Ouantitative Proteomic Analysis of Cellular Responses to a Designed Amino Acid Feed in a Monoclonal Antibody Producing Chinese Hamster Ovary Cell Line

Abstract

Background:Chinese hamster ovary (CHO) cell line is considered as the most common cell line in the biopharmaceutical industry because of its capability in performing efficient post-translational modifications and producing the recombinant proteins, which are similar to natural human proteins. The optimization of the upstream process via different feed strategies has a great impact on the target molecule expression and yield.Methods:To determine and understand the molecular events beneath the feed effects on the CHO cell, a label-free quantitative proteomic analysis was applied. The proteome changes followed by the addition of a designed amino acid feed to the monoclonal antibody producing CHO cell line culture medium were investigated.Results:The glutathione synthesis, the negative regulation of the programmed cell death, proteasomal catabolic process, and the endosomal transport pathway were up-regulated in the group fed with a designed amino acid feed compared to the control group.Conclusion:Our findings could be helpful to identify new targets for metabolic engineering.

Nutrient supplementation strategy improves cell concentration and longevity, monoclonal antibody production and lactate metabolism of Chinese hamster ovary cells

“…The development of the culture medium and feeds and their uses in bioprocesses are of the utmost importance for the cell culture experiments, as well as for biopharmaceutical work. In fact, the optimization of the composition of culture media and feeds has a direct and positive impact on the duration of the bioprocesses [7][8][9][10][11][12][13][14][15][16][17][18], productivity [7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22] and purification and quality of proteins [9][10][11]13,15,17,22], factors that ultimately impact on lower RP costs. Feeds have been designed to replenish depleted nutrients in basal medium, alleviating different cellular stresses and supporting the cell growth and RP production; they can be classified into those of general use [7,8,10,12,[15][16][17]19,20] or those specifically developed for certain clones…”

“…Feeds have been designed to replenish depleted nutrients in basal medium, alleviating different cellular stresses and supporting the cell growth and RP production; they can be classified into those of general use [7,8,10,12,[15][16][17]19,20] or those specifically developed for certain clones [3,18,21,22]. Although these supplements are usually added to culture media during the course of the bioprocess (fed-batch culture) [3,[10][11][12][14][15][16][18][19][20][21][22], they can be used at the beginning of the culture (batch culture) as well [3,7,8,13,17]. Feeds can be classified as chemically defined nutrient cocktails (CDF) or concentrated hydrolyzates; in the first case, their exact composition is generally not available excluding some exceptions [3,18,21,22], and in the second one, they remain as black boxes for their composition.…”

“…Although these supplements are usually added to culture media during the course of the bioprocess (fed-batch culture) [3,[10][11][12][14][15][16][18][19][20][21][22], they can be used at the beginning of the culture (batch culture) as well [3,7,8,13,17]. Feeds can be classified as chemically defined nutrient cocktails (CDF) or concentrated hydrolyzates; in the first case, their exact composition is generally not available excluding some exceptions [3,18,21,22], and in the second one, they remain as black boxes for their composition. They have demonstrated a positive impact on cell concentration, viability, product titer, specific and volumetric productivity and metabolic behavior during the production of different RPs such as monoclonal antibodies (MAbs) [8][9][10][11][12][13][15][16][17][18][19][20][21][22][23], blood clotting factors [14] and fusion proteins [3], or in non-transformed cells [7,23], without significant changes in protein quality [13,17,…”

“…Feeds can be classified as chemically defined nutrient cocktails (CDF) or concentrated hydrolyzates; in the first case, their exact composition is generally not available excluding some exceptions [3,18,21,22], and in the second one, they remain as black boxes for their composition. They have demonstrated a positive impact on cell concentration, viability, product titer, specific and volumetric productivity and metabolic behavior during the production of different RPs such as monoclonal antibodies (MAbs) [8][9][10][11][12][13][15][16][17][18][19][20][21][22][23], blood clotting factors [14] and fusion proteins [3], or in non-transformed cells [7,23], without significant changes in protein quality [13,17,20,22]. Although many efforts have focused on the development of supplements that can be used in several cell lines, the nutritional requirements are specific to each of them [9,12,13,15,21,22,24,25], so the benefits of feeds should be evaluated on a case-by-case basis.…”

A metabolic network-based approach for developing feeding strategies for CHO cells to increase monoclonal antibody production

“…In addition to exchange reactions, some internal metabolic reactions were found to influence mAb production. Using the proteomics data of CHO cell lines producing mAb in our lab (24), the enzyme expression levels of these metabolic reactions in high production cell line were compared to low production state. The predicted changes in the rates of these reactions in the FVSEOF list were in accordance with the enzyme expression changes (data not shown).…”