非洲猪瘟病毒K205R mRNA-LNP制备及免疫评价

doi:10.1016/j.jia.2024.03.053

Journal of Integrative Agriculture ›› 2025, Vol. 24 ›› Issue (10): 4026-4033.DOI: 10.1016/j.jia.2024.03.053

非洲猪瘟病毒K205R mRNA-LNP制备及免疫评价

收稿日期:2023-11-15 修回日期:2024-03-16 接受日期:2024-02-07 出版日期:2025-10-20 发布日期:2025-09-24

Immunogenicity and efficacy of an LNP-mRNA prepared from African swine fever virus K205R

Chuanwen Tian^1*, Yingnan Liu^1*, Dongdong Di², Zhenhua Xie¹, Yao Li¹, Rongrong Wang¹, Jie Li², Jingyi Liu^1#, Hongjun Chen^1#

¹Key Laboratory of Animal Biosafety Risk Prevention and Control (North) of Ministry of Agriculture and Rural Affairs, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China
²Spirit Jinyu Biological Pharmaceutical Co., Ltd., Hohhot 010030, China

Received:2023-11-15 Revised:2024-03-16 Accepted:2024-02-07 Online:2025-10-20 Published:2025-09-24
About author:Chuanwen Tian, E-mail: 2294875470@qq.com; Yingnan Liu, E-mail: liuyingnan@cau.edu.cn; #Correspondence Hongjun Chen, E-mail: vetchj@cau.edu.cn; Jingyi Liu, E-mail: liujingyi@shvri.ac.cn * These authors contributed equally to this study.
Supported by:
This research was funded by the National Key Research and Development Program of China (2022YFD1800500, 2021YFD1801401, and 2023YFD1802600), the Central Public-interest Scientific Institution Basal Research Fund, China (Y2022PT11), and the Shanghai Sailing Program, China (23YF1457400).

摘要/Abstract

摘要： 非洲猪瘟（Afrecian Swine Fever, ASF）是由非洲猪瘟病毒（Afrecian Swine Fever Virus, ASFV）感染引起的致死性疾病，感染ASFV的猪通常出现急性病症，表现为高热、食欲丧失、呼吸系统和神经系统功能紊乱，内脏器官广泛出血等症状，且病程短、死亡率极高。尽管ASFV一些重要蛋白的结构已经解析，但目前仍然无有效防控ASF的疫苗。ASFV K205R 基因位于病毒基因组右侧，被证实具有高抗原性，可以用于检测ASFV特异性抗体。同时，重组K205R的腺病毒载体疫苗免疫猪后能够产生极强的体液免疫水平，表明K205R具有良好的免疫原性。mRNA疫苗作为新兴的疫苗平台，具有广阔的应用价值。mRNA疫苗可以同时激活体液免疫和细胞免疫，形成双重免疫保护机制，较传统疫苗的保护作用更强。为了探究ASFV K205R mRNA的免疫原性，本研究经体外转录反应合成了K205R mRNA，并将mRNA与LNP混合，获得LNP包裹的K205R mRNA。经检测K205R mRNA-LNP的粒径大小约为86.27纳米，其封装效率为96.24%。体外转染证明其在293T和PK15细胞中能够正常表达。随后通过ELISA在小鼠和猪体内评价了K205R mRNA-LNP的免疫原性，与对照组相比，二次免疫后7天均产生了高水平的抗体，抗体效价可以达到1：512000。接下来，为了探究ASFV K205R mRNA的保护效果，我们进行了非洲猪瘟病毒攻毒实验。在第二次接种K205R mRNA后的第三周，使用5.0 HAD₅₀ ASFV-GZ株进行攻毒。对照组所有猪只在9天内全部死亡。而免疫组中，接种K205R mRNA的猪只死亡时间有所延迟，其中2头猪在第10天死亡，另外1头猪在第18天死亡。这表明K205R mRNA对ASFV有猪一定的保护作用，能够延长猪的存活时间。本研究首次评价了ASFV K205R mRNA的免疫原性，并在猪体内评估了其保护作用，为后续ASFV疫苗的研发提供参考。

Abstract:

African swine fever (ASF), caused by African swine fever virus (ASFV), is a highly contagious swine disease that has spread globally. Effective control strategies are not yet available. In this study, we prepared K205R mRNA, which was then formulated using Lipid Nanoparticle (LNP). The resulting K205R mRNA-LNP showed a particle size of approximately 86.27 nm and an mRNA encapsulation efficiency of 96.24%. Efficient expression of the K205R protein was confirmed in both HEK293T and PK15 cells. We further evaluated the immunogenicity of K205R mRNA-LNP in mice and pigs. All immunized animals developed significantly higher levels of IgG antibodies against K205R compared to the control group in the first week after the second immunization, with antibody titers reaching up to 10⁵. Challenge experiments showed that K205R mRNA delayed the time of death. Our results suggested the successful implementation of the mRNA platform in the preparation and application of ASFV mRNA.

Key words: ASFV , K205R , mRNA , immunogenicity

Chuanwen Tian, Yingnan Liu, Dongdong Di, Zhenhua Xie, Yao Li, Rongrong Wang, Jie Li, Jingyi Liu, Hongjun Chen. 非洲猪瘟病毒K205R mRNA-LNP制备及免疫评价[J]. Journal of Integrative Agriculture, 2025, 24(10): 4026-4033.

Chuanwen Tian, Yingnan Liu, Dongdong Di, Zhenhua Xie, Yao Li, Rongrong Wang, Jie Li, Jingyi Liu, Hongjun Chen. Immunogenicity and efficacy of an LNP-mRNA prepared from African swine fever virus K205R[J]. Journal of Integrative Agriculture, 2025, 24(10): 4026-4033.

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非洲猪瘟病毒K205R mRNA-LNP制备及免疫评价

Immunogenicity and efficacy of an LNP-mRNA prepared from African swine fever virus K205R

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