| Background and Objective: Pancreatic cancer is a highly common malignancy worldwide and is considered the ninth leading cause of cancer death in China. This disease is mainly characterized by rapid progression and metastasis. It is about 80% of patients with unresectable stages at diagnosis such as distant metastases or locally advanced tumors. Surgery remains the standard treatment for early-stage pancreatic cancer. With the development of medical techniques, novel treatment options can improve the prognosis of patients with unresectable pancreatic tumors. However, the 5-year survival rate for all patients with pancreatic cancer is less than 5%. Even patients who had undergone margin-negative (R0) surgical resection attained a 5-year survival rate of no more than 20%. Several studies elucidated that immune suppression of the tumor microenvironment was the key for cancer growth, metastasis, and even tumor immune escape. IDO is a rate-limiting enzyme that catabolizes tryptophan into a stable metabolite via the kynurenine pathway. IDO activity considerably influences immune regulation and exerts immunosuppressive effect. Overexpression of IDO correlates with poor clinical outcome in several cancer types. However, the expression of IDO in pancreatic cancer and its prognostic value have not been intensive studied. We designed clinical research and basic experiments to explore the effect of IDO in pancreatic cancer progression and effect of IDO related immunotherapy.Methods: (1)We performed immunohistochemical staining and Western blot analysis for IDO expression in both pancreatic cancer and normal pancreas tissues. Survival analysis was performed to correlate IDO expression and histopathologic parameters with overall survival. The Kaplan-Meier method and Cox proportional hazards regression model were conducted. (2) We evaluated the mRNA and protein expression of IDO in PANC-1, CFPAC-1, BxPC-3 and Panc02 cell lines with or without 48 h treatment by 500 units/ml IFN-?. After being incubated with the different concentrations of 1-MT, R848 and CH223191 (0, 0.01nM, 0.1 nM, 1nM, lOnM, 100nM, respectively) for 48 hours, the Panc02 cell proliferation was assessed by MTT assay. (3) A subcutaneous model of pancreatic cancer was constructed to evaluate the therapeutic effect of combination anti-tumor therapy of 1-MT and R848. And analyze the immune cells in tumor by flow cytometry. (4) CD8+T cells were depleted by anti-CD8 antibody and the function of AHR signaling pathway was blocked by AHR inhibitor. The growth of tumor was observed subsequently. DC cells in tumor draining lymph nodes of each experimental group were also analyzed.Results: (1) Compared with normal pancreatic tissues, pancreatic adenocarcinoma showed significantly higher IDO expression levels, especially among patients with high TNM stages (P = 0.017), poor histological differentiation (P = 0.017), and lymph node metastasis (P = 0.037). Kaplan-Meier survival curves showed that high IDO expression was correlated with low survival rates (P = 0.009). Multivariate analysis using Cox proportional hazards model indicated that lymph node metastasis (HR = 2.86 P = 0.01)and IDO expression (HR = 2.38 P = 0.02) were two independent prognostic predictors of pancreatic adenocarcinoma. (2) PANC-1, CFPAC-1, and BxPC-3 cell lines expressed IDO at mRNA and protein level, and the relative expression level increased after stimulation with 500 units/ml IFN- ? for 48 hours. 1-MT, CH223191 and R848 showed no significant effect on the proliferation of Panc02 cell line. (3) The subcutaneous model of pancreatic cancer was successfully completed, and the results showed that 1-MT combined with R848 in the treatment group had better effect of inhibiting tumor growth.The use of 1-MT and R848 separately can not control the growth of tumor. And more CD8+T cells infiltrating tumor were found in 1-MT combined with R848 group compared with control group and R848 group. (4) The number of cDC cells in tumor draining lymph nodes of 1-MT combined R848 group was significantly increased. 1-MT combined R848 CD8+T cell depleting group and CH223191 combined R848 group could not control the growth of tumor.Conclusion: (1) IDO expression in pancreatic adenocarcinoma was related to prognosis of patients. IDO expression was independent prognostic predictors of pancreatic adenocarcinoma. (2) The expression of IDO in pancreatic cancer was regulated by IFN-? . (3) Combined application of 1-MT and R848 can inhibit the growth of pancreatic cancer. This effect depends on more cDC presenting tumor antigens and increases the infiltration of CD8+T cells in the tumor. Inhibition of the AHR pathway does not inhibit tumor growth. |
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