Clinical Significance Of High Expression Of Inhibitory Receptor KIR2DL1 On CD8+ T Lymphocyte Surface Induced By Fusobacterium Nucleatum In Esophageal Squamous Cell Carcinoma

Background: Esophageal cancer is one of the most common malignant tumors in the world.Esophageal squamous cell carcinoma（ESCC）is the most common histopathological type of it,with the highest morbidity and mortality in Asia and Africa.The latest research shows that Fusobacterium nucleatum（Fn）is closely related to the occurrence and development of ESCC.KIR2DL1 is an important inhibitory receptor on the surface of CD8+ T cells.The immunosuppression mediated by KIR2DL1 is one of the reasons for tumor immune escape.It is also a major obstacle to tumor immunotherapy.Therefore,it is of great significance to further explore the effect of Fn on the expression of inhibitory receptor KIR2DL1 on THE surface of CD8+T cells and its mechanism in the development and development of ESCC,so as to identify new biomarkers for ESCC and effective molecular therapeutic targets.Objectives: To analyze the correlation between the induction of CD8+ T cell surface inhibitory receptor KIR2DL1 by Fusobacterium nucleatum（Fn）in cancer tissues of patients with esophageal squamous cell carcinoma,clinicopathological features and 5-year survival,and to explore its clinical significance and prognostic value.Methods:1.The induction effect of Fn on the CD8+T cell surface inhibitory receptor KIR2DL1 was analysed by Fn infection of human CD8+T cells in a co-culture system with ESCC cells.2.The effect of Fn infection on the sensitivity of CDDP treatment of tumour cells was analysed by flow cytometry to detect the apoptosis rate of tumour cells in the co-culture system after CDDP treatment.3.The effect of Fn infection on the sensitivity of CDDP to treat tumour cells was verified in vitro by observing the tumour size of mice treated with CDDP in the normal ESCC subcutaneous tumour model as well as in the Fn-infected ESCC subcutaneous tumour model.4.RNAscope and immunohistochemistry were used to detect Fn infection and KIR2DL1 expression on the surface of CD8+T cells in the cancerous and corresponding paracancerous tissues of 196 ESCC patients.5.Analysis of the induction effect of Fn on KIR2DL1 on the surface of CD8+T cells correlated with clinicopathological features.6.The impact of clinicopathological factors on the prognosis of ESCC was analysed using the COX regression method.7.Survival curves were plotted using the Kaplan-Meier method.8.The effect of induction of KIR2DL1 on the surface of CD8+ T cells on survival was analysed using a Log-rank test.Results:1.CD8+ T cells co-cultured with tumor cells or infected with Fn can cause increased KIR2DL1 expression,and the combination of the two can cause the most significant increase in KIR2DL1 expression（P < 0.05）.2.Apoptosis rates of tumour cells in co-culture systems after CDDP treatment were analysed by flow cytometry and Fn infection was found to reduce the potency of tumour cell responses to CDDP.3.By analyzing the tumor size of the ESCC subcutaneous tumor model mice treated with CDDP,we found that the tumors of the ESCC subcutaneous tumor model mice in the Fn-infected group were significantly larger than those of the normal ESCC subcutaneous tumor model mice,and their sensitivity to CDDP treatment was reduced.4.By using RNAscope and immunohistochemistry,a significant concordance between Fn infection and high expression of KIR2DL1 on the surface of CD8+T cells was found in 196 ESCC patients（P < 0.05）.The positive rate of Fn infection in ESCC patients’ cancer tissues was significantly higher than that in the corresponding paracancerous tissues,and the positive rate of KIR2DL1 expression on the surface of CD8+ T cells in cancer tissues was significantly higher than that in the paracancerous tissues（P < 0.05）.5.Patients with Fn+CD8+KIR2DL1-positive ESCC were mostly men who smoked and drank alcohol,and were also characterised by low tumour differentiation,high infiltration,frequent early lymph node metastases and more advanced clinical stage（P <0.05）.6.The analysis of prognosis revealed that alcohol consumption,degree of differentiation,depth of infiltration,lymph node metastasis,clinical stage and the Fn-induced CD8+T cell surface KIR2DL1 high expression positive group were independent risk factors affecting the prognosis of ESCC patients.7.The 5-year survival rate of patients in the Fn+CD8+KIR2DL1 positive group was23.33%,with a median survival time of（20.00±6.16）months,significantly lower than the negative group at 38.57% and（42.00±5.23）months（P <0.05）.Conclusion: Long-term smoking and drinking can lead to a harsh oral environment,and Fn is more likely to be infected and colonized in this environment,thereby inducing high expression of KIR2DL1 on the surface of CD8+ T cells,weakening the body’s anti-tumor immune response,and promoting the malignant progression of esophageal squamous cell carcinoma.