Co-expression Of TIM-3 And CEACAM1 Promote T Cell Exhaustion In Colorectal Cancer Patients And Their Effects On The Function Of CD8~+ T Lymphocytes

Part Ⅰ:TIM-3 is a potential prognostic marker in solid tumors:a systematic review and meta-analysisObjective:Accumulated studies have demonstrated the important role of T cell immunoglobulin-and mucin-domain-containing molecule-3(TIM-3)in various solid tumors and indicated its correlation with patients’ survival.To further verify the prognostic significance of TIM-3 in cancer patients and its correlation with tumor,we performed this analysis.Methods:We performed this meta-analysis including seven studies searched from PubMed,Web of Science,and Embase till July 2016.We did a quality assessment for all the enrolled studies using the NOS system firstly,then a total of 869 patients with solid tumors were used to analyze the association between TIM-3 expression and patients’ overall survival(OS).Besides,we also calculated the correlation between TIM-3 and tumor stage.Results:1.The pooled results showed that higher expression of TIM-3 was significantly correlated to shorter OS(7 studies,HR=1.89;95%CI:1.38-2.57;P<0.001).2.We further found that higher TIM-3 expression was associated with advanced tumor stage(3 studies,Ⅲ/Ⅳ vs.Ⅰ/Ⅱ,RR=2.02;95%CI:1.45-2.81;P<0.001).Conclusion:Our study highlights the role of TIM-3 as a potential prognostic marker and a promising therapeutic target in solid tumors.Part Ⅱ:Co-expression of TIM-3 and CEACAM1 promotes T cell exhaustion in colorectal cancer patientsObjective:To investigate the expression of TIM-3 and CEACAM1 on circulating CD8+ T cells and tumor infiltrating lymphocytes(TILs),and their correlation with CRC patients’ clinical features.Methods:65 diagnosed colorectal cancer(CRC)patients and 38 healthy controls were enrolled in this study,flow cytometry analysis was used to detect the expression of TIM-3 and CEACAM1 on circulating CD8+ T cells and tumor infiltrating lymphocytes(TILs).The correlation between their expression and clinical features was determined.IFN-y production was evaluated among different cell subtypes to illustrate the effect of TIM-3 and CEACAM1 on T cell exhaustion.Cox regression model was used to predict the risk factors of colorectal cancer.Results:1.TIM-3 and CEACAM1 were both highly expressed on circulating CD8+ T cells in CRC patients compared with healthy controls.2.TIM-3 and CEACAM1 were elevated on TILs compared with paraneoplastic T cells.Their co-expression is higher on TILs.3.TIM-3 and CEACAM1 is related to T cell exhaustion in CRC progression.Furthermore,TIM-3+CEACAM1+ CD8+ T cells represented the most dysfunctional population with the least IFN-y production.4.At last,the expressions of TIM-3 and CEACAM1were correlated with advanced tumor stage and could be independent risk factors for CRC.Conclusion:We for the first time to our knowledge suggested that co-expression of TIM-3and CEACAM1 can mediate T cell exhaustion and may be potential biomarkers for CRC prediction,highlighting the possibility of being immunotherapy targets.Part III:The effects of TIM-3 and CEACAM1 on the function of CD8+ T lymphocytesObjective:The purpose of this article is to investigate the effects of TIM-3 and CEACAM1 on the function of CD8+T lymphocytes.Methods:After we obtained the human peripheral blood sample,Ficoll density and gradient centrifugation was utilized to get the PBMCs from single cell suspension.Antibodies of CD8,TIM-3 and CEACAM1 were used for Flow cytometry staining.Finally we got four subgroups of cells through Fluorescence Activated Cell Sorting:TIM-3-CEACAM1-;TIM-3 CEACAM1+;TIM-3+CEACAM2-;TIM-3+CEACAM1+.Cell Counting Kit-8 was used to detect cell proliferation,while real-time PCR was used to evaluate expressions of cell cycle associated genes,apoptosis related genes and anti-tumor factor genes.Results:1.Compared with TIM-3/CEACAM1 double negative subgroup or the single positive subgroups,the double positive one expressed lower expression of CCND1,CCNE1 and CDK4,while expression of p16,p21 and p27 had no significant differences;2.The TIM-3/CEACAM1 double positive cells had weaker proliferation ability than the other three subgroups;3.The TIM-3/CEACAM1 double positive cells secreted less IL-1β,IL-8 and IL-17 than the other three groups with higher IL-6 expression;4.The production of TNF-α and IFN-γ is much less in TIM-3/CEACAM1 double positive subgroup than the other three ones.5.The expressions of anti-apoptosis protein Bcl-2 and Bcl-xl were much lower in TIM-3/CEACAM1 double positive subgroup compared with the other threes.Conclusion:TIM-3 and CEACAM1 could inhibit cell cycle of CD8+T cells and its cell proliferation ability,promote apoptosis of T lymphocytes,inhibit its production of TNF-a and IFN-γ to block the immune response of T cell on tumor cells,thus promoting tumorigenesis,tumor development and metastasis,which may provide a potential target for immunotherapy.