Westhaus, Adrian;
Cabanes Creus, Marti;
Jonker, Timo;
Sallard, Erwan;
Navarro, Renina Gale;
Zhu, Erhua;
Baltazar, Grober;
... Lisowski, Leszek; + view all
(2022)
AAV-p40 Bioengineering Platform for Variant Selection Based on Transgene Expression.
Human Gene Therapy
, 33
(11-12)
pp. 664-682.
10.1089/hum.2021.278.
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Abstract
The power of AAV directed evolution for identifying novel vector variants with improved properties is well established, as evidenced by numerous publications reporting novel AAV variants. However, most capsid variants reported to date have been identified using either replication-competent selection platforms or PCR-based capsid DNA recovery methods, which can bias the selection towards efficient replication or unproductive intracellular trafficking, respectively. A central objective of this study was to validate a functional transduction (FT)-based method for rapid identification of novel AAV variants based on AAV capsid mRNA expression in target cells. We performed a comparison of the FT platform to existing replication competent strategies. Based on the selection kinetics and function of novel capsids identified in an in vivo screen in a xenograft model of human hepatocytes, we identified the mRNA-based FT selection as the most optimal AAV selection method. Lastly, to gain insight into the mRNA-based selection mechanism driven by the native AAV-p40 promoter, we studied its activity in a range of in vitro and in vivo targets. We found AAV-p40 to be a ubiquitously active promoter that can be modified for cell type-specific expression by incorporating binding sites for silencing transcription factors, allowing for cell-type-specific library selection.
| Type: | Article |
|---|---|
| Title: | AAV-p40 Bioengineering Platform for Variant Selection Based on Transgene Expression |
| Location: | United States |
| Open access status: | An open access version is available from UCL Discovery |
| DOI: | 10.1089/hum.2021.278 |
| Publisher version: | https://doi.org/10.1089/hum.2021.278 |
| Language: | English |
| Additional information: | © Adrian Westhaus et al. 2022; Published by Mary Ann Liebert, Inc. This Open Access article is distributed under the terms of the Creative Commons License [CC-BY] (http://creativecommons.org/licenses/by/4.0). |
| Keywords: | AAV, capsid bioengineering, novel vectors, adeno-associated viral vectors, vector development, directed evolution |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > UCL GOS Institute of Child Health > Infection, Immunity and Inflammation Dept UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > UCL GOS Institute of Child Health UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences |
| URI: | https://discovery-pp.ucl.ac.uk/id/eprint/10150382 |
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