Development of inducible EIAV-based lentiviral vector packaging and producer cell lines
| Autor: | Pippa A Radcliffe, K A Mitrophanous, M A Leroux-Carlucci, C J M Sion, Hannah J Stewart |
|---|---|
| Rok vydání: | 2009 |
| Předmět: |
viruses
Genetic enhancement Blotting Western Genetic Vectors Cell Culture Techniques Biology Transfection Virus Replication Genomic Instability Viral vector Cell Line chemistry.chemical_compound Open Reading Frames Viral Envelope Proteins Genetics Animals TetR Molecular Biology Gene Membrane Glycoproteins Virus Assembly Lentivirus RNA-Directed DNA Polymerase Genetic Therapy biology.organism_classification Virology Anti-Bacterial Agents Clone Cells Fusion Proteins gag-pol Repressor Proteins chemistry Viral replication Gene Expression Regulation Vesicular stomatitis virus Doxycycline Molecular Medicine Infectious Anemia Virus Equine Plasmids |
| Zdroj: | Gene therapy. 16(6) |
| ISSN: | 1476-5462 |
| Popis: | Large-scale production of gene therapeutics comprising equine infectious anaemia virus (EIAV) -based lentiviral vectors (LVs) would benefit from the development of producer cell lines enabling the generation of larger quantities of vector than achievable by transient systems. Such cell lines would contain three vector components (Gag/Pol, VSV-G envelope and genome expression constructs). As the vesicular stomatitis virus (VSV-G) envelope protein is cytotoxic, its expression must be regulated. It is also desirable to regulate Gag/Pol expression to minimise metabolic burden on the cell. The Tet repressor (TetR) system was selected to regulate expression of VSV-G and Gag/Pol, necessitating the introduction of a fourth construct, encoding TetR, into the cell line. We have generated an inducible packaging cell line that shows tight control of the packaging components, and high-titre vector production on transient transfection of the EIAV genome. The cell line is stable for at least 7 weeks in the absence of selective pressure. To verify that this packaging cell line can support the generation of producer cell lines it was transfected stably with an EIAV genome cassette encoding ProSavin; a gene therapeutic for Parkinson's disease. Producer cell lines were generated, which on induction, yielded ProSavin with titres comparable to the transient system. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|