Toward Shortened the Time-to-Market for Biopharmaceutical Proteins: Improved Fab Protein Expression Stability Using the Cre/lox System in a Multi-Use Clonal Cell Line

Abstract

Stable gene integration and rapid selection of high-expressing clones are important when developing biopharmaceutical systems to produce a protein of interest. According to regulatory guidelines, the final production clones should be stable through multiple cell generations. To achieve long-term stable expression of Fab genes via recombinase-mediated cassette exchange (RMCE), we modified mutual configurations of the lox sequences. By inversion of the spacer orientation, we avoided the loss of the integrated gene after several dozen cycles of cell division. This feature also prevents reversible transgene integration. Although the RMCE allows us to generate transgenic lines rapidly relative to current methods, it remains difficult to obtain stable industrial cell lines for long-term culturing and for the initial development stage. In this study, we present an approach to shortening the timeline for therapeutic protein development. Our approach provides easy access to the same clonal cell line in the initial development phase, and also for the production of biopharmaceutical proteins.