title: Engineering of Escherichia coli strains for plasmid biopharmaceutical production: scale-up challenges.
abstract: Plasmid-based vaccines and therapeutics have been making their way into the clinic in the last years. The existence of cost-effective manufacturing processes capable of delivering high amounts of high-quality plasmid DNA (pDNA) is essential to generate enough material for trials and support future commercialization. However, the development of pDNA manufacturing processes is often hampered by difficulties in predicting process scale performance of Escherichia coli cultivation on the basis of results obtained at lab scale. This paper reports on the differences observed in pDNA production when using shake flask and bench-scale bioreactor cultivation of E. coli strains MG1655ΔendAΔrecA and DH5α in complex media with 20 g/L of glucose. MG1655ΔendAΔrecA produced 5-fold more pDNA (9.8 mg/g DCW) in bioreactor than in shake flask (1.9 mg/g DCW) and DH5α produced 4-fold more pDNA (8 mg/g DCW) in bioreactor than in shake flask (2 mg/g DCW). Accumulation of acetate was also significant in shake flasks but not in bioreactors, a fact that was attributed to a lack of control of pH.
authors.affiliation: MIT-Portugal Program, Portugal; Department of Bioengineering, Instituto Superior Técnico (IST), Lisbon, Portugal; IBB-Institute for Biotechnology and Bioengineering, Center for Biological and Chemical Engineering, IST, Lisbon, Portugal.
authors.forename: Geisa A L
authors.initials: GA
authors.lastname: Gonçalves
authors.affiliation: MIT-Portugal Program, Portugal; Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA, United States.
authors.forename: Kristala L J
authors.initials: KL
authors.lastname: Prather
authors.affiliation: MIT-Portugal Program, Portugal; Department of Bioengineering, Instituto Superior Técnico (IST), Lisbon, Portugal; IBB-Institute for Biotechnology and Bioengineering, Center for Biological and Chemical Engineering, IST, Lisbon, Portugal.
authors.forename: Gabriel A
authors.initials: GA
authors.lastname: Monteiro
authors.affiliation: MIT-Portugal Program, Portugal; Department of Bioengineering, Instituto Superior Técnico (IST), Lisbon, Portugal; IBB-Institute for Biotechnology and Bioengineering, Center for Biological and Chemical Engineering, IST, Lisbon, Portugal. Electronic address: miguelprazeres@ist.utl.pt.
authors.forename: Duarte M F
authors.initials: DM
authors.lastname: Prazeres
mesh_terms: D061249:Batch Cell Culture Techniques; D019149:Bioreactors; D004926:Escherichia coli / Q000235:genetics*; D005285:Fermentation; D005818:Genetic Engineering; D010957:Plasmids / Q000096:biosynthesis*; D013678:Technology, Pharmaceutical / Q000379:methods
keywords: DNA vaccine; Escherichia coli; Fermentation; Plasmid biopharmaceuticals; Scale-up process; Strain engineering