结核分枝杆菌相关抗原在基于减毒单增李斯特菌为载体的宫颈癌免疫治疗中的增强效果  

Enhanced Effect of Mycobacterium tuberculosis-Related Antigens in Cervical Cancer Immunotherapy Based on Attenuated Listeria monocytogenes

作　　者：刘晨 宋亚雯 蒋昕 孙静 江玲丽[2] 宋厚辉 程昌勇 LIU Chen;SONG Ya-Wen;JIANG Xin;SUN Jing;JIANG Ling-Li;SONG Hou-Hui;CHENG Chang-Yong(College of Animal Science and Technology/College of Veterinary Medicine/Key Laboratory of Applied Technology on Green-Eco-Healthy Animal Husbandry of Zhejiang Province/Zhejiang Provincial Engineering Research Center for Animal Health Diagnostics/Advanced Technology,Zhejiang International Science and Technology Cooperation Base for Veterinary Medicine and Health Management/China-Australia Joint Laboratory for Animal Health Big Data Analytics,Zhejiang A&F University,Hangzhou 311300,China;NingBo College of Health Sciences,Ningbo 315100,China)

机构地区：[1]浙江农林大学动物科技学院/动物医学院/浙江省畜禽绿色生态健康养殖应用技术研究重点实验室/动物健康互联网检测技术浙江省工程研究中心/浙江省动物医学与健康管理国际科技合作基地/中澳动物健康大数据分析联合实验室,杭州311300 [2]宁波卫生职业技术学院,宁波315100

出　　处：《农业生物技术学报》2023年第11期2367-2376,共10页Journal of Agricultural Biotechnology

基　　金：国家自然科学基金(31972648;32172849);浙江省自然科学基金(LY23C180002);宁波市公益类科技项目(2022S006);浙江省属高校基本科研业务费专项资金(2020KJ004)。

摘　　要：单增李斯特菌(Listeria monocytogenes)可诱导机体产生强烈的免疫反应,作为递呈载体在各种癌症和传染病免疫治疗中的研究越来越多,是一种生物安全性高的肿瘤免疫治疗疫苗研发载体。本研究在实验室前期减毒单增李斯特菌LM基础上,构建与李斯特菌溶血素O(Listeriolysin O,LLO)融合表达结核分枝杆菌(Mycobacterium tuberculosis)抗原Ag85B(Antigen 85B)和ESAT6(6 kD early secreted antigen target)的肿瘤疫苗菌株LM-Ag85B和LM-ESAT6;通过Western blot确定抗原能与LLO融合表达和正常分泌;利用细菌感染生物学方法研究LM-Ag85B和LM-ESAT6的体外生长能力及溶血活性;利用小鼠(Mus musculus)宫颈癌模型评价LM-Ag85B和LM-ESAT6与LM-E7(表达宫颈癌相关抗原E7)联合应用在小鼠宫颈癌模型中的治疗效果。结果显示,LM-Ag85B和LM-ESAT6能分别正常表达分泌携带的Ag85B和ESAT6抗原;LM-Ag85B、LM-ESAT6在体外生长能力与野生型无显著差异,溶血活性低于野生型;在小鼠宫颈癌治疗模型中,LM-Ag85B、LM-ESAT6和LM-E7联合应用试验组较对照组显著抑制肿瘤生长,治疗效果良好。本研究证实了结核分枝杆菌抗原能够增强抗肿瘤效果,为推动基于减毒细菌为载体的肿瘤免疫治疗研究提供了科学依据。Listeria monocytogenes has been increasingly studied as a delivery vector in various cancer and infectious disease immunotherapies,as it can induce robust immune responses in the host.In this study,the tumor vaccine strains,LM-Ag85B and LM-ESAT6 were constructed by fusing the Mycobacterium tuberculosis antigens Ag85B(Antigen 85B)and ESAT6(6 kD early secreted antigen target)with the virulence factor listeriolysin O(LLO),which served as the basis in the preliminary laboratory experiments.The Western blot was employed to confirm the fusion expression and normal secretion of antigens with LLO.The in vitro growth capability and hemolytic activity of LM-Ag85B and LM-ESAT6 were investigated using bacterial infection biology methods.The therapeutic effects of combined application of LM-Ag85B and LM-ESAT6 with LM-E7(expressing cervical cancer-related antigen E7)were evaluated in a mouse model of cervical cancer.The results revealed that LM-Ag85B and LM-ESAT6 were able to express and secrete the carried Ag85B and ESAT6 antigens,respectively.The in vitro growth capability of LM-Ag85B and LM-ESAT6 showed no significant difference compared to the wild-type strain,while their hemolytic activity was lower than that of the wild-type strain.In the mouse model of cervical cancer treatment,the experimental groups receiving combined application of LM-Ag85B,LM-ESAT6,and LM-E7 exhibited significant inhibition of tumor growth compared to the control group,indicating favorable therapeutic effects.This study demonstrates that the Mycobacterium tuberculosis antigens can provide enhanced tumor therapeutic efficacy for the LM-E7 based cervical cancer,which significantly promotes future applications of bacterial-based tumor immunotherapy.

关 键 词：减毒单增李斯特菌 结核分枝杆菌相关抗原 宫颈癌 疫苗载体 

分 类 号：S851[农业科学—预防兽医学]