dependence.5.After treatment with Ibrutinib on CCRF-CEM cells,the expression of PENT and BCL-2 at the protein level was significantly higher(P<0.01).ConclusionIbrutinib can act on acute T-lymphocytic leukemia,inhibit the proliferation of leukemia cells through the PI3 K pathway,and induce leukemia cell apoptosis.Ibrutinib may become a new drug for the treatment of acute T-lymphocytic leukemia.