Study On Tumor Inhibition Effect Of Radiotherapy Combined With IDO Inhibitor On Mice With Lung Cancer

Object: 1.To observe the tumor inhibition effect of radiotherapy combined with indoleamine-2,3-dioxygenase(IDO)inhibitor 1-methyltryptophan(1MT)on mice with lung cancer.2.To observe the inhibitory effect of radiotherapy combined with 1MT treatment on the immunosuppressive molecule IDO in tumor-bearing mice.3.To investigate the effects of radiotherapy combined with 1MT treatment on the mature differentiation of DCs,regulatory T cell differentiation,CD8+T cell activation,and T cell depletion receptors(PD-1,TIM3,BTLA)in tumor-bearing mice.Methods: 1.C57BL/C mice subcutaneously vaccinated Lewis lung carcinoma(LLC)cells suspension after a tumor-burdened mice model,were randomly divided into 4 groups: the control group(CON group),the 1MT group,the RT group,the combined group(1MT+RT group),were given normal saline twice a day,or 1MT 400 mg/kg lavage,the RT group and 1MT+RT group on the 13 th day after tumor single give a tumor-burdened 10 Gy X-ray irradiation in mice tumor surface,the combined group to give 1MT treatment at the same time.Tumor growth curve was drawn by observing and measuring tumor size on a daily basis.The mice were sacrificed on the 28 th day after receiving tumor,and the tumor tissue was removed and weighed.2.Some tumor tissues were taken to prepare pathological sections for HE staining,and the expression of Ki-67 in each group was observed by immunohistochemical staining.3.Expression levels of IDO,PD-1,TIM3 and BTLA in tumor tissues of each group were detected by RT-q PCR.4.Flow cytometry analyzer groups of mice was detected in spleen of DCs in mature phenotype(MHC?+,CD80+ and CD86+),CD4+CD25+Foxp3+Tregs,CD8+INF ?+ T cells express.Results: 1.The tumor growth curve showed that the tumor growth was the slowest in the 1MT+RT group,and the mean value of each group was CON group > the RT group > the 1MT group > 1MT+RT group after tumor tissue stripping was weighed on the 28 th day after inoculation(P<0.05).Compared with CON group,the tumor inhibition rate of the other three groups was the RT group 62.8% < the 1MT group 78.3% < 1MT+RT group 89.4%(P<0.05).2.Histopathological changes of the tumor: the tumor cells were arranged in diffuse sheets by HE staining,and the nuclei were varied in size,hyperchromatic,and highly heterotypic.Immunohistochemical Ki-67 positive rate: CON group(56.72±4.02)%> the RT group(21.24±3.83)% > the 1MT group(11.32±3.53)% > 1MT+RT group(8.31±2.16)%(P<0.05),1MT+RT group was 85.35%,60.88%,26.59% lower than CON group,RT group and 1MT group(P<0.05).3.RT-q PCR detection: the expression levels of IDO,PD-1,TIM3,and BTLA genes in tumor tissues were shown as follows: CON group > the RT group > the 1MT group > 1MT+RT group;The difference between CON group and the 1MT group and the 1MT+RT group was statistically significant(P < 0.05).4.Flow cytometry showed that the expression ratio of CD80+ in CD11c+ cells of spleen was CON group(56.05±1.79)% < the RT group(65.65±1.07)% < the 1MT group(72.38±1.82)% < 1MT+RT group(81.75±1.84)%.The expression rate of CD86+ in CD11c+ cells was(48.88±1.23)% < the RT group(54.9±3.36)% < the 1MT group(67.05±0.76)% < 1MT+RT group(71.23±0.58)%,and the difference between CON group and the other three groups was statistically significant(P<0.05).The proportion of MHCII+ expression in dendritic cells in each group was CON group(64.12±0.52)% < the RT group(90.23±0.45)% < the 1MT group(94.50±0.14)% < 1MT+RT group(96.78±0.43)%,and the difference between CON group and the other three groups was statistically significant(P<0.05).The proportion of CD4+CD25+Foxp3+Tregs cells in the spleen was CON group(2.58±0.05)% > the RT group(1.32±0.1)% > the 1MT group(1.02±0.1)% > 1MT+RT group(0.64±0.07)%,and the difference between CON group and the other three groups was statistically significant(P<0.05).The proportion of CD8+INF-?+ T cells in spleen was CON group(54.40±0.49)% < the RT group(61.36±1.05)% < the 1MT group(63.80±2.11)% < 1MT+RT group(69.06±0.98)%,and the difference between CON group and the other three groups was statistically significant(P<0.05).Conclusion: 1.IDO inhibitor 1MT combined with radiotherapy can effectively delay tumor growth in mice with lung cancer;2.1MT combined with radiotherapy can reduce the expression of IDO in tumor tissues of mice with lung cancer,reduce the number of peripheral Tregs cells,promote the maturation of DCs and the activation of CD8+T cells,inhibit the expression of depleted T cell receptor,and enhance the anti-tumor immune response of tumor-bearing mice.