Research On Relationship Between The Different Phenotypes Of Circulating Tumor Cells And Esophageal Squamous Cell Carcinoma Clinical Characteristics

Background and PurposeThe circulating tumor cells(CTCs)in the peripheral blood have already been used in early diagnosis and the assessment of curative effect.CTCs has been considered as a noninvasive “liquid biopsy” and was expected to replace “surgical exploration for biopsy” and “needle aspiration biopsy”.So far,there are many challenges in the technology for isolation and characterization of rare CTCs,and requires our continuing efforts that contribute to the technical progress.Therefore,it makes sense to figure out whether the different clinical characteristics would lead to the differential phenotypes of CTCs.As we know,the primary tumors with an increasingly mesenchymal phenotype may have heightened organization invasiveness and metastatic capacity,and hence the identification of CTCs with a mesenchymal phenotype may be of value in the evaluation of prognosis.The aim of this research is to evaluate the diagnostic value of CTCs with expressing epithelial-mesenchymaltransitions(EMT)biomarkers in esophageal squamous cell carcinoma(ESCC)patients and investigated the association with their clinical parameters.MethodsWe included 174 patients with ESCC as the observation group and 40 healthy volunteers as negative control group,and acquiring 5m L blood sample from everyone.The method we used for CTCs isolation and characterization as the following steps:lysed erythrocytes;centrifuge;fixed;sample filtering;in situ hybridization;signal amplification,and so on.Ultimately,we could use a fluorescent microscope and the automatic analysis system of CTC to detect and identify CTCs.Receiver Operating Characteristic(ROC)curve analysis was used to evaluate the sensitivity and specificity of this method as an early diagnose of ESCC.According to the different treatment stages,blood samples from ESCC patients were analyzed qualitatively and quantitatively for CTCs in peripheral blood.Result1.The total CTCs in 174 ESCC patients was significantly higher than that in 40 healthy volunteers,the area under ROC curve of epithelial CTCs,hybrid epithelial/mesenchymal CTCs,mesenchymal CTCs and total CTCs from the subjects were 0.813,0.961,0.782 and 0.990,respectively.The clinical diagnostic critical point of each phenotypes of CTCs were 2,1,0 and 3,respectively.As it turns out,using this method for isolation and characterization of rare CTCs has favorable effective functions for the diagnosis,the sensitivity in this research is more than 90%.2.For 174 ESCC patients,the later the clinical stage is,the more the mesenchymal CTCs(P=0.006).3.Follow-up blood specimens were obtained from neoadjuvant chemotherapy in ESCC patients.Complete response(CR),partial response(PR),stable disease(SD)and progressive disease(PD)after treatment were seen in 2,20,13 and 4 patients,respectively.Patients with a negative mesenchymal CTCs status after two cycles of chemotherapy had a good chemosensitivity(P=0.05),and the positive rate of mesenchymal CTCs in SD/PD(70.6%)was significantly higher than that in CR/PR(36.4%)group.4.We compare the changes in CTCs between preoperative and postoperative of47 ESCC patients,and the analysis revealed that the total CTCs of postoperative patients were decreasing(P=0.005),however,mesenchymal CTCs were increasing dramatically(P=0.006).ConclusionThis research applied a novel concept and approach for isolation and characterization of rare CTCs in ESCC,our practice is to combine the two approaches of “isolation by size of tumor cells” and “fluorescence in situ hybridization(FISH)”,and achieve a higher sensitivity of this approach.Mesenchymal CTCs as a biochemical marker of ESCC has a certain value in diagnosis,which is significantly correlated with the clinical stage and chemosensitivity of the tumor.