Comparison of l‐tyrosine containing dipeptides reveals maximum ATP availability for l‐prolyl‐l‐tyrosine in CHO cells

Abstract

Increasing markets for biopharmaceuticals, including monoclonal antibodies, have triggered a permanent need for bioprocess optimization. Biochemical engineering approaches often include the optimization of basal and feed media to improve productivities of Chinese hamster ovary (CHO) cell cultures. Often, l‐tyrosine is added as dipeptide to deal with its poor solubility at neutral pH. Showcasing IgG1 production with CHO cells, we investigated the supplementation of three l‐tyrosine (TYR, Y) containing dipeptides: glycyl‐l‐tyrosine (GY), l‐tyrosyl‐l‐valine (YV), and l‐prolyl‐l‐tyrosine (PY). While GY and YV led to almost no phenotypic and metabolic differences compared to reference samples, PY significantly amplified TYR uptake thus maximizing related catabolic activity. Consequently, ATP formation was roughly four times higher upon PY application than in reference samples.

Exploring metabolic effects of dipeptide feed media on CHO cell cultures by in silico model-guided flux analysis

“…Additionally, the 0.5 × GY condition achieved the highest final titer, while the 0.25 × , 1.0 × , and 2.0 × conditions showed relatively lower final titers of 25.7%, 20.5%, and 25.24%, respectively. Most conditions displayed consistent Qp, except for the 0.5 × GY condition, indicating a strong relationship between the higher SGR observed during the exponential phase and enhanced productivity, which is in agreement with previous observations (Verhagen et al 2020 ). In fact, the most effective condition, 0.5 × GY, showed significantly reduced levels of toxic byproducts such as lactate and ammonia throughout the culture duration, fostering a more favorable environment conducive to cell growth.…”

“…This adaptation aligns with a key market trend, meeting the demands of biologics biomanufacturing in terms of the concepts of process simplification and intensification. In this regard, various Tyr-containing dipeptides were investigated to demonstrate their beneficial effects in reducing the formation of toxic byproducts and increasing cell viability and titer (Kang et al 2012 ; Sánchez-Kopper et al 2016 ; Verhagen et al 2020 ). However, the supplementation of Tyr-containing dipeptides in the media increases production costs primarily due to the high expense involved in synthesizing short peptides.…”

Compartment-specific metabolome labeling enables the identification of subcellular fluxes that may serve as promising metabolic engineering targets in CHO cells

“…To date, the compartment-specific analytical approach has shown its suitability for multiple metabolomic studies investigating CHO cells under in vivo-like conditions [8,15,[24][25][26][27][28][29][30]. The latter is enabled by fast and standardized metabolism inactivation.…”

Compartment‐specific 13C metabolic flux analysis reveals boosted NADPH availability coinciding with increased cell‐specific productivity for IgG1 producing CHO cells after MTA treatment

“…However, a beneficial link to heterologous protein formation exists. Flux balance analysis using the simplified metabolic model [62] published by Verhagen et al (2020c) considered that the maximum NADPH supply should be supported by high cytosolic malic enzyme flux and high oxidative PPP activity (Supplementary Material 3). Interestingly, MTA treatment provides an optimal flux distribution to maximize the NADPH supply.…”