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Ahn, Junki

Baek, Young-Ok

Hong, Na-young

Kim, Kyung-ho

Kim, Rock-ki

Kim, Yong-Jin

Lee, Joon-Chul

Park, Jeanho

English

Engineering Conferences International

 Poster Number 7 FAST TRACK FED-BATCH CULTURE DEVELOPMENT FOR COVID-19 VACCINE CLINICAL STUDY  Joon-Chul Lee, Korea Institute of Industrial Technology jclee@kitech.re.kr Jeanho Park, Korea Institute of Industrial Technology Junki Ahn, Korea Institute of Industrial Technology Yong-Jin Kim, Korea Institute of Industrial Technology  Na-young Hong, Korea Institute of Industrial Technology Rock-ki Kim, Eubiologics Kyung-ho Kim, Eubiologics Young-Ok Baek, Eubiologics   Key Words: fed-batch culture, cell culture media, productivity increase, media fortification, DOE  We have been fighting an epidemic called SARS-CoV-2, a COVID-19 virus, for the past 3 years. Many infections resulted in death and sequelae. In reality, vaccine platforms using mRNA are the mainstay, but some people cannot get this vaccine because of an underlying disease. Therefore, developing a subunit vaccine with fewer side effects is still needed.  A COVID-19 virus enters the host cell through the spike protein, which is tightly bound to sugar chains. The spike protein exists as a trimer and binds to the host cell’s receptor through RBD (Receptor Binding Domain). So, the ACE2 binding site (Receptor Binding Motif, RBM) was used as our vaccine target. The antigen was composed of a total of 229 amino acid residues by binding 6 histidine residues to the C-terminus of the RBD amino acid sequence. This produces an antibody against the ACE2 binding site to block the invasion pathway and induces defense against the virus.   When producing phase 1 clinical RBD vaccine, we consumed a lot of culture time and costs because of the low RBD concentration in the CHO cell culture process. Therefore, it was necessary to increase the RBD concentration. This study was carried out to rapidly increase productivity within 3 months, including fortifying the commercial medium used in the past.  Firstly, an experiment was conducted on the effect of feeding medium, culture temperature, feeding start day, and feeding volume. As a result of the experiment, it was found that specific feed media, culture temperature of 36 °C, feed start day 4th day, and feeding volume of 2.5% were the optimal conditions. And a temperature shift experiment was performed at 32°C and 34°C, and as a result, it showed a high concentration at 34 °C. So far, the concentration has risen 5 times compared to the control  Subsequently, we used the addition of media components to the enriched medium using a design of experiment (DOE) methodology. An experiment of 10 medium component addition was conducted on the commercial base medium used in the past for screening DOE using the fractional factorial design method, and as a result, we could determine that niacinamide, ATA, and i-inositol are significant factors. Using these factors, we could fasten the optimal concentrations using the Box-Behnken design method. We confirmed the results of the DOE study by a verification experiment in a 50L Single Use bioreactor, We showed that the optimal amount of niacinamide, ATA, and i-inositol were expressed at the highest RBD concentration. It was possible to increase the RBD concentration by almost 4.5-folds in a short-time DOE study. In conclusion, we successfully and effectively increased the productivity of the fed-batch culture in a short period to produce a vaccine for a phase 2 study.  Reference  1. Christos P. Design of Experiment as a Tool for Optimization in Recombinant Protein Biotechnology: From Constructs to Crystals. Molecular Biotechnology, 2019. 61: 873-891 2. Nicola McGillcuddy. et. al. Examining the sources of variability in cell culture media used for biopharmaceutical production. Biotechnol. Lett. 2018. 40: 5-21 3. Roisin O’Flaherty et. al. Mammalian cell culture for production of recombinant proteins: A review of the critical steps in their biomanufacturing. Biotechnol. Advances. 2020. 43 

Fast track fed-batch culture development for COVID-19 vaccine clinical study

https://dc.engconfintl.org/cgi/viewcontent.cgi?filename=0&article=1059&context=ccexviii&type=additional

Fast track fed-batch culture development for COVID-19 vaccine clinical study

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Engineering Conferences International