(547d) Model-Based Optimization of Continuous Manufacturing with the Baculovirus Expression Vector System

The baculovirus expression vector system (BEVS) is a promising platform for high yield production of recombinant proteins and more complex biologics, such as virus-like particles and viral vectors for gene therapy.^1–3 Within the BEVS, insect cells, usually derived from Spodoptera frugiperda (Sf9), are infected with recombinant baculoviruses that carry the genetic information for the expression of the product of interest.^4,5 The BEVS is typically employed in batch mode, with Sf9 cells first grown to the desired density and then infected with recombinant baculovirus(es). Within the current interest in transitioning from the traditional batch manufacturing to continuous processing to increase the BEVS productivity, several challenges need to be addressed.^6,7 In this work, a mathematical model is used to design the flowsheet and the operation of a setup for continuous drug substance manufacturing in the BEVS. Considering a case study involving the production of recombinant adeno associated virus (rAAV) vectors for gene therapy applications, the mathematical model is used to assess the optimal configuration of cell growth and infection/production bioreactors. Both perfusion and fully continuous design alternatives are evaluated. An optimal control study is carried out to maximize the process productivity and to limit the formation of baculovirus defective interfering particles that can severely compromise the process efficiency.⁸ The designed control system is implemented and validated on lab-scale bioreactors.

References

1.Cecchini S, Virag T, Kotin RM. Reproducible high yields of recombinant adeno-associated virus produced using invertebrate cells in 0.02-to 200-liter cultures (2011). Hum Gene Ther 22(8), 1021-1030.

2. Smith GE, Summers MD, Fraser MJ. Production of human beta interferon in insect cells infected with a baculovirus expression vector (1983). Mol Cell Biol 24, 434-443.

3. Li J, Li R, Zhang Q, Peng P, Wang X, Gu M, Hu Z, Jiao X, Peng D, Hu J, Liu X (2021). H7N9 influenza virus-like particle based on BEVS protects chickens from lethal challenge with highly pathogenic H7N9 avian influenza virus. Vet Microbiol 258, 109106.

4. Rohrmann GF (2019). Baculovirus Molecular Biology. National Center for Biotechnology Information (US), Bethesda, MD.

5. Smith RH, Levy JR, Kotin RM (2009). A simplified baculovirus-AAV expression vector system coupled with one-step affinity purification yields high-titer rAAV stocks from insect cells. Mol Ther 17(11), 1888-1896.

6. Clément N, Grieger JC. Manufacturing of recombinant adeno-associated viral vectors for clinical trials (2016). Mol Ther Methods Clin Dev 3, 16002.

7. Pijlman GP, De Vrij J, Van Den End FJ, Vlak JM, Martens DE (2004). Evaluation of baculovirus expression vectors with enhanced stability in continuous cascaded insect-cell bioreactors. Biotechnol Bioeng 87(6), 743-753.

8. Giri L, Feiss MG, Bonning BC, Murhammer DW (2012). Production of baculovirus defective interfering particles during serial passage is delayed by removing transposon target sites in fp25k. J Gen Virol 93(2), 389-399.