Inhibition of hepatitis C virus NS3-mediated cell transformation by recombinant intracellular antibodies

Zemel, Romy; Berdichevsky, Yevgeny; Bachmatov, Larisa; Benhar, Itai; Tur–Kaspa, Ran

doi:10.1016/j.jhep.2004.02.026

Cited by 10 publications

(16 citation statements)

References 40 publications

(48 reference statements)

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“…29,36 However, we found that the maltose-binding protein (MBP) fusion technology that we developed for efficient cytoplasmic expression of scFvs 40 could be adopted to improve the expression of scNS3 without compromising its activity (our unpublished results). Thus, the current study was carried out using MBP-scNS3.…”

Section: Genetic Screen For Protease Inhibitorsmentioning

confidence: 99%

“…All 25 clones regained their blue phenotype, confirming that the phenotypic change was indeed a direct result of scFvs expression. A sample of four of the clones (10, 11, 35, and 171) is shown in Figure 1(e) in comparison to four control scFvs, three of which (37Y, 44Y, and 53Y) that were isolated from a human synthetic antibody library, 36 bind NS3 without inhibiting it and one (Anti Tac) totally irrelevant. 48 Characterization of specificity and binding affinity of selected scFvs…”

Section: Genetic Screen For Protease Inhibitorsmentioning

confidence: 99%

“…As a control, we used the non-inhibitory, NS3 binding scFv 53Y. 36 As shown in Figure 2(a), seven of the isolated scFvs bound scNS3 specifically. Although MBP-scNS3 was used for immunization and scFv identification, the binding studies that were carried out with unfused scNS3 confirm that MBP did not play a role in their antigen binding.…”

Section: Genetic Screen For Protease Inhibitorsmentioning

confidence: 99%

“…Unfortunately, these scFvs were non-inhibitory and upon their expression as intracellular antibodies (intrabodies) in NS3-expressing cells they only partially inhibited cell transformation by diverting NS3 from the cell cytoplasm to the nucleus. 36 All the NS3-inhibiting antibodies isolated thus far have been isolated by virtue of NS3 binding and subsequent testing for inhibition of catalysis. We opted for an approach that would facilitate the direct isolation of NS3-inhibitory scFvs by virtue of their inhibition of catalysis.…”

Section: Introductionmentioning

confidence: 99%

“…Recently, we reported that the expression of NS3 in the cytoplasm of rat fibroblasts enhances cell proliferation as evident from increased incorporation of [ 3 H]thymidine. 36 We applied this system as a model for preliminary evaluation of the ability of our scFvs to perform as NS3 inhibitors when expressed as cytoplasmic intrabodies. We show that the three scFvs we tested effectively neutralize NS3-mediated enhancement of cell proliferation.…”

Section: Introductionmentioning

confidence: 99%

See 4 more Smart Citations

HCV NS3 Serine Protease-neutralizing Single-chain Antibodies Isolated by a Novel Genetic Screen

Gal-Tanamy

Zemel

Berdichevsky

et al. 2005

Journal of Molecular Biology

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Section: Genetic Screen For Protease Inhibitorsmentioning

confidence: 99%

Section: Genetic Screen For Protease Inhibitorsmentioning

confidence: 99%

Section: Genetic Screen For Protease Inhibitorsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

HCV NS3 Serine Protease-neutralizing Single-chain Antibodies Isolated by a Novel Genetic Screen

Gal-Tanamy

Zemel

Berdichevsky

et al. 2005

Journal of Molecular Biology

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Inhibition of hepatitis C virus RNA replicons by peptide aptamers

Trahtenherts¹,

Gal-Tanamy²,

Zemel³

et al. 2008

Antiviral Research

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Inhibition of protease-inhibitor-resistant hepatitis C virus replicons and infectious virus by intracellular intrabodies

et al. 2010

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Hepatitis C virus (HCV) infection is a common cause of chronic liver disease and a serious threat to human health. The HCV NS3/4A serine protease is necessary for viral replication and innate immune evasion, and represents a well-validated target for specific antiviral therapy. We previously reported the isolation of single-chain antibodies (scFvs) that inhibit NS3/4A protease activity in vitro. Expressed intracellularly (intrabodies), these scFvs blocked NS3-mediated proliferation of NS3-transfected cells. Here we show that anti-NS3 scFvs suppress HCV RNA replication when expressed intracellularly in Huh7 hepatoma cells bearing either subgenomic or genome-length HCV RNA replicons. The expression of intrabodies directed against NS3 inhibited the autonomous amplification of HCV replicons resistant to small molecule inhibitors of the NS3/4A protease, and replicons derived from different HCV genotypes. The combination of intrabodies and interferon-α had an additive inhibitory effect on RNA replication in the replicon model. Intrabody expression also inhibited production of infectious HCV in a cell culture system. The NS3 protease activity was inhibited by the intrabodies in NS3-expressing cells. In contrast, cell-free synthesis of HCV RNA by preformed replicase complexes was not inhibited by intrabodies, suggesting that the major mode of inhibition of viral replication is inhibition of NS3/4A protease activity and subsequent suppression of viral polyprotein processing.

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Inhibition of hepatitis C virus NS3-mediated cell transformation by recombinant intracellular antibodies

Cited by 10 publications

References 40 publications

HCV NS3 Serine Protease-neutralizing Single-chain Antibodies Isolated by a Novel Genetic Screen

HCV NS3 Serine Protease-neutralizing Single-chain Antibodies Isolated by a Novel Genetic Screen

Inhibition of hepatitis C virus RNA replicons by peptide aptamers

Inhibition of protease-inhibitor-resistant hepatitis C virus replicons and infectious virus by intracellular intrabodies

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