Advancements in CHO metabolomics: techniques, current state and evolving methodologies.

Investigating the metabolic behaviour of different cellular phenotypes, i.e., good/bad grower and/or producer, in production culture is important to identify the key metabolite(s)/pathway(s) that regulate cell growth and/or recombinant protein production to improve the overall yield. Currently, LC-MS, GC-MS and NMR are the most used and advanced technologies for investigating the metabolome. Although contributed significantly in the domain, each technique has its own biasness towards specific metabolites or class of metabolites due to various reasons including variability in the concept of working, sample preparation, metabolite-extraction methods, metabolite identification tools, and databases. As a result, the application of appropriate analytical technique(s) is very critical. This review provides a state-of-the-art technological insights and overview of metabolic mechanisms involved in regulation of cell growth and/or recombinant protein production for improving yield from CHO cultures. In this review, the advancements in CHO metabolomics over the last 10 years are traced based on a bibliometric analysis of previous publications and discussed. With the technical advancement in the domain of LC-MS, GC-MS and NMR, metabolites of glycolytic and nucleotide biosynthesis pathway (glucose, fructose, pyruvate and phenylalanine, threonine, tryptophan, arginine, valine, asparagine, and serine, etc.) were observed to be upregulated in exponential-phase thereby potentially associated with cell growth regulation, whereas metabolites/intermediates of TCA, oxidative phosphorylation (aspartate, glutamate, succinate, malate, fumarate and citrate), intracellular NAD+/NADH ratio, and glutathione metabolic pathways were observed to be upregulated in stationary-phase and hence potentially associated with increased cell-specific productivity in CHO bioprocess. Moreover, each of technique has its own bias towards metabolite identification, indicating their complementarity, along with a number of critical gaps in the CHO metabolomics pipeline and hence first time discussed here to identify their potential remedies. This knowledge may help in future study designs to improve the metabolomic coverage facilitating identification of the metabolites/pathways which might get missed otherwise and explore the full potential of metabolomics for improving the CHO bioprocess performances.