Chinese hamster ovary (CHO) cells are the predominant host for producing complex recombinant therapeutic proteins. N6-methyladenosine (m6A) regulates recombinant protein production in CHO cells by influencing RNA stability and translation; however, the role of the m6A demethylase ALKBH5 remains poorly characterized in this context. Here, ALKBH5 overexpression increased the titers of three recombinant products by up to 2.87-fold, enhanced CHO cell proliferation, and reduced oxidative stress. Co-immunoprecipitation supported an association between ALKBH5 and ITGB5, and structure-guided mutagenesis identified Tyr205 of ITGB5 as functionally important for this association and ITGB5 protein stability. ALKBH5 overexpression did not measurably alter ITGB5 mRNA stability or m6A enrichment, supporting a mechanism that does not involve detectable m6A changes on the ITGB5 transcript, while not excluding m6A-dependent effects on other targets. FAK inhibition attenuated but did not abolish the production advantage, indicating that FAK signaling contributes to, but may not fully account for, the ALKBH5−ITGB5-associated phenotype. Metabolic and redox measurements further linked ALKBH5 overexpression to increased antioxidant capacity and ATP abundance, together with altered nutrient consumption and by-product formation. These findings identify ALKBH5 as a candidate host-cell engineering target for improving recombinant protein production and support further validation under industry-relevant fed-batch conditions.
Chinese hamster ovary (CHO) cells represent the dominant host system for the production of recombinant therapeutic proteins. In recent decades, extensive research has focused on process/media optimization and cell line engineering to improve both the productivity and quality of biopharmaceutical proteins produced in CHO cells. Nevertheless, the inherent complexity of biological pathways and the heterogeneous cellular responses to different environmental conditions have posed substantial challenges to traditional methodologies. Recent advances in omics technologies have enabled comprehensive characterization of CHO cell physiology, providing multidimensional molecular and phenotypic insights that facilitate the enhancement of recombinant protein production. This review first summarizes the methodologies and advances in CHO omics research, including genomics, transcriptomics, proteomics, metabolomics, and epigenomics. It then examines contemporary approaches to integrate and analyze multi-omics data in CHO cells. The review further elucidates how these multi-omics datasets can be strategically applied across various developmental stages, including cell line selection, genetic engineering, expression vector design, and bioprocess optimization. Finally, we explore the transformative potential of integrating multi-omics with artificial intelligence and discuss promising future research directions in CHO cell studies. These emerging paradigms offer novel opportunities for data-driven cell engineering and bioprocess optimization in CHO-based biomanufacturing.
Yuan Shen, Lei Shi, Xi Zhang et al.· Bioresources and Bioprocessi...· 0 citations
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