Monoclonal antibodies (mAbs) are a key component of the pharmaceutical industry, accounting for half of the top selling pharmaceuticals in 2016 with total worldwide sales projected to increase from $75 billion in 2013 to $125 billion in 2020. Historically, Chinese Hamster Ovary (CHO) cells have been used extensively for a number of reasons, including their secretory nature and ability to grow in serum-free media. However, CHO cell metabolism is not naturally efficient, producing large amounts of lactate and ammonia that can hinder growth and affect the quality of the final product. MAbs are large and energetically demanding to express; therefore, high-producing CHO cell lines that are metabolically efficient are needed to meet the increasing demand for mAb production. Our previous studies have identified that high-producing cell lines possess a different metabolic phenotype when compared to low-producing cell lines. In particular, it was found that mAb production is correlated to lactate consumption and elevated TCA cycle flux. We hypothesized that increasing flux through the TCA cycle could lead to increased mAb titers. In our current studies, we have overexpressed genes that are related to mitochondrial metabolism in order to increase TCA cycle flux and studied the response to these metabolic changes. In cell lines that overexpress target metabolic genes of interest, we have observed increased oxygen consumption, indicating increased mitochondrial metabolism, as well as increased mAb specific productivity. We have also carried out <sup>13</sup>C metabolic flux analysis (MFA) to quantify how these genes affect metabolic fluxes. This presentation will highlight how cell lines can be systematically engineered to increase metabolic efficiency, which has broad industrial applications for production in mammalian cell culture.

Metabolic engineering of CHO cells for increased mAb production

2019 Spring National Meeting

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