Primary Resistance Of EGFR-TKIs In NSCLC Induced By MDM2 Amplification

Lung cancer,as a highly malignant disease,ranks first in China in incidence rate and mortality rate.At the same time,the five-year survival rate of lung cancer in China is lower than 20%.Based on different pathological features,lung cancer could be directly divided into small cell lung cancer(SCLC)and non-small cell lung cancer(NSCLC)while NSCLC accounts for 85% of all clinical cases.In recent years,the rapid development of targeted sequencing lead to the resultant wildly use EGFR-TKIs in NSCLC.However,20%-30% of NSCLC patients with EGFR sensitive mutations demonstrated the primary resistance to EGFR-TKIs,which leads to failure of EGFR-TKIs treatment.The mechanisms remain unclear which might involve multiple molecules and signaling pathways.Therefore,it is necessary to further study the mechanism of EGFR-TKIs primary resistance in NSCLC patients..Objective The research aims to study the significance of MDM2 amplification in the treatment of EGFR-TKIs among NSCLC patients.Through the verification of experiments on cell lines,the effects of MDM2 amplification and MDM2 protein overexpression on the therapeutic sensitivity of non-small cell lung cancer cell line EGFR-TKIs were estimated,so as to provide theoretical basis for further exploring the possible molecular mechanism of primary resistance to EGFR-TKIs.Methods 141 NSCLC cases in our center were involved and 4 patients were detected harboring the EGFR sensitive mutations and MDM2 amplification through the next generation sequencing.4 patients administrated with EGFR-TKIs were followed up to determine the effect of MDM2 amplification in the treatment.Regularly cell culture and plasmid transfection were used to construct MDM2 overexpression non-small cell lung cancer cell line and the vector control cell line as well.MTT method was used to calculate the inhibitory rate of erlotinib,the first generation of EGFR-TKI drug,on cell proliferation of the cell lines.The IC50 of the drug in the treatment for NSCLC cell line was calculated.In the meantime,the expression of the corresponding proteins was detected by Western blotting.The changes of MDM2 related signaling pathways were identified by enrichment analysis and other bioinformatic methods.Results 4 NSCLC patients with EGFR sensitive mutation and MDM2 amplification demonstrated primary resistance to EGFR-TKIs,and the median progression free survival was reduced to 6.3 months(5.1-8.5 months).In vitro,MDM2 amplification lead to erlotinib resistance in NSCLC.MDM2 overexpression increased the IC50 value of erlotinib in HCC2279 cell line [from 36.5μm(43.4μm-79.6μm)to 57.6μm(24.1μm-58.7μm)],and decreased the inhibition of cell proliferation induced by erlotinib.MDM2 amplification did not change the expression level of EGFR protein,but significantly up-regulated the expression level of the downstream ERK1/2 protein.It suggests that MDM2 amplification leads to the activation of downstream pathways in EGFR signaling pathway.In addition,MDM2 amplification predicted poor prognosis in NSCLC patients.Progression free survival(P=0.007)and overall survival(P<0.001)were at a disadvantage in patients with high MDM2 expression.At the same time,MDM2 amplification may shorten the progression free survival of patients with NSCLC treated with EGFR-TKIs(P<0.001).Through the follow-up analysis,we found that ERBB2 signaling pathway might be the potential pathway of MDM2 amplification leading to EGFR TKIs primary drug resistance,which needs further experimental verification.Conclusion1.MDM2 amplification could induce the primary resistance to EGFR-TKIs in NSCLC patients with EGFR sensitive mutations;2.MDM2 protein could be used as a novel marker to predict the poor prognosis of patients with NSCLC;3.MDM2 amplification can be used as a novel biomarker and therapeutic target for patients with NSCLC.