Developing a potentially immunologically inert tetracycline-regulatable viral vector for gene therapy in the peripheral nerve.
Viral vector-mediated gene transfer of neurotrophic factors is an emerging and promising strategy to promote the regeneration of injured peripheral nerves. Unfortunately, the chronic exposure to neurotrophic factors results in local trapping of regenerating axons or other unwanted side effects. Therefore, tight control of therapeutic gene expression is required. The tetracycline/doxycycline-inducible system is considered to be one of the most promising systems for regulating heterologous gene expression. However, an immune response directed against the transactivator protein rtTA hampers further translational studies. Immunogenic proteins fused with the Gly-Ala repeat of the Epstein-Barr virus Nuclear Antigen-1 protein have been shown to successfully evade the immune system. In this article, we used this strategy to demonstrate that a chimeric transactivator, created by fusing the Gly-Ala repeat with rtTA and embedded in a lentiviral vector (i) retained its transactivator function in vitro, in muscle explants, and in vivo following injection into the rat peripheral nerve, (ii) exhibited a reduced leaky expression, and (iii) had an immune-evasive advantage over rtTA as shown in a novel bioassay for human antigen presentation. The current findings are an important step toward creating a clinically applicable potentially immune-evasive tetracycline-regulatable viral vector system.
1] Department of Neuroregeneration, Netherlands Institute for Neuroscience, Royal Netherlands Academy of Arts and Sciences (KNAW), Amsterdam, The Netherlands [2] Department of Neurosurgery, Leiden University Medical Center, Leiden, The Netherlands.
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Hoyng
1] Department of Neuroregeneration, Netherlands Institute for Neuroscience, Royal Netherlands Academy of Arts and Sciences (KNAW), Amsterdam, The Netherlands [2] Neuroscience Institute of the Cavalieri Ottolenghi Foundation (NICO), Department of Clinical and Biological Sciences, University of Turin, Turin, Italy.
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Gnavi
1] Department of Neuroregeneration, Netherlands Institute for Neuroscience, Royal Netherlands Academy of Arts and Sciences (KNAW), Amsterdam, The Netherlands [2] Department of Neurosurgery, Leiden University Medical Center, Leiden, The Netherlands.
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de Winter
Department of Neuroregeneration, Netherlands Institute for Neuroscience, Royal Netherlands Academy of Arts and Sciences (KNAW), Amsterdam, The Netherlands.
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Eggers
Department of Chemistry, School of Science, University of Tokyo, Tokyo, Japan.
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Ozawa
Department of Molecular Cell Biology, Leiden University Medical Center, Leiden, The Netherlands.
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Zaldumbide
Department of Molecular Cell Biology, Leiden University Medical Center, Leiden, The Netherlands.
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Hoeben
1] Department of Neuroregeneration, Netherlands Institute for Neuroscience, Royal Netherlands Academy of Arts and Sciences (KNAW), Amsterdam, The Netherlands [2] Department of Neurosurgery, Leiden University Medical Center, Leiden, The Netherlands.
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MJ
Malessy
1] Department of Neuroregeneration, Netherlands Institute for Neuroscience, Royal Netherlands Academy of Arts and Sciences (KNAW), Amsterdam, The Netherlands [2] Center for Neurogenomics and Cognitive Research, Vrije Universiteit Amsterdam, Amsterdam, The Netherlands.
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Verhaagen
D000818:Animals; D001483:Base Sequence; D005260:Female; D005786:Gene Expression Regulation; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors / Q000235:genetics / Q000276:immunology / Q000494:pharmacology*; D057809:HEK293 Cells; D006801:Humans; D066298:In Vitro Techniques; D016086:Lentivirus / Q000235:genetics; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000502:physiology; D010525:Peripheral Nerves / Q000187:drug effects*; D017208:Rats, Wistar; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology; D013752:Tetracycline / Q000494:pharmacology*; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism
D015534:Trans-Activators; D013752:Tetracycline