Process Development of Recombinant Adeno-Associated Virus Production Platform Results in High Production Yield and Purity

Zhao, Huiren; Meisen, Hans; Wang, Songli; Lee, Ki Jeong

doi:10.1089/hum.2022.153

Cited by 4 publications

(1 citation statement)

References 21 publications

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“…Parameters including culture duration and harvest time should also be considered. Although rAAV was found to secret into cell culture media (Vandenberghe et al, 2010), a cell lysis step with detergents (e.g., Triton X‐100, Tween‐20) is widely implemented as the secretion is largely serotype‐dependent, and significant amount of rAAV product could remain intracellular at the end of production (Zhao et al, 2023). Virus harvest is normally carried out 48–72 h posttransfection (hpt) due to limited culture life resulting from virus‐mediated cell lysis (Table 4).…”

Section: Production Process Optimizationmentioning

confidence: 99%

Recent advances in upstream process development for production of recombinant adeno‐associated virus

Ou,

Tang,

et al. 2023

Biotech & Bioengineering

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Recombinant adeno‐associated virus (rAAV) is rapidly emerging as the preferred delivery vehicle for gene therapies, with promising advantages in safety and efficacy. Key challenges in systemic in‐vivo rAAV gene therapy applications are the gap in production capabilities versus potential market demand and complex production process. This review summarizes current available information on rAAV upstream manufacturing processes and proposed optimizations for production. The advancements in rAAV production media were reviewed with proposals to speed up the cell culture process development. Furthermore, major methods for genetic element delivery to host cells were summarized with their advantages, limitations, and future directions for optimization. In addition, culture vessel selection criteria were listed based on production cell system, scale, and development stage. Process control at the production step was also outlined with an in‐depth understanding of production kinetics and quality control.

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Section: Production Process Optimizationmentioning

confidence: 99%

Recent advances in upstream process development for production of recombinant adeno‐associated virus

Ou,

Tang,

et al. 2023

Biotech & Bioengineering

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Biomanufacturing Aspects of Gene Therapy

Warnock

2023

Cell Engineering

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Comprehensive Comparison of AAV Purification Methods: Iodixanol Gradient Centrifugation vs. Immuno-Affinity Chromatography

Lam,

Mulcrone,

Frabutt

et al. 2023

Advances in Cell and Gene Therapy

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Recombinant adeno-associated viruses (AAVs) have emerged as a widely used gene delivery platform for both basic research and human gene therapy. To ensure and improve the safety profile of AAV vectors, substantial efforts have been dedicated to the vector production process development using suspension HEK293 cells. Here, we studied and compared two downstream purification methods, iodixanol gradient ultracentrifugation versus immuno-affinity chromatography (POROS™ CaptureSelect™ AAVX column). We tested multiple vector batches that were separately produced (including AAV5, AAV8, and AAV9 serotypes). To account for batch-to-batch variability, each batch was halved for subsequent purification by either iodixanol gradient centrifugation or affinity chromatography. In parallel, purified vectors were characterized, and transduction was compared both in vitro and in vivo in mice (using multiple transgenes: Gaussia luciferase, eGFP, and human factor IX). Each purification method was found to have its own advantages and disadvantages regarding purity, viral genome (vg) recovery, and relative empty particle content. Differences in transduction efficiency were found to reflect batch-to-batch variability rather than disparities between the two purification methods, which were similarly capable of yielding potent AAV vectors.

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Process Development of Recombinant Adeno-Associated Virus Production Platform Results in High Production Yield and Purity

Cited by 4 publications

References 21 publications

Recent advances in upstream process development for production of recombinant adeno‐associated virus

Recent advances in upstream process development for production of recombinant adeno‐associated virus

Biomanufacturing Aspects of Gene Therapy

Comprehensive Comparison of AAV Purification Methods: Iodixanol Gradient Centrifugation vs. Immuno-Affinity Chromatography

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