A Flexible Hybrid Site-Specific Integration-Based Expression System in CHO Cells for Higher-Throughput Evaluation of Monoclonal Antibody Expression Cassettes
Date
2025-01-20
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
Biotechnology Journal
Abstract
The implementation of site-specific integration (SSI) systems in Chinese hamster ovary (CHO) cells for the production of monoclonal antibodies (mAbs) can alleviate concerns associated with production instability and reduce cell line development timelines. SSI cell line performance is driven by the interaction between genomic integration location, clonal background, and the transgene expression cassette, requiring optimization of all three parameters to maximize productivity. Systematic comparison of these parameters has been hindered by SSI platforms involving low-throughput enrichment strategies, such as cell sorting. This study presents a recombinase-mediated cassette exchange (RMCE)-capable SSI system that uses only chemical selection to enrich for transgene-expressing RMCE pools in less than one month. The system was used to compare eight mAb expression cassettes containing two novel genetic regulatory elements, the Azin1 CpG island and the Piggybac transposase 5’ terminal repeat, in various orientations to improve the expression of two therapeutic mAbs from two genomic loci. Similar patterns of productivity and mRNA expression were observed across sites and mAbs, and the best performing cassette universally increased mAb productivity by 7- to 11-fold. This flexible system allows for higher-throughput comparison of expression cassettes from a consistent clonal and transcriptional background to optimize RMCE-derived cell lines for industrial production of mAbs.
Description
This article was originally published in Biotechnology Journal. The version of record is available at: https://doi.org/10.1002/biot.202400520.
© 2025 The Author(s). Biotechnology Journal published by Wiley-VCH GmbH.
This is an open access article under the terms of the Creative Commons Attribution-NonCommercial License (http://creativecommons.org/licenses/by-nc/4.0/), which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.
Keywords
bioprocessing, cell line development, CHO cells, monoclonal antibodies, recombinase-mediated cassette exchange
Citation
Szkodny, A.C. and Lee, K.H. (2025), A Flexible Hybrid Site-Specific Integration-Based Expression System in CHO Cells for Higher-Throughput Evaluation of Monoclonal Antibody Expression Cassettes. Biotechnol. J., 20: e202400520. https://doi.org/10.1002/biot.202400520