Preventing pyruvate kinase muscle expression in Chinese hamster ovary cells curbs lactogenic behavior by altering glycolysis, gating pyruvate generation, and increasing pyruvate flux into the TCA cycle,Biotechnology Progress

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Preventing pyruvate kinase muscle expression in Chinese hamster ovary cells curbs lactogenic behavior by altering glycolysis, gating pyruvate generation, and increasing pyruvate flux into the TCA cycle
Biotechnology Progress ( IF 2.9 ) Pub Date : 2021-07-20 , DOI: 10.1002/btpr.3193
Danming Tang ₁ , Wendy Sandoval ₂ , Peter Liu ₂ , Cynthia Lam ₁ , Brad Snedecor ₁ , Shahram Misaghi ₁

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Deletion of the pyruvate kinase muscle (PKM) gene, which is involved in conversion of phosphoenolpyruvate to pyruvate, has been shown to curb lactogenic behavior in Chinese hamster ovary (CHO) cells. This study describes the generation of pyruvate kinase muscle isoforms 1 and 2 knockout (PKM-KO) and pyruvate kinase muscle isoform-1 knockout (PKM1-KO) CHO host cells to understand metabolic shifts that reduce lactate secretion in these cells. Glucose and amino acids uptake levels in wild-type (WT), PKM-KO, and PKM1-KO stable cell lines, expressing two different antibodies, were analyzed in 14-day fed-batch production assays using different vessels. PKM-KO and PKM1-KO cells consumed more glucose per cell, altered amino acids metabolism, had higher flux of pyruvate into the tricarboxylic acid (TCA) cycle, and as previously shown reduced lactate secretion levels compared with the WT cells. Additionally, both PKM-KO and PKM1-KO cells had higher specific productivity and lower cell growth rates compared with the WT cells. Our findings suggest that rewiring the flux of pyruvate to the TCA cycle by deletion of PKM or PKM1 reduced cell growth and increased specific productivity in CHO cells. Overall, PKM1-KO cells had similar product quality and comparable or better titers relative to the WT cells, hence, targeted deletion of this isoform for curbing lactogenic behavior in CHO cells is suggested.

更新日期：2021-07-20

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