High ‐yield recombinant adeno‐associated viral vector production by multivariate optimization of bioprocess and transfection conditions

AbstractRecombinant adeno-associated viral vectors (rAAVs) are one of the most used vehicles for gene therapy, with five rAAV therapeutics commercially approved by the FDA. To improve product yield, we optimized the suspension production process of rAAV8 vectors carrying a proprietary transgene using a commercially available transfection reagent, FectoVIR-AAV. Using a miniaturized automated 250  mL scale bioreactor system, we generated models of vector genome (vg) titer, capsid (cp) titer, and Vg:Cp percentage from two multivariate design of experiment studies, one centered around bioreactor operating parameters, and another based on the transfection conditions. Using the optimized proce ss returned from these models, the vector genome titer from the bioreactor was improved to beyond 1 × 1012 vg/mL. Five critical parameters were identified that had large effects on the pre-purification vector quantity—the transfection pH, production pH, complexation time, viable cell density at transfection, and transfection reagent to DNA ratio. The optimized process was further assessed for its pe rformance extending to six AAV serotypes, namely AAV1, AAV2, AAV5, AAV6, AAV8, and AAV9 carrying a transgene encoding for green fluorescent protein (GFP). Five of the six serotypes returned higher vector genome titers than the control condition. These data suggest that the choice of transfection rea gent is a major factor in improving vector yield. The multivariate design of experiment ...
Source: Biotechnology Progress - Category: Biotechnology Authors: Tags: RESEARCH ARTICLE Source Type: research