Intracellular Antibody Fragment Against Hepatitis B Virus X Protein Does Not Inhibit Viral Replication

Abstract

Replication of the hepatitis B virus is suppressed by deficiency of the X protein. Although several molecules that block cellular targets of X protein reduce the production of hepatitis B virus progeny, the effect of a specific inhibitor of X protein on viral replication has not been investigated. To block X protein specifically, we adopted an intracellular expression approach using H7 single chain variable fragment (H7scFv), an antibody fragment against X protein. We previously demonstrated that cytoplasmic expression of H7scFv inhibits X protein-induced tumorigenicity and transactivation. In this study, intracellular H7scFv expression inhibits reporter gene transactivation but not viral replication determined by endogenous hepatitis B virus polymerase activity assay and real-time PCR. Our findings imply that intracellular expression of antibody fragment against X protein may not be an alternative therapeutic modality for inhibition of hepatitis B virus replication.

Expression and Evaluation of Chikungunya Virus E1 and E2 Envelope Proteins for Serodiagnosis of Chikungunya Virus Infection

“…Three days post-infection, cells were harvested and whole-cell lysates were analyzed by a discontinuous sodium dedecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) system, and the gels were stained with Coomassie blue. Proteins resolved by SDS-PAGE were electrophoretically transferred onto a nitrocellulose membrane for Western blot analysis 18. The membrane was incubated in PBS containing 2% skim milk and then reacted with 1 : 100 dilution of CHIKV IgM positive serum, which was prepared by pooling 40 CHIKV IgM positive sera.…”

“…Proteins resolved by SDS-PAGE were electrophoretically transferred onto a nitrocellulose membrane for Western blot analysis. 18 The membrane was incubated in PBS containing 2% skim milk and then reacted with 1 : 100 dilution of CHIKV IgM positive serum, which was prepared by pooling 40 CHIKV IgM positive sera. After 1 hour, the membrane was washed several times and subsequently treated with horseradish peroxidase (HRP) conjugated goat anti-human IgM antibodies (Santa Cruz Biotechnology, Santa Cruz, CA, USA) at a 1 : 2,000 dilution for 1 hour at room temperature.…”

Intracellular-delivery of a single-chain antibody against hepatitis B core protein via cell-penetrating peptide inhibits hepatitis B virus replication in vitro

“…Small molecular compounds phenopropenamides (11) and dihydroarylpyrimidines, such as Bay 4-4109 (12), inhibit the maturation of HBV nucleocapsids and subsequent replication, although challenged by the potential hepatotoxicity due to fatty acid metabolism disorder and mitochondrial inability (13). Both the internal fragment of HBc (HBc78-117) (14) and the intracellular single-chain variable fragment (scFv) antibody against HBc, but not that against HBx, delivered by plasmid or lentiviral vector were all capable of inhibiting HBV replication in vitro by interfering with the function of HBc (15,16). Collectively, these data indicate the prospects of encapsidation of HBV genome as an attractive target for antiviral design, and, presumably, the scFv antibodies against HBc are preferable alternatives for this purpose, considering their well-known advantages of specificity for antiviral therapeutics.…”