Overexpression of CHOP alone and in combination with chaperones is effective in improving antibody production in mammalian cells

Abstract

Secretory capacities including folding and assembly are believed to be limiting factors in the establishment of mammalian cell lines producing high levels of recombinant therapeutic proteins. To achieve industrial success, it is also important to improve protein folding, assembly, and secretory processes in combination with increasing transcription and translation. Here, we identified the expression of CHOP/Gadd153 and GRP78, which are unfolded protein response (UPR)-related genes, correlated with recombinant antibody production in stable CHO cells. Subsequently, CHOP overexpression resulted in increasing recombinant antibody production in some mammalian cell lines, and in addition a threefold further enhancement was obtained by combining expression with UPR-related genes or ER chaperones in transient assays. Overexpression of CHOP had no effect on the biochemical characteristics of the product. These results suggest overexpression of CHOP and its combinations may be an effective method to efficiently select a single cell line with a high level of antibody production in the development of cell lines for manufacturing.

Improved antibody production in Chinese hamster ovary cells by ATF4 overexpression

“…The relative expression levels of heavy and light chain mRNAs in DP-12-ATF4-16 did not change significantly through the time course of batch culture compared with the mock-transfected cell line. UPR is thought to be associated with the production of most recombinant proteins (Feige et al 2010;Nishimiya et al 2013). In order to obtain productive cells with higher specific production rates, it is important to engineer the secretory pathway (Hou et al 2012;Le Fourn et al 2013).…”

“…ATF4 and CHOP were significantly elevated in clone DP-12-ATF4-16, which revealed 2.5 times increased productivity with no change in growth rate. Recently, it was reported that CHOP might help to improve antibody secretion (Nishimiya et al 2013). Although CHOP is considered to be a pre-apoptotic gene, it did not affect the growth of any of the confirmed clones in this study.…”

Transcriptional and post-transcriptional targeting for enhanced transient gene expression in CHO cells

“…We have previously shown that DMA addition can lead to improved expression Rajendra et al 2014). In this study, we identified four genes in UPR that may improve expression, namely XBP1S, ATF4, CHOP and GRP78 (Tigges and Fussenegger 2006;Ohya et al 2008;Nishimiya et al 2013;Cain et al 2013;Haredy et al 2013) and evaluated them in combination with DMA for expression of multiple proteins including human and mouse monoclonal antibodies, human bispecific antibodies and Fc-fusion proteins.…”

“…Many UPR genes enhance protein expression in stable cell lines and some genes have a positive impact on transient expression albeit under low expression conditions (Tigges and Fussenegger 2006;Ku et al 2008;Ohya et al 2008;Nishimiya et al 2013;Cain et al 2013;Haredy et al 2013). The goal of this study was to improve transient protein expression in CHO cells without resorting to host cell line or plasmid vector engineering.…”

A protein chimera strategy supports production of a model “difficult‐to‐express” recombinant target

“…Limitations in the secretion of recombinant proteins can impact both protein quality and yield, which can have a negative impact on downstream processes. Published reports have focused on the characterization of such ‘difficult‐to‐express’ recombinant proteins and described the modification of culture conditions 6, 7, 8, 9, 10, 11, 12, 13, 14, 15 or the design of appropriate cell/protein engineering strategies to overcome restrictions in their production 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26. However, little is known regarding the mechanisms underpinning poor recombinant protein production, particularly between proteins of high sequence similarity.…”