High-Level rAAV Vector Production by rAdV-Mediated Amplification of Small Amounts of Input Vector
The successful application of recombinant adeno-associated virus (rAAV) vectors for long-term transgene expression in clinical studies requires scalable production methods with genetically stable components. Due to their simple production scheme and the high viral titers achievable, first generation...
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MDPI AG
2022-12-01
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Series: | Viruses |
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Online Access: | https://www.mdpi.com/1999-4915/15/1/64 |
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author | Stefan Weger |
author_facet | Stefan Weger |
author_sort | Stefan Weger |
collection | DOAJ |
description | The successful application of recombinant adeno-associated virus (rAAV) vectors for long-term transgene expression in clinical studies requires scalable production methods with genetically stable components. Due to their simple production scheme and the high viral titers achievable, first generation recombinant adenoviruses (rAdV) have long been taken into consideration as suitable tools for simultaneously providing both the helper functions and the AAV <i>rep</i> and <i>cap</i> genes for rAAV packaging. So far, however, such rAdV-<i>rep/cap</i> vectors have been difficult to generate and often turned out to be genetically unstable. Through ablation of <i>cis</i> and <i>trans</i> inhibitory function in the AAV-2 genome we have succeeded in establishing separate and stable rAdVs for high-level AAV serotype 2 Rep and Cap expression. These allowed rAAV-2 production at high burst sizes by simple coinfection protocols after providing the AAV-ITR flanked transgene vector genome either as rAAV-2 particles at low input concentrations or in form of an additional rAdV. With characteristics such as the ease of producing the required components, the straightforward adaption to other transgenes and the possible extension to further serotypes or capsid variants, especially the rAdV-mediated rAAV amplification system presents a very promising candidate for up-scaling to clinical grade vector preparations. |
first_indexed | 2024-03-09T11:03:44Z |
format | Article |
id | doaj.art-f9328b85c77843f483ba0ad162cd8a6a |
institution | Directory Open Access Journal |
issn | 1999-4915 |
language | English |
last_indexed | 2024-03-09T11:03:44Z |
publishDate | 2022-12-01 |
publisher | MDPI AG |
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series | Viruses |
spelling | doaj.art-f9328b85c77843f483ba0ad162cd8a6a2023-12-01T01:07:43ZengMDPI AGViruses1999-49152022-12-011516410.3390/v15010064High-Level rAAV Vector Production by rAdV-Mediated Amplification of Small Amounts of Input VectorStefan Weger0Charité–Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Clinic for Neurology with Experimental Neurology, Gene Therapy Group, Campus Benjamin Franklin, Hindenburgdamm27, 12203 Berlin, GermanyThe successful application of recombinant adeno-associated virus (rAAV) vectors for long-term transgene expression in clinical studies requires scalable production methods with genetically stable components. Due to their simple production scheme and the high viral titers achievable, first generation recombinant adenoviruses (rAdV) have long been taken into consideration as suitable tools for simultaneously providing both the helper functions and the AAV <i>rep</i> and <i>cap</i> genes for rAAV packaging. So far, however, such rAdV-<i>rep/cap</i> vectors have been difficult to generate and often turned out to be genetically unstable. Through ablation of <i>cis</i> and <i>trans</i> inhibitory function in the AAV-2 genome we have succeeded in establishing separate and stable rAdVs for high-level AAV serotype 2 Rep and Cap expression. These allowed rAAV-2 production at high burst sizes by simple coinfection protocols after providing the AAV-ITR flanked transgene vector genome either as rAAV-2 particles at low input concentrations or in form of an additional rAdV. With characteristics such as the ease of producing the required components, the straightforward adaption to other transgenes and the possible extension to further serotypes or capsid variants, especially the rAdV-mediated rAAV amplification system presents a very promising candidate for up-scaling to clinical grade vector preparations.https://www.mdpi.com/1999-4915/15/1/64rAAV packagingadeno-associated virusrecombinant adenovirusRIS-Ad |
spellingShingle | Stefan Weger High-Level rAAV Vector Production by rAdV-Mediated Amplification of Small Amounts of Input Vector Viruses rAAV packaging adeno-associated virus recombinant adenovirus RIS-Ad |
title | High-Level rAAV Vector Production by rAdV-Mediated Amplification of Small Amounts of Input Vector |
title_full | High-Level rAAV Vector Production by rAdV-Mediated Amplification of Small Amounts of Input Vector |
title_fullStr | High-Level rAAV Vector Production by rAdV-Mediated Amplification of Small Amounts of Input Vector |
title_full_unstemmed | High-Level rAAV Vector Production by rAdV-Mediated Amplification of Small Amounts of Input Vector |
title_short | High-Level rAAV Vector Production by rAdV-Mediated Amplification of Small Amounts of Input Vector |
title_sort | high level raav vector production by radv mediated amplification of small amounts of input vector |
topic | rAAV packaging adeno-associated virus recombinant adenovirus RIS-Ad |
url | https://www.mdpi.com/1999-4915/15/1/64 |
work_keys_str_mv | AT stefanweger highlevelraavvectorproductionbyradvmediatedamplificationofsmallamountsofinputvector |