Single Domain Antibody (sdAb) Expression in Mammalian Cell
Mammalian Cell System for sdAb Production
In the past decades, the production levels for functional antibodies in mammalian cell expression systems have improved dramatically. Mammalian cells are preferred hosts for the production of secreted proteins harboring posttranslational modifications that are often necessary for their optimal biological activity. Transient or stable transfection of different cell lines can be used for sdAbs expression; at the same time, the expression vector engineering could also increase the expression levels to several hundred milligrams per liter. Because of the improvements in expression hosts, expression vectors, cell culture media, and production processes, transient expression in mammalian cells has been used more and more widely to produce large quantities of recombinant antibodies in a timely and cost-effective way.
Basic Strategies for Mammalian Cell System
High producing cell lines can be isolated after stable integration of transgenes into a transcriptionally active site of host genome using vectors containing the exogenous gene of interest. Production of antibody fragments in mammalian cells has made advances from the early times of low-density adherent cultures to the recent high-density cultures of suspension growing cells. If large-scale Ab production in suspension cells is carried out, cells should be adapted to suspension culture conditions after successful transfection.
Thereinto, suspension-adapted Chinese hamster ovary (CHO) cells have been rapidly growing as the most popular expression systems for expression of sdAbs; besides, sdAbs expression levels can reach up to 100 mg/L in shake flasks. CHO cell lines have recently become the preferred hosts for transient mammalian expression to manufacture a majority of the clinically approved biologics. Compared with the stable expression produced in CHO cells, a transient and semi-stable expression produced in human embryonic kidney 293 (HEK293) cells are also the most frequently used for large-scale transient expression of antibody fragments. Coupled with process optimizations and co-transfection of various expression-enhancing molecules, expression levels in HEK293 cell lines can reach 1 g/L on a routine basis.
Fig. 1 The typical process of mammalian cell expression system.1
Brief Workflow for sdAb Production
Advantages of Mammalian Cell System
Expression of sdAbs in mammalian cells is a desirable strategy to ensure the production of biologically functional sdAbs. Because it presents native leader sequences and appropriate folding factors to ensure successful secretion and has the capacity for suitable posttranslational modifications. Several approaches, including cell host engineering, expression vector engineering, production process, and media optimizations have been used to improve the mammalian cell expression system. As a result, the productivity of sdAbs in mammalian cells continues to increase. VHH-Fc antibodies composed of sdAbs and the Fc domain prolong the serum half-life and increase functional affinity, and these VHH-Fc antibodies require a eukaryotic protein folding machinery. Thus, mammalian cells that effectively promote correct posttranslational modifications are the most widely used expression platform under the circumstances.
Reference
- Lai, Tingfeng, Yuansheng Yang, and Say Kong Ng. "Advances in mammalian cell line development technologies for recombinant protein production." Pharmaceuticals 6.5 (2013): 579-603. Distributed under Open Access license CC BY 3.0, without modification.
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