Production of cell culture (MDCK) derived live attenuated influenza vaccine (LAIV) in a fully disposable platform process

Abstract

The majority of influenza vaccines are manufactured using embryonated hens' eggs. The potential occurrence of a pandemic outbreak of avian influenza might reduce or even eliminate the supply of eggs, leaving the human population at risk. Also, the egg-based production technology is intrinsically cumbersome and not easily scalable to provide a rapid worldwide supply of vaccine. In this communication, the production of a cell culture (Madin-Darby canine kidney (MDCK)) derived live attenuated influenza vaccine (LAIV) in a fully disposable platform process using a novel Single Use Bioreactor (SUB) is presented. The cell culture and virus infection was maintained in a disposable stirred tank reactor with PID control of pH, DO, agitation, and temperature, similar to traditional glass or stainless steel bioreactors. The application of this technology was tested using MDCK cells grown on microcarriers in proprietary serum free medium and infection with 2006/2007 seasonal LAIV strains at 25-30 L scale. The MDCK cell growth was optimal at the agitation rate of 100 rpm. Optimization of this parameter allowed the cells to grow at a rate similar to that achieved in the conventional 3 L glass stirred tank bioreactors. Influenza vaccine virus strains, A/New Caledonia/20/99 (H1N1 strain), A/Wisconsin/67/05 (H3N2 strain), and B/Malaysia/2506/04 (B strain) were all successfully produced in SUB with peak virus titers > or =8.6 log(10) FFU/mL. This result demonstrated that more than 1 million doses of vaccine can be produced through one single run of a small bioreactor at the scale of 30 L and thus provided an alternative to the current vaccine production platform with fast turn-around and low upfront facility investment, features that are particularly useful for emerging and developing countries and clinical trial material production.

Safety, immunogenicity and infectivity of new live attenuated influenza vaccines

“…While this technology is extremely robust, the use of established cell lines rather than fertilized hens' eggs for vaccine manufacture could have significant advantages, for example, better consistency of production, easier scaling-up, independence from egg supply and maintenance of chicken flocks, a reduced risk of microbial contamination, a non-animal origin process, a more homogeneous virus yield and the possibility of amplifying viruses that grow poorly in eggs [67,68]. The ability of cold-adapted LAIV strains to grow in MDCK cells has been evaluated in several studies, using small-scale and large-scale production processes, for both licensed LAIV backbones [69][70][71][72][73][74][75]. MedImmune established its own MDCK cell line, which allowed the growth of LAIV viruses to high titers with no significant impact on antigenic and growth properties [71,73].…”

Single-Use Bioreactors for Animal and Human Cells

“…African green monkey kidney-derived Vero cells, Madin-Darby Canine Kidney cells or PBS-1 cells) (Josefsberg and Buckland 2012;Whitford and Fairbank 2011). Both stirred (Chaubard et al 2010;George et al 2010) and wave-mixed (Genzel et al 2010;Magnusson et al 2011) SU systems operated with microcarriers as well as fixed bed bioreactors with fiber carriers (Moncaubeig 2013) have proven themselves.…”

“…Different studies, where MDCK and Vero cells were grown on microcarriers, revealed that tenfold higher cell densities and virus titers can easily be achieved when using wave-mixed and stirred bioreactors for virus production (e.g. Influenza, Polio, mink enteritis virus) (Genzel et al 2004(Genzel et al , 2006Hundt et al 2007;George et al 2010;Schouwenberg et al 2010;Thomassen et al 2012).…”

Cell culture-based influenza vaccines: A necessary and indispensable investment for the future

“…The cells can be easily adapted to and be grown in serum-free media, and in suspension, as well as on various microcarriers maintained under various bioreactor conditions. 93,[101][102][103][104][105][106][107][108] Subclones of MDCK cells adopted to grow in suspension and support robust virus production have also been described, 91,108,109 although adherent MDCK cells appear to support more robust virus production than suspension MDCK cells. 110 Influenza vaccines derived from MDCK cells are also safe and immunogenic.…”