Establishment Of A Novel EGFR Mutations Detection Method Based On PCR-GoldMag?Lateral Flow Assays And Its Applications For Genotyping Of EGFR

Lung cancer is the most common cause of cancer death,accounting for one third of all deaths from cancer worldwide with a 5-year survival rate at only around 15%.Non-small-cell lung cancer(NSCLC)is the major type of lung cancer,constituting about 75-85%of lung cancer cases.More than 70%of patients with NSCLC are diagnosed with advanced stage disease,lost the chance of operation.Recently,the important driving gene of NSCLC-epidermal growth factor receptor(EGFR)as a molecular target for cancer treatment has received widespread attention.Studies have revealed that activating mutations in the EGFR gene correlates to sensitivity to TKIs.EGFR mutation is the key predictor of the response to TKIs in NSCLC.However,the effect of TKIs is limited in time because of the emergence of drug resistance.The commonest cause of acquired mechanism of resistance associated with a T790M mutation in EGFR.Therefore,testing of EGFR mutations(including sensitive mutations and drug-resistant mutations)is of great significance for diagnosis and treatment guideline in NSCLC.This study aims to develop a novel method for rapid detection of EGFR mutations,which was integrated allele-specific PCR with gold magnetic nanoparticles(GoldMag)-based lateral flow assay(LFA).It includes the detection of two most common mutations(delE746-A750 and L858R)and a drug resistance mutation(T790M)in EGFR.First,the positive and negative controls were constructed and newly designed mutation-specific primers were used to optimize the experimental conditions and evaluated its specificity and sensitivity.A total of 40 formalin-fixed paraffin-embedded(FFPE)tissue samples from lung cancer patients were collected and detected the EGFR gene mutations by using our PCR-LFA system,then the results were verified with direct sequencing and qPCR method.The results indicated that our method has obvious advantages to analyze of clinical samples and offer a more sensitive alternative to direct sequencing for the detection of EGFR mutations.