From single‐cell cloning to high‐yield influenza virus production – implementing advanced technologies in vaccine process development

Abstract

AbstractInnovations in viral vaccine manufacturing are crucial for pandemic preparedness and to meet ever‐rising global demands. For influenza, however, production still mainly relies on technologies established decades ago. Although modern production shifts from egg‐based towards cell culture technologies, the full potential has not yet been fully exploited. Here, we evaluate whether implementation of state‐of‐the‐art technologies for cell culture‐based recombinant protein production are capable to challenge outdated approaches in viral vaccine process development. For this, a fully automated single‐cell cloning strategy was established to generate monoclonal suspension Madin‐Darby canine kidney (MDCK) cells. Among selected cell clones, we could observe distinct metabolic and growth characteristics, with C59 reaching a maximum viable cell concentration of 17.3 × 106 cells/mL and low doubling times in batch mode. Screening for virus production using a panel of human vaccine‐relevant influenza A and B viruses in an ambr15 system revealed high titers with yields competing or even outperforming available MDCK cell lines. With C113, we achieved cell‐specific virus yields of up to 25,000 virions/cell, making this cell clone highly attractive for vaccine production. Finally, we confirmed process performance at a 50‐fold higher working volume. In summary, we present a scalable and powerful approach for accelerated development of high‐yield influenza virus production in chemically defined medium starting from a single cell.