Study On The Correlation And Clinical Significance Of Circulating Tumor Cells Undergoing The Epithelial–Mesenchymal Transition And Cytokeratin19-Positive Hepatocellular Carcinoma

Objective: To investigate the relationship between epithelial-mesenchymal transition（Epithelial-Mesenchymal Transition,EMT）of circulating tumor cells（circulating tumor cells,CTCs）of preoperative peripheral blood and cytokeratin19（cytokeratin 19,CK19）positive in hepatocellular carcinoma（hepatocellular carcinoma,HCC）,and the potential influence of that relationship on prognosis.Methods:1.Can Patrol^TMCTC-enrichment technique and in situ hybridization（ISH）were used to enrich and classify CTC according to the expression of EMT-related markers.Blood was collected from HCC patients at 1or 2 days before resection.2.HCC tissues were stained with HE and CK19 immunohistochemistry.Results of CK19 expression were collected and collated.3.Comparison of CTCs 、 CK19 expression differences in clinical data of different patients.4.Analyze the correlation between CK19 expression and CTC count and its different phenotypes,and their influence on the prognosis of patients after surgery.Results: 1.A total of 153 HCC patients（91 males and 14 females）with a mean age of 46.2 years（range: 20-72 years）meeting the enrollment criteria were included.In total,44 of 153（28.8%）HCC patients were CK19-positive,109 of 153（71.2%）HCC patients were CK19-negative.CK19-positive patients had significantly lower tumor-free survival（TFS）than CK19-negative patients（median TFS:5 vs 10 months,P=0.006）.2.In total,10（6.5%）patients failed to detect CTCs,143（93.5%）patients showed pre-surgery peripheral blood CTCs.The preoperative median CTC count was 6（range,0-76）.The cut-off value of 6for CTC count predicted early recurrence with sensitivity of 65.5%,specificity of 78.4% and AUC of 0.709.CTCs≥6 in 84 patients（54.9%）and CTCs<6 in 69patients（45.1%）.3.CK19-positive patients with CTC count ≥6 showed significantly higher percentage than CK19-negative ones（77.8% vs 46.2%）.CK19-negative patients with CTC count <6 showed significantly higher percentage than CK19-negative ones（53.8% vs 46.2%）,P=0.004.4.The distribution difference of CK19 expression in CTC subtypes（M-CTC 、E/M-CTC、E-CTC）: the proportion of M-CTC in the CK19 positive group was significantly higher than that in the CK19 negative group（mean rank: 90.07 vs71.72,P=0.0113）.The proportion of E/M-CTC in the CK19 positive group was significantly higher than that in the CK19 negative group（mean rank: 88.33 vs72.43,P=0.043）.However,there was no significant difference in E-CTC between the two groups.5.HCC patients were divided into four groups: CTCs≥6 CK19+,CTCs ≥6 CK19-,CTCs <6 CK19+ and CTCs <6 CK19-.Their median tumor-free survival time was 4 months,6 months,7 months and 18months（P <0.001）.Conclusion:CK19-positive HCC cells show a high degree of malignancy and short tumor-free survival time.CK19-positive HCC patients have more CTCs in their peripheral blood and are prone to EMT transformation,which may be an important reason for the low tumor-free survival rate after surgery.Preoperative CTC analysis combined with postoperative analysis of CK19 expression may be a new method for predicting prognosis of HCC patients after surgery.