Evaluation of screening platforms for virus-like particle production with the baculovirus expression vector system in insect cells

Recombinant protein and virus-like particle (VLP) production based on the baculovirus expression vector system is fast, flexible, and offers high yields. Independent from the product, a multitude of parameters are screened during process development/optimisation. Early development acceleration is a key requirement for economic efficiency, and µ-scale bioreactor systems represent an attractive solution for high-throughput (HTP) experimentation. However, limited practical knowledge is available on the relevance and transferability of screening data to pilot scales and manufacturing. The main goal of the present study was to evaluate a HTP µ-bioreactor platform with respect to its aptitude as a screening platform mainly based on transferability of results to benchtop bioreactors representing the conventional production regime. Second question was to investigate to what extent the online sensors of the µ-bioreactor contribute to process understanding and development. We demonstrated that transferability of infection screening results from the HTP µ-bioreactor scale to the benchtop bioreactor was equal or better than that from shaker cultivation. However, both experimental setups turned out to be sub-optimal solutions that only allowed for a first and rough ranking with low relevance in the case of absolute numbers. Bioreactor yields were up to one order of magnitude higher than the results of screening experiments.

[1]  Vicente Bernal,et al.  Cell density effect in the baculovirus‐insect cells system: A quantitative analysis of energetic metabolism , 2009, Biotechnology and bioengineering.

[2]  L. Miller,et al.  Control of baculovirus polyhedrin gene expression by very late factor 1. , 1998, Virology.

[3]  L. Nielsen,et al.  Effect of the peak cell density of recombinant AcMNPV-infected Hi5 cells on baculovirus yields , 2014, Applied Microbiology and Biotechnology.

[4]  Christian Blesken,et al.  The microfluidic bioreactor for a new era of bioprocess development , 2016 .

[5]  T. Park,et al.  Two - phase cultivation of insect cells for production of recombinant protein , 1995 .

[6]  K. Bayer,et al.  Fed-batch like cultivation in a micro-bioreactor: screening conditions relevant for Escherichia coli based production processes , 2015, SpringerPlus.

[7]  E. Boles,et al.  Parallelised online biomass monitoring in shake flasks enables efficient strain and carbon source dependent growth characterisation of Saccharomycescerevisiae , 2016, Microbial Cell Factories.

[8]  P F Greenfield,et al.  Modeling and optimization of the baculovirus expression vector system in batch suspension culture , 1994, Biotechnology and bioengineering.

[9]  G. Greller,et al.  Single‐use wave‐mixed versus stirred bioreactors for insect‐cell/BEVS‐based protein expression at benchtop scale , 2014 .

[10]  A. Kamen,et al.  Bioprocessing of baculovirus vectors: a review. , 2010, Current Gene Therapy.

[11]  M. Ng,et al.  A novel platform for virus-like particle-display of flaviviral envelope domain III: induction of Dengue and West Nile virus neutralizing antibodies , 2013, Virology Journal.

[12]  S. Meyer,et al.  Identification of Essential Genetic Baculoviral Elements for Recombinant Protein Expression by Transactivation in Sf21 Insect Cells , 2016, PloS one.

[13]  Randal J. Schoepp,et al.  Evaluation of ViroCyt® Virus Counter for Rapid Filovirus Quantitation , 2015, Viruses.

[14]  S. Kain,et al.  Optimized codon usage and chromophore mutations provide enhanced sensitivity with the green fluorescent protein. , 1996, Nucleic acids research.

[15]  D. Murhammer,et al.  Comparison of Trichoplusia ni BTI-Tn-5B1-4 (high five) and Spodoptera frugiperda Sf-9 insect cell line metabolism in suspension cultures. , 1997, Biotechnology and bioengineering.

[16]  Laura Sander,et al.  Using cell size kinetics to determine optimal harvest time for Spodoptera frugiperda and Trichoplusia ni BTI-TN-5B1-4 cells infected with a baculovirus expression vector system expressing enhanced green fluorescent protein , 2007, Cytotechnology (Dordrecht).

[17]  Robert C. Wolpert,et al.  A Review of the , 1985 .

[18]  Marc C. Johnson,et al.  The stoichiometry of Gag protein in HIV-1 , 2004, Nature Structural &Molecular Biology.

[19]  A. Teixeira,et al.  Combining stable insect cell lines with baculovirus-mediated expression for multi-HA influenza VLP production. , 2017, Vaccine.

[20]  James Pettitt,et al.  Comparison of the plaque assay and 50% tissue culture infectious dose assay as methods for measuring filovirus infectivity. , 2013, Journal of virological methods.

[21]  A. Jungbauer,et al.  Globular Head-Displayed Conserved Influenza H1 Hemagglutinin Stalk Epitopes Confer Protection against Heterologous H1N1 Virus , 2016, PloS one.

[22]  J. Vlak,et al.  Baculovirus Expression System , 2020, Definitions.

[23]  F. Gòdia,et al.  Nanoscale characterization coupled to multi-parametric optimization of Hi5 cell transient gene expression , 2018, Applied Microbiology and Biotechnology.

[24]  L. Nielsen,et al.  Low multiplicity infection of insect cells with a recombinant baculovirus: The cell yield concept , 2000, Biotechnology and bioengineering.

[25]  R. Speck,et al.  Ultrasensitive quantitative HIV-1 p24 antigen assay adapted to dried plasma spots to improve treatment monitoring in low-resource settings. , 2006, Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology.

[26]  J. van den Heuvel,et al.  Fast plasmid based protein expression analysis in insect cells using an automated SplitGFP screen , 2016, Biotechnology and bioengineering.

[27]  J E Bailey,et al.  Modeling the population dynamics of baculovirus‐infected insect cells: Optimizing infection strategies for enhanced recombinant protein yields , 1992, Biotechnology and bioengineering.

[28]  A. Kamen,et al.  Stability of Serum‐Free and Purified Baculovirus Stocks under Various Storage Conditions , 2006, Biotechnology progress.

[29]  P. Palese,et al.  An immuno-assay to quantify influenza virus hemagglutinin with correctly folded stalk domains in vaccine preparations , 2018, PloS one.