Multigene engineering of secretion in mammalian cell factories

Chinese hamster ovary cells are widely used in the production of biopharmaceuticals such as monoclonal antibodies. Since their first use for this application in the 1980s much engineering of manufacturing processes and the CHO cell itself has been undertaken, leading to increased cell productivity, product specificity and overall product titres and yields. Much CHO cell engineering has thus far focused on increasing transcription and translation levels, with the engineering of protein folding and modification also being targeted. Whilst this has increased the protein production capacity of the CHO cell it has also had the effect of increasing pressure on the CHO biosynthetic and secretory pathways. The secretory pathway of the CHO cell is not adapted to high levels of secretion as is seen in plasma cells, the main function of which is to produce and secret high levels of antibodies. As such an increase in secretory load upon the CHO cell has introduced bottlenecks that limit the amount of recombinant product that can be transported out of the cell. Engineering strategies to ease these bottlenecks have so far focused on expression of single genes or two genes in combination. Two engineering strategies were utilised to engineer the CHO cell secretory pathway and enhance CHO cell productivity. Firstly a literature- and 'omics-driven approach was used to select genes with which to engineer the CHO secretory pathway. Engineering of the secretory and biosynthetic pathways, using single genes and multiple genes in combination, showed that this approach can increase CHO productivity levels through enhancing specific productivity and overall titre. Secondly, directed evolution was used as a method to bring about myriad fine global changes within the CHO biosynthetic and secretory pathways. From these results we hypothesise that a more holistic, global approach to biosynthetic and secretory pathway engineering proves more successful in increasing CHO cell productivity when compared to more directed techniques.