Construction and characterization of an intracellular single-chain human antibody to hepatitis C virus non-structural 3 protein

Deborah E. Sullivan, Mario U. Mondelli, David T. Curiel, Viktor Krasnykh, Gallina Mikheeva, Paul Gaglio, Cindy B. Morris, Srikanta Dash, Michael A. Gerber

Research output: Contribution to journalArticle

20 Citations (Scopus)

Abstract

Background/Aims: We developed a single-chain antibody fragment (scFv) to the non-structural 3 protein (NS3) of hepatitis C virus (HCV) and tested its ability to interfere with the HCV replication cycle in infected hepatocytes. Methods: The variable regions of the human monoclonal antibody CM3.B6 that recognizes a conformational epitope within the helicase domain of NS3 were introduced into adenoviral vectors for expression in mammalian hepatocytes. Expression and binding properties of the scFv were analyzed by immunological assays. Effects of intracellular expression of the scFv on HCV replication were assessed in primary hepatocytes isolated from explanted livers of patients with chronic HCV infection by reverse transcription-polymerase chain reaction. Results: Transduction of HepG2 cells by the recombinant adenoviruses resulted in stable, efficient expression of scFv in the cytoplasm that was non-toxic to the cells. The scFv specifically bound to its cognate antigen. Significantly, intracellular expression of scFv resulted in a decrease in HCV genomic RNA in HCV infected hepatocytes. Conclusions: These results indicate that specific binding of a scFv to NS3 may inhibit one or more functions of this essential viral protein thus interfering with the HCV replication cycle.

Original languageEnglish
Pages (from-to)660-668
Number of pages9
JournalJournal of Hepatology
Volume37
Issue number5
DOIs
Publication statusPublished - Nov 1 2002

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Single-Chain Antibodies
Hepacivirus
Hepatocytes
Virus Replication
Proteins
Immunoglobulin Fragments
Hep G2 Cells
Viral Proteins
Chronic Hepatitis C
Virus Diseases
Protein C
Adenoviridae
Reverse Transcription
Epitopes
Cytoplasm
Monoclonal Antibodies
RNA
Antigens
Polymerase Chain Reaction
Liver

Keywords

  • Adenovirus
  • Gene therapy
  • Helicase
  • Hepatitis C virus
  • Non-structural 3 protein
  • Single-chain antibody

ASJC Scopus subject areas

  • Gastroenterology

Cite this

Construction and characterization of an intracellular single-chain human antibody to hepatitis C virus non-structural 3 protein. / Sullivan, Deborah E.; Mondelli, Mario U.; Curiel, David T.; Krasnykh, Viktor; Mikheeva, Gallina; Gaglio, Paul; Morris, Cindy B.; Dash, Srikanta; Gerber, Michael A.

In: Journal of Hepatology, Vol. 37, No. 5, 01.11.2002, p. 660-668.

Research output: Contribution to journalArticle

Sullivan, Deborah E. ; Mondelli, Mario U. ; Curiel, David T. ; Krasnykh, Viktor ; Mikheeva, Gallina ; Gaglio, Paul ; Morris, Cindy B. ; Dash, Srikanta ; Gerber, Michael A. / Construction and characterization of an intracellular single-chain human antibody to hepatitis C virus non-structural 3 protein. In: Journal of Hepatology. 2002 ; Vol. 37, No. 5. pp. 660-668.
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