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Chinese hamster ovary (CHO) cells are the preferred system for expression of therapeutic proteins and the majority of all biotherapeutics are being expressed by these cell lines. CHO expression systems are readily scalable, resistant to human adventitious agents, and have desirable post‐translational modifications, such as glycosylation. Regardless, drug development as a whole is a very costly, complicated, and time‐consuming process. Therefore, any improvements that result in reducing timelines are valuable and can provide patients with life‐saving drugs earlier. Here we report an effective method (termed SPEED‐MODE, herein) to speed up the Cell line Development (CLD) process in a targeted integration (TI) CHO CLD system. Our findings show that (1) earlier single cell cloning (SCC) of transfection pools, (2) speeding up initial titer screening turnaround time, (3) starting suspension adaptation of cultures sooner, and (4) maximizing the time CHO cultures spend in the exponential growth phase can reduce CLD timelines from ~4 to ~3 months. Interestingly, SPEED‐MODE timelines closely match the theoretical minimum timeline for CHO CLD assuming that CHO cell division is the rate limiting factor. Clones obtained from SPEED‐MODE CLD yielded comparable titer and product quality to those obtained via a standard CLD process. Hence, SPEED‐MODE CLD is advantageous for manufacturing biotherapeutics in an industrial setting as it can significantly reduce CLD timelines without compromising titer or product quality. [ABSTRACT FROM AUTHOR]

Chinese hamster ovary (CHO) cells are the predominant host of choice for recombinant monoclonal antibody (mAb) expression. Recent advancements in gene editing technology have enabled engineering new CHO hosts with higher growth, viability, or productivity. One approach involved knock out (KO) of BCAT1 gene, which codes for the first enzyme in the branched chain amino acid (BCAA) catabolism pathway; BCAT1 KO reduced accumulation of growth inhibitory short chain fatty acid (SCFA) byproducts and improved culture growth and titer when used in conjunction with high‐end pH‐controlled delivery of glucose (HiPDOG) technology and SCFA supplementation during production. Accumulation of SCFAs in the culture media is critical for metabolic shift toward higher specific productivity and hence titer. Here we describe knocking out BCKDHa/b genes (2XKO), which act downstream of the BCAT1, in a BAX/BAK KO CHO host cell line background to reduce accumulation of growth‐inhibitory molecules in culture. Evaluation of the new 4XKO CHO cell lines in fed‐batch production cultures (without HiPDOG) revealed that partial KO of BCKDHa/b genes in an apoptosis‐resistant (BAX/BAK KO) background can achieve higher viabilities and mAb titers. This was evident when SCFAs were added to boost productivity as such additives negatively impacted culture viability in the WT but not BAX/BAK KO cells during batch production. Altogether, our findings suggest that SCFA addbacks can significantly increase productivity and mAb titers in the context of apoptosis‐attenuated CHO cells with partial KO of BCAA genes. Such engineered CHO hosts can offer productivity advantages for expressing biotherapeutics in an industrial setting. [ABSTRACT FROM AUTHOR]