Insight into the roles of tyrosine on rCHO cell performance in fed-batch cultures
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Tyrosine (Tyr), as one of the least soluble amino acids, is essential to monoclonal antibody (mAb) production in recombinant Chinese hamster ovary (rCHO) cell cultures since its roles on maintaining the specific productivity (qmAb) and avoiding Tyr sequence variants. To understand the effects of Tyr on cell performance and its underlying mechanisms, rCHO cell–producing mAbs were cultivated at various cumulative Tyr addition concentrations (0.6 to 5.5 mM) in fed-batch processes. Low Tyr concentrations gave a much lower peak viable cell density (VCD) during the growth phase and also induced rapid cell death and pH decrease during the production phase, resulting in a low efficient fed-batch process. Autophagy was initiated following the inhibition of mTOR under the Tyr starvation condition. Excessive autophagy subsequently induced autophagic cell death, which was found as the major type of cell death in this study. Additionally, the results obtained here demonstrate that the decrease in culture pH under the Tyr starvation condition was associated with the autophagy and such pH drop might be attributed to the lysosome acidification and cell lysis.
KeywordsChinese hamster ovary cells Tyrosine Cell death Autophagy Culture pH
This work was supported by the Fundamental Research Funds for the Central Universities (No. 22221818014).
Compliance with ethical standards
This article does not contain any studies with human participants or animals performed by any of the authors.
Conflict of interest
The authors declare that they have no conflict of interest.
- Aggarwal SR (2014) What's fueling the biotech engine—2012 to 2013. Nat Biotechnol 32(1):32–39Google Scholar
- Castilho L, Maroes A, Augusto E, Butler M (2008) Animal Cell Technology: From Biopharmaceuticals to Gene Therapy. Taylor & Francis Group, New YorkGoogle Scholar
- Figueroa B, Ailor E, Osborne D, Hardwick JM, Reff M, Betenbaugh MJ (2007) Enhanced cell culture performance using inducible anti-apoptotic genes E1B-19K and Aven in the production of a monoclonal antibody with chinese hamster ovary cells. Biotechnol Bioeng 97(4):877–892. https://doi.org/10.1002/bit.21222 Google Scholar
- Fu T, Zhang C, Jing Y, Jiang C, Li Z, Wang S, Ma K, Zhang D, Hou S, Dai J, Kou G, Wang H (2016) Regulation of cell growth and apoptosis through lactate dehydrogenase C over-expression in Chinese hamster ovary cells. Appl Microbiol Biotechnol 100(11):5007–5016. https://doi.org/10.1007/s00253-016-7348-4 Google Scholar
- Hansen HA, Emborg C (1994) Extra- and intracellular amino acid concentrations in continuous Chinese hamster ovary cell culture. Appl Microbiol Biotechnol 41(5):560–564Google Scholar
- Jayapal KR, Wlaschin KF, Hu WS, Yap MGS (2007) Recombinant protein therapeutics from CHO cells - 20 years and counting. Chem Eng Prog 103(10):40–47Google Scholar
- Kim NS, Lee GM (2002) Response of recombinant Chinese hamster ovary cells to hyperosmotic pressure: effect of Bcl-2 overexpression. J Biotechnol 95(3):237–248Google Scholar
- Kroemer G, Galluzzi L, Vandenabeele P, Abrams J, Alnemri ES, Baehrecke EH, Blagosklonny MV, El-Deiry WS, Golstein P, Green DR, Hengartner M, Knight RA, Kumar S, Lipton SA, Malorni W, Nunez G, Peter ME, Tschopp J, Yuan J, Piacentini M, Zhivotovsky B, Melino G, Nomenclature Committee on Cell D (2009) Classification of cell death: recommendations of the Nomenclature Committee on Cell Death 2009. Cell Death Differ 16(1):3–11. https://doi.org/10.1038/cdd.2008.150 Google Scholar
- Meneses-Acosta A, Mendonca RZ, Merchant H, Covarrubias L, Ramirez OT (2001) Comparative characterization of cell death between Sf9 insect cells and hybridoma cultures. Biotechnol Bioeng 72(4):441–457. https://doi.org/10.1002/1097-0290(20000220)72:4<441::Aid-Bit1006>3.0.Co;2-3 Google Scholar
- Muthing J, Kemminer SE, Conradt HS, Sagi D, Nimtz M, Karst U, Peter-Katalinic J (2003) Effects of buffering conditions and culture pH on production rates and glycosylation of clinical phase I anti-melanoma mouse IgG3 monoclonal antibody R24. Biotechnol Bioeng 83(3):321–334. https://doi.org/10.1002/bit.10673 Google Scholar
- Sharma A, Sharma R, Singh SP, Khinchi M (2017) A Brief Review on Apoptosis. Asian J Pharm Res Dev:1–10Google Scholar
- Trummer E, Fauland K, Seidinger S, Schriebl K, Lattenmayer C, Kunert R, Vorauer-Uhl K, Weik R, Borth N, Katinger H, Muller D (2006) Process parameter shifting: Part I. Effect of DOT, pH, and temperature on the performance of Epo-Fc expressing CHO cells cultivated in controlled batch bioreactors. Biotechnol Bioeng 94(6):1033–1044. https://doi.org/10.1002/bit.21013 Google Scholar
- von Hagen J, Hecklau C, Seibel R, Pering S, Schnellbaecher A, Wehsling M, Eichhorn T (2017) Simplification of Fed-Batch Processes with a Single-Feed Strategy. BioProcess InternationalGoogle Scholar
- Yang JD, Lu C, Stasny B, Henley J, Guinto W, Gonzalez C, Gleason J, Fung M, Collopy B, Benjamino M, Gangi J, Hanson M, Ille E (2007) Fed-batch bioreactor process scale-up from 3-L to 2,500-L scale for monoclonal antibody production from cell culture. Biotechnol Bioeng 98(1):141–154. https://doi.org/10.1002/bit.21413 Google Scholar
- Yu L, Lenardo MJ, Baehrecke EH (2004) Autophagy and caspases: a new cell death program. Cell Cycle 3(9):1122–1124Google Scholar
- Zanghi JA, Schmelzer AE, Mendoza TP, Knop RH, Miller WM (1999) Bicarbonate concentration and osmolality are key determinants in the inhibition of CHO cell polysialylation under elevated pCO2 or pH. Biotechnol Bioeng 65(2):182–191Google Scholar
- Zhang X, Tang H, Sun Y-T, Liu X, Tan W-S, Fan L (2015) Elucidating the effects of arginine and lysine on a monoclonal antibody C-terminal lysine variation in CHO cell cultures. Appl Microbiol Biotechnol 99(16):6643–6652. https://doi.org/10.1007/s00253-015-6617-y
- Zhu MM, Goyal A, Rank DL, Gupta SK, Boom TV, Lee SS (2005) Effects of elevated pCO2 and osmolality on growth of CHO cells and production of antibody-Fusion Protein B1: a case study. Biotechnol Prog 21(1):70–77Google Scholar
- Zustiak MP, Pollack JK, Marten MR, Betenbaugh MJ (2008) Feast or famine: autophagy control and engineering in eukaryotic cell culture. Curr Opin Biotechnol 19(5):518–526. https://doi.org/10.1016/j.copbio.2008.07.007