Inhibition of hepatitis C virus NS3-mediated cell transformation by recombinant intracellular antibodies.
OBJECTIVE
Hepatitis C virus (HCV) infection is a major worldwide health problem, causing chronic hepatitis, liver cirrhosis and primary liver cancer. In addition to its role in the viral polyprotein-processing, the viral NS3 serine protease has been implicated in interactions with various cell constituents resulting in phenotypic changes including malignant transformation. NS3 is currently regarded a prime target for anti-viral drugs thus specific inhibitors of its activities should be of importance. With the aim of inhibiting NS3-mediated cell transformation, we used antibody-phage display to isolate NS3-specific single-chain antibodies (scFv).


METHODS
We have isolated and characterized eight anti-NS3 scFvs. We investigated the phenotypic changes that NS3-expressing cells undergo upon intracellular expression of these antibodies in different subcellular compartments (intracellular immunization), assayed by their proliferation rate and their ability to grow anchorage independently. The intracellular location of NS3 and the scFvs were analyzed by immunofluorescent staining using confocal microscopy.


RESULTS
Nuclear targeted anti-NS3 intrabodies shuttle NS3 from the cytosol to the nucleus with concomitant inhibition of cell proliferation and loss of the transformed phenotype.


CONCLUSIONS
Intracellular immunization-based gene therapy strategies may emerge as a promising antiviral approach to interfere with the life cycle and tumorigenicity of HCV.
Zemel
Romy
R
Molecular Hepatology Research Laboratory, Felsenstein Medical Research Center, Sackler School of Medicine, Tel-Aviv University, Petah Tikva, Israel.
Berdichevsky
Yevgeny
Y
Bachmatov
Larisa
L
Benhar
Itai
I
Tur-Kaspa
Ran
R
D000595:Amino Acid Sequence; D000818:Animals; D002472:Cell Transformation, Viral; D002478:Cells, Cultured; D003001:Cloning, Molecular; D016174:Hepacivirus / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D008969:Molecular Sequence Data; D010957:Plasmids; D016133:Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000235:genetics*; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
D017361:Viral Nonstructural Proteins; C084422:NS3 protein, hepatitis C virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases