Recent developments in miRNA based recombinant protein expression in CHO

Bazaz, Masoume; Abdolzadeh, Ahmad; Azizi, Mohammad; Mahboudi, Fereidoun; Davoudi, Noushin

doi:10.1007/s10529-022-03250-1

Cited by 5 publications

(2 citation statements)

References 77 publications

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“…Previously, we reviewed recent developments in miRNA engineering in CHO cells. (Bazaz et al 2022 ). In this study, we selected a known upregulated microRNA in cancer as a novel candidate to improve the productivity of CHO cells.…”

Section: Discussionmentioning

confidence: 98%

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Overexpression of miR-32 in Chinese hamster ovary cells increases production of Fc-fusion protein

et al. 2023

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The demand for industrial genetically modified host cells were increased with the growth of the biopharmaceutical market. Numerous studies on improving host cell productivity have shown that altering host cell growth and viability through genetic engineering can increase recombinant protein production. During the last decades, it was demonstrated that overexpression or downregulation of some microRNAs in Chinese Hamster Ovary (CHO) cells as the host cell in biopharmaceutical manufacturing, can improve their productivity. The selection of microRNA targets has been based on their previously identified role in human cancers. MicroRNA-32 (miR-32), which is conserved between humans and hamsters (Crisetulus griseus), was shown to play a role in the regulation of cell proliferation and apoptosis in some human cancers. In this study, we investigated the effect of miR-32 overexpression on the productivity of CHO-VEGF-trap cells. Our results indicated that stable overexpression of miR-32 could dramatically increase the productivity of CHO cells by 1.8-fold. It also significantly increases cell viability, batch culture longevity, and cell growth. To achieve these results, following the construction of a single clone producing an Fc-fusion protein, we transfected cells with a pLexJRed-miR-32 plasmid to stably produce the microRNA and evaluate the impact of mir-32 overexpression on cell productivity, growth and viability in compare with scrambled control. Our findings highlight the application of miRNAs as engineering tools and indicated that miR-32 could be a target for engineering CHO cells to increase cell productivity.

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Section: Discussionmentioning

confidence: 98%

“…Several studies indicate that deregulation of microRNAs in CHO cells can increase cell productivity (see Table 1 ) by interfering with a variety of cellular mechanisms such as proliferation, post-translational modification, secretion, and apoptosis. (Singh et al 2022 ; Bazaz et al 2022 ; Koh et al 2009 ).…”

Section: Introductionmentioning

confidence: 99%

Overexpression of miR-32 in Chinese hamster ovary cells increases production of Fc-fusion protein

et al. 2023

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CHO cell engineering via targeted integration of circular miR-21 decoy using CRISPR/RMCE hybrid system

Adibzadeh,

Amiri,

Barkhordari

et al. 2024

Appl Microbiol Biotechnol

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Chinese hamster ovary (CHO) cells, widely acknowledged as the preferred host system for industrial recombinant protein manufacturing, play a crucial role in developing pharmaceuticals, including anticancer therapeutics. Nevertheless, mammalian cell-based biopharmaceutical production methods are still beset by cellular constraints such as limited growth and poor productivity. MicroRNA-21 (miR-21) has a major impact on a variety of malignancies, including glioblastoma multiforme (GBM). However, reduced productivity and growth rate have been linked to miR-21 overexpression in CHO cells. The current study aimed to engineer a recombinant CHO (rCHO) cell using the CRISPR-mediated precise integration into target chromosome (CRIS-PITCh) system coupled with the Bxb1 recombinase-mediated cassette exchange (RMCE) to express a circular miR-21 decoy (CM21D) with five bulged binding sites for miR-21 sponging. Implementing the ribonucleoprotein (RNP) delivery method, a landing pad was inserted into the genome utilizing the CRIS-PITCh technique. Subsequently, the CM21D cassette flanked by Bxb1 attB was then retargeted into the integrated landing pad using the RMCE/Bxb1 system. This strategy raised the targeting efficiency by 1.7-fold, and off-target effects were decreased. The miR-21 target genes (Pdcd4 and Atp11b) noticed a significant increase in expression upon the miR-21 sponging through CM21D. Following the expression of CM21D, rCHO cells showed a substantial decrease in doubling time and a 1.3-fold increase in growth rate. Further analysis showed an increased yield of hrsACE2, a secretory recombinant protein, by 2.06-fold. Hence, we can conclude that sponging-induced inhibition of miR-21 may lead to a growth rate increase that could be linked to increased CHO cell productivity. For industrial cell lines, including CHO cells, an increase in productivity is crucial. The results of our research indicate that CM21D is an auspicious CHO engineering approach. Key points • CHO is an ideal host cell line for producing industrial therapeutics manufacturing, and miR-21 is downregulated in CHO cells, which produce recombinant proteins. • The miR-21 target genes noticed a significant increase in expression upon the miR-21 sponging through CM21D. Additionally, sponging of miR-21 by CM21D enhanced the growth rate of CHO cells. • Productivity and growth rate were increased in CHO cells expressing recombinant hrs-ACE2 protein after CM21D knocking in. Graphical abstract

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Engineering Saccharomyces cerevisiae for efficient production of recombinant proteins

Yang,

Song,

Wang

et al. 2024

Engineering Microbiology

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Recent developments in miRNA based recombinant protein expression in CHO

Cited by 5 publications

References 77 publications

Overexpression of miR-32 in Chinese hamster ovary cells increases production of Fc-fusion protein

Overexpression of miR-32 in Chinese hamster ovary cells increases production of Fc-fusion protein

CHO cell engineering via targeted integration of circular miR-21 decoy using CRISPR/RMCE hybrid system

Engineering Saccharomyces cerevisiae for efficient production of recombinant proteins

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