Refined methods for propagating vesicular stomatitis virus vectors that are defective for G protein expression
| Autor: | Christopher L. Parks, Susan E. Witko, J. Erik Johnson, R. Michael Hendry, George N. Pavlakis, Stephen A. Udem, Narender K. Kalyan, Barbara K. Felber, Maninder K. Sidhu |
|---|---|
| Rok vydání: | 2009 |
| Předmět: |
Virus Cultivation
G protein viruses Genetic Vectors Molecular Sequence Data gag Gene Products Human Immunodeficiency Virus Epitope Virus Article Viral Envelope Proteins Virology Chlorocebus aethiops Animals Mononegavirales Gene Vero Cells Membrane Glycoproteins biology Sequence Analysis DNA Vesiculovirus Rhabdoviridae biology.organism_classification Molecular biology Vesicular stomatitis virus Vero cell RNA Viral |
| Zdroj: | Journal of virological methods. 164(1-2) |
| ISSN: | 1879-0984 |
| Popis: | Propagation-defective vesicular stomatitis virus (VSV) vectors that encode a truncated G protein (VSV-Gstem) or lack the G gene entirely (VSV-DeltaG) are attractive vaccine vectors because they are immunogenic, cannot replicate and spread after vaccination, and do not express many of the epitopes that elicit neutralizing anti-VSV immunity. To consider advancing non-propagating VSV vectors towards clinical assessment, scalable technology that is compliant with human vaccine manufacturing must be developed to produce clinical trial material. Accordingly, two propagation methods were developed for VSV-Gstem and VSV-DeltaG vectors encoding HIV gag that have the potential to support large-scale production. One method is based on transient expression of G protein after electroporating plasmid DNA into Vero cells and the second is based on a stable Vero cell line that contains a G gene controlled by a heat shock-inducible transcription unit. Both methods reproducibly supported production of 1 x 10(7) to 1 x 10(8) infectious units (I.U.s) of vaccine vector per milliliter. Results from these studies also showed that optimization of the G gene is necessary for abundant G protein expression from electroporated plasmid DNA or from DNA integrated in the genome of a stable cell line, and that the titers of VSV-Gstem vectors generally exceeded VSV-DeltaG. |
| Databáze: | OpenAIRE |
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