Construction strategies for developing expression vectors for recombinant monoclonal antibody production in CHO cells

Abstract

Recent years have seen the use of recombinant proteins in the treatment of different diseases. Among them, monoclonal antibodies (mAbs) are currently the fastest growing class of bio-therapeutic recombinant proteins. Chinese hamster ovary (CHO) cells are the most commonly used host cells for production of these recombinant mAbs. Expression vectors determine the expression level and quality of recombinant mAbs. Currently, few construction strategies for recombinant mAbs expression vectors in CHO cells have been developed, including monocistronic vector, multiple-promoter expression vector, and tricistronic vector mediated by internal ribosome entry site (IRES) or Furin-2A element. Among them, Furin-2A-mediated vector is an effective approach due to advantages of high "self-cleavage" efficiency, and equal expression of light and heavy chains from a single open reading frame. Here, we have reviewed the progress in development of different strategies for constructing recombinant mAb expression vectors in CHO cells and its potential advantages and disadvantages.

Cell line development for continuous high cell density biomanufacturing: Exploiting hypoxia for improved productivity

“…In the current study, we created a novel hypoxia responding cell system in CHO production cells based on HRE elements and cellular oxygen sensing proteins. We were able to demonstrate substantial improvement of recombinant protein production under hypoxic process conditions using a vector based system which can be used without time- and cost-intensive host cell modifications in industrial settings ( Li et al, 2018 ). Specifically, we first verified the conservation of the hypoxia-responsive pathway in CHO cell lines, functionally analyzed HREs from VEGF and EPO genes and then created hypoxia-responsive expression vectors.…”

Bioprocessing of mAb (Monoclonal Antibodies) using Chinese Hamster Ovary (CHO) Cells: A Review

“…Controlling the LC:HC peptide ratio is essential to the kinetics of mAb formation. It was reported that excess LC and an LC:HC ratio of more than 1.5 are favored since incompletely formed HCs can cause the mAb clones to aggregate and have inconsistent glycosylation (Ahmadi et al, 2017;Ho et al, 2013a;Ho et al, 2013-b;Li et al, 2018). The DNA fragments are then joined together to the plasmid by the enzyme ligase, such as T4 DNA ligase, or by overlap extension PCR and cloned (Haryadi et al, 2015;Voronina et al, 2016).…”

Progress of Transposon Vector System for Production of Recombinant Therapeutic Proteins in Mammalian Cells

“…Compared with prokaryotic, yeast and insect cells, CHO cells have the following advantages ( Khan 2013 ): 1) CHO cells can grow under the adhere and suspension state with high cell density, which are suitable for large-scale industrial production ( Lai et al, 2013 ): 2) CHO cells are less sensitive to human virus infection ( O'Flaherty et al, 2020 ): 3) The expressed proteins have high similarity with natural proteins in terms of molecular structure, physicochemical properties and biological functions, and the glycosylation is also more similar to that of human-derived cells due to the lack of immunogenic α-galactose epitopes ( Ghaderi et al, 2012 ): 4) CHO cells are fibroblasts with low endogenous protein secretion, which is beneficial to the isolation and purification of recombinant proteins. ( Mohan et al, 2008 ; Li et al, 2018 ). In addition, by constructing DNA methyltransferase-deficient CHO cells, the stability of expression for the recombinant protein can be enhanced by inactivating DNA methylation ( Jia et al, 2018 ; Wang et al, 2019 ).…”