Effect Of Dyrk1B Inhibitor AZ191 On Apoptosis And Cell Cycle Of Ovarian Cancer Cells

Objective: To detect effect of dual specificity tyrosine phosphorylation regulated kinase 1B(Dyrk1B)inhibitor AZ191 on the proliferation,cell apoptosis and cell cycle,to study the mechanisms of Dyrk1 B in cell apoptosis and cell cycle of ovary cancer cells.Methods:(1)Methyl thiazolyl tetrazo-lium(MTT)method assay was used to determine the influence of AZ191 on the growth of OVCAR3 and OV2008 cells.(2)Cell clone assay was used to investigate the effect of AZ191 on ovarian cancer cells.(3)The apoptotic rate of OVCAR3 and OV2008 cells was detected by flowcytometry with Annexin V/PI binding assay.(4)Flow cytometry with propidium iodide staining was used to determine the influence of AZ191 on cell cycle of OVCAR3 and OV2008.(5)The expressions of apoptosis and cell cycle related protein were detected by western blot after treatment of AZ191 in ovarian cancer cells.Results:(1)Different concentration of AZ191(0,1,2.5,5,10 ?M)for 24,48,72 hours acting on OVCAR3 and OV2008 cells inhibited the cellular proliferation in a time-and concentration-dependent manner.The values of IC50 of AZ191 on OVCAR3 and OV2008 are 9.78 ?M,6.08 ?M,5.46 ?M and 9.05 ?M,5.47 ?M,3.00 ?M,respectively.(2)The clony formation of both OVCAR3 and OV2008 cells treated by AZ191(5 ?M)for 2 weeks was significantly lower than the control group(0 ?M)(P<0.05).(3)The apoptotic rate of both OVCAR3 and OV2008 cells was increased significantly(P<0.05)after 48 h-treatment of AZ191(0,5,10 ?M)and in a concentration-dependent manner.(4)The increased cleavage of PARP in a dose-dependent manner was measuresd after treatment of AZ191(0,5,10?M)for 48 hours in OVCAR3 cells,concomitant with upregulation of Bim,a Bcl-2 family member.Aslo,increased protein levels of phosphorylated H2 AX and CHK2 were also found.(5)OVCAR3 and OV2008 cells were arrested at the G2/M-phase of cell cycle detected by flow cytometry with propidium iodide staining after treated by different concentrations of AZ191(0,5,10?M)for 48 hours,coincident with an increase in cyclinB1 and cyclinD1 as well as decrease in P27kip1.Conclusion: Dyrk1 B inhibitor AZ191 can inhibit cellular proliferation in a timeand concentration-dependent manner in ovary cancer cells.In addition,it induced cell apoptosis and a G2/M-phase cell-cycle arrest.Moreover,The data showed that AZ191 increased the amount of cleaved PARP,accompanied by alterations of cyclinB1,cyclinD1 and P27kip1 as well as activations of Bim,p-H2 AX and p-CHK2.In conclusion,the present study showed that Dyrk1 B inhibitor AZ191 induced cell apoptosis and cell cycle arrest.The function may be related to the Bcl-2 family,ATM/CHK2 pathway and cell cycle regulation.Therefore,Dyrk1 B possibly be a candidate target for the treatment of ovarian cancer in future.