The vaccine manufacturing industry is continuously seeking more flexible and time-saving solutions for a quicker response to pandemic outbreaks (1, 2). For influenza, vaccine is traditionally produced in fertilized hen eggs and purified in fixed steel equipment. Here we describe the scale-up of a downstream purification process of cell-based influenza virus, from a 10 L cell culture to the purification from a 50 L culture using ReadyToProcess single-use technology. Single-use technology reduces the need for time-consuming device preparations, such as cleaning and cleaning validation, and minimizes the risk for cross-contamination, all essential parameters in traditional vaccine manufacturing.

Virus Purification

Influenza virus was propagated in Vero cells grown on microcarriers (3). Virus particles were subsequently purified using robust and scalable single-use filtration and chromatography techniques. In the laboratory-scale experiment, standard equipment was used for the downstream purification of influenza virus from a 10 L cell culture. The laboratory-scale purification process was subsequently scaled up 62.5-fold. In the scaled-up process, ReadyToProcess single-use equipment was used for downstream virus purification from a 50 L cell culture (Figure 1).

Figure 1:  ()

The outcome from the process steps of the influenza purification from both 10 L and 50 L production scales is shown in Figure 2. A more detailed description of the purification is given in Application note 29-04-35-49 (4).

Figure 2:  ()

Comparison to Regulatory Specifications

The output from the scaled-up production is compared with a commercially available specification for a nasal LAIV and a specification for egg-based, split-inactivated influenza vaccine from WHO (5, 6) (Table 1). Assuming a nasal route ofadministration and doses of 10⁷ infectious particles per 0.2 mL dose, the amounts of host cell DNA and protein were shown to be below the acceptance levels defined by WHO. The outcome from the scaled-up production indicates that it is possible to obtain approximately 3000 doses/L harvest, corresponding to harvests of 325 L for 1 million doses (calculations based on specification for nasal LAIV) and 175 doses/L harvest corresponding to harvests of 5760 L for 1 million doses (calculations based on specification for split-inactivated vaccine).

Table 1: 

Table 1: 194; ()

Conclusions

In In this case study, the downstream purification of cell-based influenza virus from a 50 L cell culture using ReadyToProcess single-use technology demonstrates high productivity and efficient removal of host cell–derived impurities, with maintained virus infectivity. Similar process yield and purity was obtained from the scaled-up virus purification from a 50 L cell culture, as from the purification from a 10 L cell culture.

About the Author

Author Details
Peder Bergvall and Jakob Liderfelt are scientists, and Christine Sund Lundström is a senior research engineer at GE Healthcare Bio-Sciences AB, Björkgatan 30, 751 84 Uppsala, Sweden.

REFERENCES

1.) Pau, MG. 2001. The Human Cell Line PER.C6 Provides a New Manufacturing System for the Production of Influenza Vaccines. Vaccine 19:2716-2721.

2.) Kistner, O. 1998. Development of a Mammalian Cell (Vero) Derived Candidate Influenza Virus Vaccine. Vaccine 16:960-968.

3.) Application Note 29-0435 48, and AA Edition. 2013.Scale-Up of Adherent Vero Cells Grown on Cytodex Microcarriers Using ReadyToProcess Equipment, GE Healthcare, Uppsala.

4.) Application Note 29-0435-49, and AA Edition. 2013.Downstream Scale-Up Purification of Influenza Virus Using ReadyToProcess Equipment, GE Healthcare, Uppsala.

5.) 2005. Recommendations for the Production and Control of Influenza Vaccine (Inactivated). World Health Organization Tech. Rep. Ser 927:103.

6.) White Paper 29-0435-51, and AA Edition. 2013.Overview of a Scale-Up of a Cell Based Influenza Virus Production Process Using ReadyToProcess Equipment, GE Healthcare, Uppsala.