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A cell-penetrating whole molecule antibody targeting intracellular HBx suppresses hepatitis B virus via TRIM21-dependent pathway.

ABSTRACT: Rationale: Monoclonal antibodies (mAbs) mostly targeting extracellular or cell surface molecules have been widely used in the treatment of various diseases. However, mAbs cannot pass through the cell membrane as efficiently as small compounds, thus limiting their use against intracellular targets. Methods to shuttle antibodies into living cells may largely expand research and application in areas based on mAbs. Hepatitis B virus X protein (HBx) is an important intracellular multi-functional viral protein in the life cycle of hepatitis B virus (HBV). HBx plays essential roles in virus infection and replication and is strongly associated with HBV-related carcinogenesis. Methods: In this study, we developed a cell-penetrating whole molecule antibody targeting HBx (9D11-Tat) by the fusion of a cell penetrating peptide (CPP) on the C-terminus of the heavy chain of a potent mAb specific to HBx (9D11). The anti-HBV effect and mechanism of 9D11-Tat were investigated in cell and mouse models mimicking chronic HBV infection. Results: Our results demonstrated that the recombinant 9D11-Tat antibody could efficiently internalize into living cells and significantly suppress viral transcription, replication, and protein production both in vitro and in vivo. Further analyses suggested the internalized 9D11-Tat antibody could greatly reduce intracellular HBx via Fc binding receptor TRIM21-mediated protein degradation. This process simultaneously stimulated the activations of NF-?B, AP-1, and IFN-?, which promoted an antiviral state of the host cell. Conclusion: In summary, our study offers a new approach to target intracellular pathogenesis-related protein by engineered cell-penetrating mAb expanding their potential for therapeutic applications. Moreover, the 9D11-Tat antibody may provide a novel therapeutic agent against human chronic HBV infection.

SUBMITTER: Zhang JF

PROVIDER: S-EPMC5743566 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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A cell-penetrating whole molecule antibody targeting intracellular HBx suppresses hepatitis B virus via TRIM21-dependent pathway.

Zhang Jun-Fang JF Xiong Hua-Long HL Cao Jia-Li JL Wang Shao-Juan SJ Guo Xue-Ran XR Lin Bi-Yun BY Zhang Ying Y Zhao Jing-Hua JH Wang Ying-Bin YB Zhang Tian-Ying TY Yuan Quan Q Zhang Jun J Xia Ning-Shao NS

Theranostics 20180101 2

<b>Rationale:</b> Monoclonal antibodies (mAbs) mostly targeting extracellular or cell surface molecules have been widely used in the treatment of various diseases. However, mAbs cannot pass through the cell membrane as efficiently as small compounds, thus limiting their use against intracellular targets. Methods to shuttle antibodies into living cells may largely expand research and application in areas based on mAbs. Hepatitis B virus X protein (HBx) is an important intracellular multi-function ...[more]

PMID: 29290826

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Project description:ObjectivesHepatitis B virus X (HBx) is closely associated with HBV-related hepatocarcinogenesis via the inactivation of tumour suppressors. Protein phosphatase 2A (PP2A) regulatory subunit B56 gamma (B56γ), as a tumour suppressor, plays a critical role in regulating cellular phosphorylation signals via dephosphorylation of signalling proteins. However, the underlying mechanism that B56γ involved in regulating HBx-associated hepatocarcinogenesis phenotypes and mediating anti-HBx antibody-mediated tumour suppression remains unknown.Materials and methodsWe used bioinformatics analysis, paired HCC patient specimens, HBx transgenic (HBx-Tg) mice, xenograft nude mice, HBV stable replication in the HepG2.2.15 cells, and anti-HBx antibody intervention to systematically evaluate the biological function of protein kinase B (AKT) dephosphorylation through B56γ in HBx-associated hepatocarcinogenesis.ResultsBioinformatics analysis revealed that AKT, matrix metalloproteinase 2 (MMP2), and MMP9 were markedly upregulated, while cell migration and viral carcinogenesis pathways were activated in HBV-infected liver tissues and HBV-associated HCC tissues. Our results demonstrated that HBx-expression promotes AKT phosphorylation (p-AKTThr308/Ser473 ), mediating the migration and invasion phenotypes in vivo and in vitro. Importantly, in clinical samples, HBx and B56γ were downregulated in HBV-associated HCC tumour tissues compared with peritumor tissues. Moreover, intervention with site-directed mutagenesis (AKTT308A , AKTS473A ) of p-AKTThr308/Ser473 mimics dephosphorylation, genetics-based B56γ overexpression, and intracellular anti-HBx antibody inhibited cell growth, migration, and invasion in HBx-expressing HCC cells.ConclusionsOur results demonstrated that B56γ inhibited HBV/HBx-dependent hepatocarcinogenesis by regulating the dephosphorylation of p-AKTThr308/Ser473 in HCC cells. The intracellular anti-HBx antibody and the activator of B56γ may provide a multipattern chemopreventive strategy against HBV-related HCC.

Dataset Information

A cell-penetrating whole molecule antibody targeting intracellular HBx suppresses hepatitis B virus via TRIM21-dependent pathway.

Publications

A cell-penetrating whole molecule antibody targeting intracellular HBx suppresses hepatitis B virus via TRIM21-dependent pathway.

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