The Reverse Effect Of Everolimus On Lapatinib Resistance In Breast Cancer And Its Molecular Mechanism

Objective To investigate the reversal effect and mechanism of drug resistance of everolimus on lapatinib-resistant cells in breast cancer.Methods The lapatinib-resistant breast cancer cell SKBR3-LR,which was established in the early stage of the laboratory,was treated with different concentrations of everolimus,and the appropriate concentration was selected according to its growth inhibition rate.The optimal concentration of everolimus was used to combine with lapatinib in the treatment of lapatinib-resistant cell SKBR3-LR.The cell viability of the combination of everolimus with lapatinib or lapatinib alone was decected by CCK-8,and calculate IC₅₀0 value.The drug-resistant cell SKBR3-LR was treated with the optimal concentration of everolimus in combination with different concentrations of lapatinib.CompuSyn software was used to calculate the combined effect index?CI?value of each group.The resistant cells SKBR3-LR were divided into blank control group,lapatinib treatment group,everolimus treatment group and everolimus combined with lapatinib treatment group;flow cytometry was used to detect the cell cycle of SKBR3-LR in each group;Cell scratch was used to detect migration in each group;Western blot was used to analyse the expression of AKT,mTOR,S6 and related protein.Results The combination of everolimus with lapatinib reduced the IC₅₀0 of SKBR3-LR to lapatinib from?27.13±0.3?to?6.79±0.04?;The CI index of 20nmol/L everolimus combined with 5?mol/L,10?mol/L,20?mol/L,40?mol/L lapatinib were 0.24367,0.27861,0.28219,0.36679,0.41667;The percentage of G1 phase cells in lapatinib combined with everolimus group?69.84±0.93?%was significantly higher than that in control group?52.91±1.64?%?P<0.001?,which was also significantly higher than that in lapatinib group?59.77±0.52?%?P<0.001?,and everolimus group?58.73±0.64?%?P<0.001?;Wound healing assay showed that compared with the healing rate of the control group?35.76%±5.35?,the healing ability of the lapatinib group?24.04%±3.25?,the everolimus group?23.88%±0.41?and the combined group?15.17%±3.14?was significantly reduced?P<0.05?;Western blot analysis showed that compared with SKBR3,p-mTOR and vimentin was up-regulated,Ecadeherin was down-regulated in SKBR3-LR;the SKBR3-LR treated with 20nmol/L everolimus or 10?mol/L lapatinib,or combination surely degrated the expression of p-mTOR,p-AKT and p-S6.Conclusion Everolimus can partially reverse the resistance of breast cancer cell SKBR3-LR to lapatinib,which may be related to down-regulation of AKT/mTOR/S6 pathway.