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Gas-inducible transgene expression in mammalian cells and mice

Abstract

We describe the design and detailed characterization of a gas-inducible transgene control system functional in different mammalian cells, mice and prototype biopharmaceutical manufacturing. The acetaldehyde-inducible AlcR-PalcA transactivator-promoter interaction of the Aspergillus nidulans ethanol-catabolizing regulon1 was engineered for gas-adjustable transgene expression in mammalian cells. Fungal AlcR retained its transactivation characteristics in a variety of mammalian cell lines and reversibly adjusted transgene transcription from chimeric mammalian promoters (PAIR) containing PalcA-derived operators in a gaseous acetaldehyde-dependent manner. Mice implanted with microencapsulated cells engineered for acetaldehyde-inducible regulation (AIR) of the human glycoprotein secreted placental alkaline phosphatase showed adjustable serum phosphatase levels after exposure to different gaseous acetaldehyde concentrations. AIR-controlled interferon-β production in transgenic CHO-K1-derived serum-free suspension cultures could be modulated by fine-tuning inflow and outflow of acetaldehyde-containing gas during standard bioreactor operation. AIR technology could serve as a tool for therapeutic transgene dosing as well as biopharmaceutical manufacturing.

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Figure 1: Molecular configuration of AIR components.
Figure 2: AIR-adjustable rheostat-like and reversible transgene expression in mammalian cells.
Figure 3: AIR-inducible transgene expression in mice and during bioreactor operation.

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Acknowledgements

We thank Béatrice Felenbok for providing p35S-alcR/pUC and pAlcA, Eva Niederer for FACS sorting, Beat P. Kramer, Alessandro Usseglio Viretta and Laetitia Malphettes for critical comments on the manuscript as well as Harvey Bialy for advice. The work was supported by the Swiss National Science Foundation (grant no. 631-065946) and Cistronics Cell Technology (Einsteinstrasse, P.O.B. 145, CH-8093 Zurich, Switzerland).

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Correspondence to Martin Fussenegger.

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Supplementary information

Supplementary Table 1

Interference studies of acetaldehyde-inducible, macrolide-, streptogramin- and tetracycline-responsive expression systems in CHO-K1 cells (PDF 7 kb)

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Weber, W., Rimann, M., Spielmann, M. et al. Gas-inducible transgene expression in mammalian cells and mice. Nat Biotechnol 22, 1440–1444 (2004). https://doi.org/10.1038/nbt1021

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