CtDNAs Sequencing Facilitate Clinical Diagnosis And Therapy Of Non-small Cell Lung Cancer Patient

Background:Lung cancer is a malignant tumor which has the highest incidence and mortality rate among all cancer types worldwide.Non-small cell lung cancer(NSCLC)is the most predominant subtype of lung cancer.Molecular targeting therapy has been shown great success in the treatment of advanced NSCLC.Thus,an easy,sensitive,and specific way of recognizing therapeutic gene targets would help to select effective treatments,to improve physical condition and increase patient survival.Currently,tissue biopsy is the gold standard for assessing tumor molecule changes.However,due to most of NSCLC patients were diagnosed at later stages,it made surgery or biopsy difficult and dangerous.Therefore,novel non-invasive and comprehensive investigation and examination methods are urgently need for NSCLC patients.Nowadays,especially blood ctDNAs detection offers the new opportunity to aid diagnosing,monitoring tumor progression,and guiding the clinical patient treatment,by a non-invasive,real-time,and high-throughput way.Purpose:We wish to establish a non-invasive molecular diagnostic gene panel and use it to analyze ctDNAs in tumor patients.It is expected that we can utilize ctDNAs to monitor the dynamic evolution of non-small cell lung cancer during development,reveal tumor molecular heterogeneity;screen targeted drug targets,and guide clinical personalized precision treatment.Compared sensitivity and specificity of different molecular diagnostic methods provide reference for the selection of clinical diagnostic methods and evaluate the clinical therapeutic effects.Finally,it is possible that an economical gene panel was optimized for early diagnosis,clinical treatment and evolutionary monitoring of lung cancer.Methods:In this study,we collected plasma ctDNAs(circulating tumor DNAs),blood cell gDNAs,psDNAs(pleural effusion supernatant DNAs)and ppDNAs(pleural effusion pellet DNAs)from NSCLC patients,using methods of next-generation sequencing(NGS),droplet digital PCR(ddPCR),and Amplification Refractory Mutation System(ARMS).In addition,pleural effusion cells were precipitated by IHC(immunohistochemistry)for EGFR gene expression level staining.Results:The results showed that the EGFR L858 R mutation could be identified by NGS,ddPCR and ARMS.However,the EGFR T790 M mutation was identified only by NGS and ddPCR,but not ARMS.This indicates that ARMS,as an auxiliary diagnostic clinical method,has a lower sensitivity and is less reliable than NGS and ddPCR.Comparison of pleural effusion supernatant DNAs and pleural effusion pellet DNAs,pleural effusion supernatant DNAs reflected tumor changes earlier after drug treatment.In ctDNAs analysis of NGS,we also found a novel rare mutation KIT-R634 W.Conclusion:ctDNAs analysis,based on NGS of high throughput and/or ddPCR of highly sensitive,providing better diagnostic and treatment options for NSCLC patients in a non-invasive and highly sensitive manner.