Engineered and Natural Promoters and Chromatin‐Modifying Elements for Recombinant Protein Expression in CHO Cells

Abstract

Promoters play a pivotal role in integrating and processing the signals related to transcription initiation. Strong natural viral promoters, such as hCMV or SV40E, have been routinely employed to achieve a high rate of gene expression in ubiquitously used Chinese hamster ovary (CHO) cells. However, viral promoters are susceptible to epigenetic silencing and lack precise regulation levers. This has paved the way to more sensible control elements: endogenous, inducible, and synthetic promoters. In this review we summarize and discuss the use of natural viral, mammalian, and endogenous promoters, as well as recent advances in synthetic promoters and inducible systems for protein expression in CHO cells. Not only the level of transcription, but its long-term stability is crucial for recombinant protein production. Epigenetic chromatin-modifying elements, such as ubiquitously acting chromatin opening elements (UCOEs), matrix attachment regions (MARs), insulators and stabilizing anti-repressors (STARs) significantly improve transcription levels over extended cultivation time and are also discussed here. This review provides up-to date information to facilitate the choice of a suitable promoter and adjacent chromatin-modifying elements to maximize transgene expression as well as ensure long-term expression stability in CHO cell culture.

Bioinformatic Identification of Chinese Hamster Ovary (CHO) Cold‐Shock Genes and Biological Evidence of their Cold‐Inducible Promoters

“…When cells are not pressured by production of the desired protein, they are able to proliferate faster and quickly achieve high densities. Inducing higher production only at that point can improve final yield, product, and clonal stability . The specific productivity of tumor necrosis factor receptor‐immunoglobulin G1 Fc fusion was enhanced around fivefold at 30 °C.…”

“…The choice of a promoter to drive the expression of the gene of interest is an important step in cell line development. Usually, heterologous viral promoters, like CMV and SV40, are used in the mammalian expression cassette . But high expression of the gene of interest by these promoters presents stress for cells which can cause gene silencing, incorrect folding, apoptosis, and clonal instability .…”

Recombinant hemagglutinin produced from Chinese Hamster Ovary (CHO) stable cell clones and a PELC/CpG combination adjuvant for H7N9 subunit vaccine development

“…As the antigenic structures of influenza HA proteins are complex and require extensive N-linked glycosylation to maintain their immunogenicity, one approach is to obtain engineered stable clones of Chinese Hamster Ovary (CHO) cells for the long-term expression of recombinant HA antigens in culture supernatants. Such a production method may allow semi-continuous or continuous harvesting of recombinant glycoproteins within an extended period time, thereby providing the advantage of the development of more cost-effective industrial processes [27,28]. Using CHO cell technology, we have recently obtained the high-yield producer stable cell clones for production of glycan-masking Pan-H5 vaccines [29].…”

Two human MARs effectively increase transgene expression in transfected CHO cells

“…More than 70% of the recombinant therapeutic proteins were produced by the Chinese hamster ovaries (CHO) cells . Low expression level and transgene silencing of recombinant protein in CHO cells are common problems in the production of recombinant protein …”