LAUSR

Chinese hamster ovary (CHO) cells are currently the primary host cell lines used in biotherapeutic manufacturing of monoclonal antibodies (mAbs) and other biopharmaceuticals. Cellular energy metabolism and endoplasmic reticulum (ER) stress are known to greatly impact cell growth, viability, and specific productivity of a biotherapeutic. However, specific molecular mechanisms responsible for these changes are not fully understood. In a previous paper, we employed multi-omics profiling to investigate the impact of a reduction in cysteine (Cys) feed concentration in a fed batch process and found that disruption of the redox balance in the CHO cells led to a substantial decline in cell viability and titer. Here, we expand our multi-omics findings and explore the significant impact redox imbalance has on ER stress, mitochondrial homeostasis, and lipid metabolism. The reduced Cys feed led to intracellular Cys depletion which activated the amino acid response (AAR), increased mitochondrial stress and initiated gluconeogenesis. Multi-omics analysis revealed that together, ER stress and AAR signaling shifted the cellular energy metabolism to rely primarily on anaplerotic reactions, consuming amino acids and producing lactate, in order to maintain energy generation. Furthermore, we demonstrate the pathways in which this shift in metabolism led to a substantial decline in specific productivity and altered mAb glycosylation. Through this work, meaningful bioprocess markers are identified, and targets for genetic engineering for enhanced productivity are suggested. This article is protected by copyright. All rights reserved.