| Objective: Cholangiocarcinoma is a heterogeneous malignant tumor originating from the bile duct epithelium,accounting for 50%?60% of the malignant tumors of the biliary tract.It is a rare form of cancer in the population.The accurate diagnosis of cholangiocarcinoma is difficult.The disease progresses quickly which is easy to invade and metastasize.The malignant degree is high.The prognosis of the patients is poor,and the 5-year survival rate is only 5%?15%.Clinically,surgery is the most important method for the treatment of cholangiocarcinoma,but the anatomic location of cholangiocarcinoma is relatively special.Tumor size is small,so it is difficult to obtain tumor tissue.Tumor heterogeneity is obvious,resulting in low clinical diagnostic accuracy and difficulty in evaluating the surgical effect.Currently,the tumor markers for clinical diagnosis and surgical effect evaluation are mainly CA199 and CEA,but the low sensitivity and specificity cannot meet the requirements.Therefore,it is particularly important to develop non-invasive diagnostic and surgical effect evaluation methods.In this study,high-depth targeted sequencing was conducted on the genomic DNA(g DNA)of plasma cfDNA and white blood cells of patients with confirmed cholangiocarcinoma and suspected cancer patients,so as to improve the preoperative diagnostic accuracy,postoperative surgical effect and prognosis assessment of patients.Methods: Blood samples of 19 patients with cholangiocarcinoma and 17 patients with suspected cholangiocarcinoma were collected from the department of pancreatic and bile surgery,Shanghai Ren Ji hospital affiliated to Shanghai Jiao Tong university from July 2017 to August 2019.The highly sensitive UMIs(unique molecular identifiers)technology was used for the deep sequencing of 93 gene targets(20000×)with a detection limit of about 0.05%.Combined with the clinical information of patients,data analysis was conducted to distinguish cancer patients from suspected cancer patients by comparing the number and mutation types of SNVs of cfDNA.ROC curve was drawn to determine the sensitivity and specificity of the diagnosis.The overall Deviation of mutation Frequency between cfDNA and white blood cell g DNA before and after operation was calculated by Allele Frequency Deviation(AFD)algorithm to predict the surgical effect.Results: the diagnostic sensitivity of cfDNA was 70.6% and the specificity was 83.3%.CA199: sensitivity: 86.7% specificity: 81.8%;CEA sensitivity: 73.3% specificity: 100%.As an auxiliary diagnostic marker,cfDNA is combined with the original tumor markers,which can make the sensitivity and specificity of clinical diagnosis reach more than 95%.Furthermore,the dynamic relationship between cfDNA changes and tumor markers in the samples was good.At the level of analysis of a single gene,the sensitivity and specificity of the effect of EGFR and ROS1 on the cancer combination in the suspected cancer group reached more than 90%,which is a good marker gene for the differentiation.Combined with the overall survival of cancer patients,it was found that patients with a smaller AFD value had better surgical outcomes,while patients with a larger AFD value had worse surgical outcomes.The AFD value of cfDNA can indicate the prognosis of cancer patients,and the combination of other tumor markers can improve the prognosis of cancer patients.Conclusion: As an auxiliary diagnostic biomarker of cholangiocarcinoma,cfDNA can effectively improve the sensitivity and specificity of preoperative diagnosis.A priori evaluation can be made on the surgical effect of some patients and the prognosis of cancer patients.In a cholangiocarcinoma diagnostic test,cfDNA sequencing should be considered if tissue sampling is unsafe,infeasible,or unsuccessful.The AFD value of cfDNA can indicate the prognosis of cancer patients,and the combination of other tumor markers can improve the prognosis of cholangiocarcinoma cancer patients. |
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