Prediction Of Radiochemosensitivity By Next Generation Sequencing In Locally Advanced Rectal Cancer Patients Receiving Neoadjuvant Radiochemotherapy

Background:Neoadjuvant radiotherapy(RCT)combined with radical surgery has become the preferred treatment for locally advanced rectal cancer(LARC).Studies have shown that patients with locally advanced rectal cancer undergo a standard comprehensive approach would suffer from more serious complications,sequelae,more therapeutic toxicity side effects and the burden of treatment.More and more studies have shown that there is a significant difference in radiotherapy sensitivity between different rectal cancer patients.Tumor regression grades(TRG)is used to characterize treatment responsiveness,patients with high TRG scores have been shown to receive no benefit from surgery.This study focuses on identifing low-TRG scores by circulating tumor DNA(ctDNA)detection to reduce over-treatment and improve quality of life of the patients with late-stage locally advanced rectal cancer.Methods:The single-center retrospective clinical cohort study was used to collect 16 patients with locally advanced rectal patients with definite surgical resection and a clear pathologic diagnosis of tumor regression grade and divide into RCT sensitive group(pathological complete response,pCR group)and RCT none-sensitive group(none-pathological complete response,none-PCR group).The inclusion criteria were included in the standard Preoperative radiotherapy and chemotherapy(Xeloda and radiotherapy,radical surgery and adjuvant chemotherapy).Oncogene driven mutations were detected by using of next-generation sequencing to screen between the two groups.Prospective collected the plasma of 28 locally advanced rectal cancer patients at three time points which are before the new adjuvant chemotherapy,treatment and after the end of the treatment,and the fresh tumor tissue and adjacent tissue before neoadjuvant chemotherapy.Both of them detected genes mutations by using of next-generation sequencing.At the same time we collected the same period of clinical information and serological examination to analysis.Results:16 patients with locally advanced rectal cancer were examined for a total of 2774 mutations of 165 genes.There were 144 genes with the frequencies difference associated with chemoradiation sensitivity in the radiotherapy sensitive group(pCR group)and radiotherapy none-sensitive group(none-pCR group).144 genes were verified by target sequencing.The ROC curve showed that the AUC of the two detection methods was 0.972.GO enrichment and KEGG pathway was analysised.Ultimately we identified 61 genes which were closely related to chemoradiotherapy sensitivity in local advanced rectal cancer.A total of 95 mutations of 24 rectal cancer-driven genes were detected in ctDNA at the three time points.The frequencies of gene mutations in APC,KRAS and TP53 in ctDNA were different between the group(TRG grade 0-2)and the group(TRG grade 3-4)after neoadjuvant radiotherapy.The prediction accuracy of the TP53 gene,KRAS gene and APC gene were 58.3%,57.1%and 100%.The prediction accuracy of the traditional serological tumor markers CEA and CA19-9 was 38%and 34%.Conclusions:More accurate genes associated with chemoradiation sensitivity were derected by using NGS.Neoadjuvant chemoradiotherapy sensitivity of the relevant target sequencing region concentrated to the entire genome size,so that subsequent deep sequencing can be achieved.Our findings suggest ctDNA mutation profiling may be a powerful tool for predicting TRG and chemoradiotherapy sensitivity in LARC patients undergoing RCT.ctDNA testing can identify patients who can be spared from unnecessary surgical treatment.