抑瘤肠菌联合抗程序性细胞死亡蛋白1单抗治疗微卫星稳定型结直肠癌的实验研究

Experimental study of tumor-suppressing enterobacteria combined with anti-programmed cell death protein-1 monoclonal antibody therapy microsatellite stable colorectal cancer

目的 探讨抑瘤肠菌(TSE)与抗程序性细胞死亡蛋白1(PD-1)单抗联合治疗对微卫星稳定(MSS)型结直肠癌(CRC)疗效的影响。方法 选取BALB/c小鼠构建CT26荷瘤小鼠模型后,随机分为对照组、TSE单药组(TSE组)、αPD-1单药组(αPD-1组)、TSE联合αPD-1组(联合组),每组10只。定期测量小鼠皮下移植瘤体积并监测体重,给药14天后处死获取皮下移植瘤,比较各组肿瘤体积大小;采用苏木精-伊红(HE)染色观察肿瘤细胞的形态结构;采用免疫组化法检测肿瘤组织CD8^(+)T细胞数量;采用流式细胞术检测皮下移植瘤组织和脾脏组织中CD4^(+)T细胞和CD8^(+)T细胞占CD45^(+)细胞的比例。结果 对照组、TSE组、αPD-1组及联合组小鼠肿瘤平均体积依次减小(P<0.05)。免疫组化结果显示,对照组、TSE组、αPD-1组及联合组小鼠肿瘤组织中CD8^(+)T细胞数量依次升高(P<0.001)。肿瘤组织流式细胞术结果显示,αPD-1组小鼠CD3^(+)CD8^(+)T细胞占CD45^(+)细胞的比例高于对照组,联合组小鼠CD3^(+)CD8^(+)T细胞占CD45^(+)细胞的比例高于对照组、TSE组及αPD-1组(P<0.05)。结论 TSE联合抗PD-1单抗可明显提高免疫单药治疗的抑瘤作用,可能通过增加肿瘤组织中浸润CD8^(+)T细胞而促进抗PD-1单抗治疗CRC的疗效。

Objective To investigate the effect of tumor-suppressing enterobacteria(TSE) combined treatment with anti-programmed cell death protein-1(PD-1) monoclonal antibody on the efficacy of microsatellite stable(MSS) colorectal cancer(CRC).Methods BALB/c mice were selected to construct CT26 tumor-bearing mouse models and randomly divided into control group,TSE single drug group(TSE group),αPD-1 single drug group(αPD-1 group),TSE combined αPD-1 group(combination group),10 in each group.The subcutaneous tumor volume of the mice were measured regularly and the body weight were monitored.After 14 days of administration,the mice were killed and subcutaneous tumors were obtained,and the tumor volume of each group was compared.Hematoxylin-eosin(HE) staining was used to observe the morphological structure of tumor cells.The number of CD8~+ T cells in tumor tissues was investigated by immunohistochemistry.The proportion of CD4~+T cells and CD8~+T cells in CD45~+ cells in tumor tissues and spleen tissues were detected by flow cytometry.Results Average tumor volume of control group,TSE group,αPD-1 group and combination group was gradually decreased in turn(P<0.05).Immunohistochemical results showed that the number of CD8~+T cells in tumor tissue of control group,TSE group,αPD-1 group and combination group increased in turn(P<0.001).The flow cytometry results of tumor tissue showed that the proportion of CD3~+CD8~+T cells in CD45~+ cells of αPD-1 group was higher than that in control group,and the proportion of CD3~+CD8~+T cells in CD45~+ cells of combination group was higher than that in control group,TSE group and αPD-1 group(P<0.05).Conclusion TSE combined with anti-PD-1 monoclonal antibody can significantly enhance the anti-tumor effect of single immune drug therapy,and may promote the efficacy of anti-PD-1 monoclonal antibody in the treatment of CRC by increasing the infiltration of CD8~+ T cells in tumor tissues.