Study On The Effect Of FTO Inhibitor Combined With PD-1/PD-L1 Inhibitor In Non-small Cell Lung Cance

Objective(s):To clarify the role of FTO inhibitor combined with PD-1/PD-L1 inhibitor in non-small cell lung cancer and explore its possible mechanism of action,so as to provide a theoretical basis for the search for a new treatment of non-small cell lung cancer.Method(s):PCR was used to detect the relative expression of FTO in various NSCLC cells(A549,H1299,H1975,PC9)and human normal bronchial epithelial cells(BEAS-2B),so as to screen suitable cell lines(H1299,A549).Non-small cell lung cancer cells were treated with different concentrations of PD-1/PD-L1 inhibitor(Carelizumab)and FTO inhibitor(FB23-2).The cell proliferation rate was detected by CCK-8 method,and IC50(half inhibitory concentration)was determined for both cells.According to IC50,appropriate concentration gradient was set for drug combination experiment,and CCK-8 experiment was used to determine the effect of drug combination on cell proliferation.The cells were divided into four groups: FTO inhibitor group;PD-1 inhibitor group;FTO inhibitor +PD-1 inhibitor group;Blank group.The migration and invasion of each group were detected by scratch test and Transwell invasion test.NSCLC cells were co-cultured with mouse splenic lymphocytes.Result(s):In the four non-small cell lung cancer cell lines detected,A549,H1299,H1975 and PC9,FTO expression was higher than that of normal bronchial epithelial cells BEAS-2B(P<0.05).NSCLC cell line H1299/A549 was treated with FTO inhibitor(FB23-2)at concentrations of 0.02,0.04,0.06,0.08,0.1mg/ml and PD-1 inhibitor(Carelizumab)at concentrations of 5,10,15,20,25mg/ml,respectively After h,OD value at 450 nm is detected.The results showed that the proliferation ability of H1299/A549 cell line was significantly inhibited with the increase of FB23-2concentration and the extension of the action time.In H1299 cell line,when the two drugs were completely cross acted for 24 h,the combination of high concentration and cell inhibition rate > 85%,the two drugs had synergistic effect,while at low concentration combination and cell inhibition rate;Antagonism was found in 85% of the cases.When the two drugs were completely cross acted for 48 h,at high concentration,the cell inhibition rate >75%,the two drugs showed antagonistic action,while at the cell inhibition rate <75%,there is synergy.After 72 h of complete cross-action,the two drugs showed synergistic effect in both high concentration combination and low concentration combination.In the sequential administration experiment,compared with the simultaneous administration,when FB23-2 was given 6,12,24 and 36 h in advance,the two drugs combined for 42,36,24 and 12 h had a stronger inhibitory effect on cells.And when FB23-2 was given 6h in advance,the inhibitory effect of FB23-2 on H1299 was greater than that of Carelizumab given 6h in advance(P<0.05).In A549 cell line,when the two drugs were fully cross acted for 48 h,the combination of high concentration and cell inhibition rate;The synergistic effect of the two drugs was observed at 95%.They were antagonistic 95% of the time.When the two drugs were fully cross treated for 72 h,the combination of the two drugs in high concentration and the inhibition rate of cells.At 95%,the two drugs showed synergistic effect,when the cell inhibition rate > 95% of the time,the two drugs showed antagonistic action.Transwell and scratch tests showed that compared with Carelizumab alone,Carelizumab combined with FB23-2 significantly inhibited the invasion and migration of non-small cell lung cancer cell lines H1299 and A549(P<0.05).Conclusion(s):In vitro experiments,compared with PD-1/PD-L1 inhibitor alone,FTO inhibitor combined with PD-1/PD-L1 inhibitor can significantly inhibit the proliferation,migration and invasion ability of non-small cell lung cancer cells.