抗乙型肝炎病毒X蛋白的广谱反应性单克隆抗体的特征与工程

StephenW

Sci Rep. 2019 Dec 30;9(1):20323. doi: 10.1038/s41598-019-56819-8.
Characterization and engineering of broadly reactive monoclonal antibody against hepatitis B virus X protein that blocks its interaction with DDB1.
Tao S1, Pan S1,2, Gu C1, Wei L1, Kang N1, Xie Y3, Liu J4,5.
Author information

1
    Key Laboratory of Medical Molecular Virology (MOE/NHC/CAMS), Department of Microbiology and Parasitology, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai, China.
2
    National Clinical Research Center for Aging and Medicine, Huashan Hospital, Fudan University, Shanghai, China.
3
    Key Laboratory of Medical Molecular Virology (MOE/NHC/CAMS), Department of Microbiology and Parasitology, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai, China. [email protected].
4
    Key Laboratory of Medical Molecular Virology (MOE/NHC/CAMS), Department of Microbiology and Parasitology, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai, China. [email protected].
5
    Key Laboratory of Medical Epigenetics and Metabolism, Institutes of Biomedical Sciences, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai, China. [email protected].

Abstract

Hepatitis B virus (HBV) X protein (HBx) plays diverse roles in both viral life cycle and HBV-related carcinogenesis. Its interaction with DNA damage-binding protein 1 (DDB1) was shown to be essential for engendering cellular conditions favorable for optimal viral transcription and replication. Previously, we described a mouse monoclonal antibody against HBx (anti-HBx 2A7) recognizing HBx encoded by representative strains from 7 of 8 known HBV genotypes. In this work, we further characterized 2A7 in order to explore its potential usefulness in HBx-targeting applications. We demonstrated that 2A7 recognizes a linear epitope mapped to L89PKVLHKR96 on HBx, a segment that is highly conserved across genotypes and coincidentally overlaps with the DDB1-interacting segment. HBx-DDB1 binding could be inhibited by 2A7 in vitro, suggesting therapeutic potential. Nucleic acid and amino acid sequences of 2A7 were then obtained, which allowed construction of recombinant antibody and single chain variable fragments (scFv). 2A7-derived recombinant antibody and scFv recapitulate 2A7's HBx-binding capacity and epitope specificity. We also reported preliminary results using cell-penetrating peptide for delivering 2A7 antibody across cell membrane to target intracellular HBx. Anti-HBx 2A7 and 2A7-derived scFv characterized here may give rise to novel HBx-targeting diagnostics and therapeutics for HBV- and HBx-related pathologies.

PMID:
    31889135
DOI:
    10.1038/s41598-019-56819-8

StephenW

Sci Rep.2019年12月30日; 9（1）：20323。 doi：10.1038 / s41598-019-56819-8。
抗乙型肝炎病毒X蛋白的广谱反应性单克隆抗体的特征与工程化，可阻断其与DDB1的相互作用。
陶S1，潘S1,2，顾C1，韦L1，康N1，谢Y3，刘J4,5。
作者信息

1个
    复旦大学上海医学院基础医学院，医学分子病毒学重点实验室（MOE / NHC / CAMS），微生物学和寄生虫学系，上海
2
    复旦大学附属华山医院国家老龄医学临床研究中心，上海
3
    复旦大学上海医学院基础医学院，医学分子病毒学重点实验室（MOE / NHC / CAMS），微生物学和寄生虫学系，上海[email protected]。
4
    复旦大学上海医学院基础医学院，医学分子病毒学重点实验室（MOE / NHC / CAMS），微生物学和寄生虫学系，上海[email protected]。
5
    复旦大学上海医学院基础医学学院，生物医学科学研究所，医学表观遗传与代谢重点实验室，上海[email protected]。

抽象

乙型肝炎病毒（HBV）X蛋白（HBx）在病毒生命周期和HBV相关的癌变过程中均发挥着不同的作用。它与DNA损伤结合蛋白1（DDB1）的相互作用被证明对于产生有利于最佳病毒转录和复制的细胞条件至关重要。以前，我们描述了一种针对HBx的小鼠单克隆抗体（抗HBx 2A7），该抗体识别由8种已知HBV基因型中的7种代表株编码的HBx。在这项工作中，我们进一步对2A7进行了表征，以探讨其在HBx靶向应用中的潜在用途。我们证明2A7识别映射到HBx上的L89PKVLHKR96的线性表位，该段在基因型之间高度保守，并且与DDB1相互作用段重叠。 HBx-DDB1结合可在体外被2A7抑制，表明具有治疗潜力。然后获得2A7的核酸和氨基酸序列，其允许构建重组抗体和单链可变片段（scFv）。 2A7衍生的重组抗体和scFv概括了2A7的HBx结合能力和表位特异性。我们还报告了使用细胞穿透肽跨细胞膜递送2A7抗体靶向细胞内HBx的初步结果。此处表征的抗HBx 2A7和2A7衍生的scFv可能会引起针对HBV和HBx相关病理的新型HBx靶向诊断和治疗。

PMID：
    31889135
DOI：
    10.1038 / s41598-019-56819-8

StephenW

https://www.nature.com/articles/s41598-019-56819-8.pdf