title: Gas-inducible product gene expression in bioreactors.
abstract: Inducible transgene expression technologies are of unmatched potential for biopharmaceutical manufacturing of unstable, growth-impairing and cytotoxic proteins as well as conditional metabolic engineering to improve desired cell phenotypes. Currently available transgene dosing modalities which rely on physical parameters or small-molecule drugs for transgene fine-tuning compromise downstream processing and/or are difficult to implement technologically. The recently designed gas-inducible acetaldehyde-inducible regulation (AIR) technology takes advantage of gaseous acetaldehyde to modulate product gene expression levels. At regulation effective concentrations gaseous acetaldehyde is physiologically inert and approved as food additive by the Federal Drug Administration (FDA). During standard bioreactor operation, gaseous acetaldehyde could simply be administered using standard/existing gas supply tubing and eventually eliminated by stripping with inducer-free air. We have determined key parameters controlling acetaldehyde transfer in three types of bioreactors and designed a mass balance-based model for optimal product gene expression fine-tuning using gaseous acetaldehyde. Operating a standard stirred-tank bioreactor set-up at 10 L scale we have validated AIR technology using CHO-K1-derived serum-free suspension cultures transgenic for gas-inducible production of human interferon-beta (IFN-beta). Gaseous acetaldehyde-inducible IFN-beta production management was fully reversible while maintaining cell viability at over 95% during the entire process. Compatible with standard bioreactor design and downstream processing procedures AIR-based technology will foster novel opportunities for pilot and large-scale manufacturing of difficult-to-produce protein pharmaceuticals.
authors.affiliation: Institute for Chemical and Bio-Engineering, Swiss Federal Institute of Technology Zurich, ETH Hoenggerberg HCI F115, Wolfgang-Pauli-Strasse 10, CH-8093 Zurich, Switzerland.
authors.forename: Wilfried
authors.initials: W
authors.lastname: Weber
authors.forename: Markus
authors.initials: M
authors.lastname: Rimann
authors.forename: François-Nicolas
authors.initials: FN
authors.lastname: de Glutz
authors.forename: Eric
authors.initials: E
authors.lastname: Weber
authors.forename: Klaus
authors.initials: K
authors.lastname: Memmert
authors.forename: Martin
authors.initials: M
authors.lastname: Fussenegger
mesh_terms: D000079:Acetaldehyde / Q000494:pharmacology*; D000818:Animals; D019149:Bioreactors; D016466:CHO Cells; D018929:Cell Culture Techniques / Q000379:methods; D003198:Computer Simulation; D006224:Cricetinae; D003412:Cricetulus; D005740:Gases / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D016899:Interferon-beta / Q000096:biosynthesis* / Q000235:genetics; D008954:Models, Biological; D010865:Pilot Projects; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000096:biosynthesis*
chemical_list: D005740:Gases; D011994:Recombinant Proteins; D016899:Interferon-beta; D000079:Acetaldehyde