Development of an Inducible, Replication-Competent Assay Cell Line for Titration of Infectious Recombinant Adeno-Associated Virus Vectors

Zion Lee, Min Lu, Eesha A Irfanullah, Morgan Soukup, Daniel Schmidt, Wei Shou Hu

Research output: Contribution to journalArticlepeer-review

2 Scopus citations

Abstract

An important quality attribute of a recombinant adeno-associated virus (rAAV) as a therapeutic vector is its infectivity. Current assays to quantify infectious rAAV rely on coinfection with a helper virus such as adenovirus (Ad), which requires helper virus preparation and introduces additional variability. A simple method that has high sensitivity and removes the need for helper virus would improve assay consistency and facilitate high-throughput applications such as rAAV producer cell line development. In this study, we describe a stable assay cell line that was generated by integrating the coding sequences for AAV Rep68 and Ad E4orf6 and DNA binding protein under the control of inducible promoters. The Rep68 protein expression was further modulated by a ligand-responsive destabilization domain. In several benchmarks, the cell line gave comparable titers with those obtained using a classical Ad coinfection method. The cell line was also used to titer vectors of multiple rAAV serotypes. This cell line has the potential to serve as an effective and robust tool for quantifying infectious rAAV titers to advance gene therapy vector biomanufacturing.

Original languageEnglish (US)
Pages (from-to)162-170
Number of pages9
JournalHuman gene therapy
Volume34
Issue number3-4
DOIs
StatePublished - Feb 1 2023

Bibliographical note

Funding Information:
This study was funded internally.

Publisher Copyright:
ª 2023 by Mary Ann Liebert, Inc.

Keywords

  • infectious rAAV titer
  • rAAV
  • synthetic biology

PubMed: MeSH publication types

  • Journal Article
  • Research Support, Non-U.S. Gov't

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