Advertisement

The effect of hyperosmolality application time on production, quality, and biopotency of monoclonal antibodies produced in CHO cell fed-batch and perfusion cultures

  • Jinyan Qin
  • Xiang Wu
  • Zhigang Xia
  • Zheng Huang
  • Ying Zhang
  • Yanchao Wang
  • Qiang Fu
  • Chen Zheng
Biotechnological products and process engineering

Abstract

Hyperosmolality has been commonly investigated due to its effects on the production and quality characteristics of monoclonal antibodies (mAbs) produced in CHO cell fed-batch cultures. However, the application of hyperosmolality at different times and its effect on biopotency have seldom been researched, especially in perfusion culture. In our study, different degrees of hyperosmolality induced by sodium chloride were investigated in anti-IgE rCHO cell fed-batch cultures and anti-CD52 rCHO cell perfusion cultures during the initial and stable phases. The results showed that the initial hyperosmolality group (IHG) in fed-batch and early phase of perfusion cultures exhibited significant suppression of the viable cell density yet an enhancement in specific productivity, whereas the stable hyperosmolality group (SHG) achieved higher mAb production in both fed-batch and perfusion cultures. Additionally, the SHG produced less aggregates and acidic charge variants than IHG in fed-batch culture, which differed from perfusion cultures. However, the contents of non-glycosylation heavy chain (NGHC) and man5 were higher in SHG than in IHG in fed-batch cultures at plus 60 and 120 mOsm/kg, which was similar to perfusion cultures. Furthermore, the biopotency in the IHG was higher than in the SHG at plus 60 and 120 mOsm/kg in fed-batch cultures, which is similar to complement-dependent cytotoxicity (CDC) efficacy in perfusion cultures. The biopotency of all group was acceptable, except FI3. Thus, the study shows that hyperosmolality at a certain level could be beneficial for both mAb production, quality and biopotency, which could play an important role in process development for commercial production.

Keywords

Chinese hamster ovary (CHO) Hyperosmolality Fed-batch Perfusion Biopotency 

Notes

Acknowledgements

This project was sponsored by Shanghai Taiyin Biotech Co. Ltd. We thank everyone who helped with this research.

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.

References

  1. Ashton DS, Beddell CR, Cooper DJ, Craig SJ, Oliver RW, Smith MA (1995) Mass spectrometry of the humanized monoclonal antibody CAMPATH 1H. Anal Chem 67(5):835–842CrossRefGoogle Scholar
  2. Bertrand V, Vogg S, Villiger TK, Stettler M, Broly H, Soos M, Morbidelli M (2018) Proteomic analysis of micro-scale bioreactors as scale-down model for a mAb producing CHO industrial fed-batch platform. J Biotechnol 279:27–36.  https://doi.org/10.1016/j.jbiotec.2018.04.015 CrossRefPubMedGoogle Scholar
  3. Boswell CA, Tesar DB, Mukhyala K, Theil F-P, Fielder PJ, Khawli LA (2010) Effects of charge on antibody tissue distribution and pharmacokinetics. Bioconjug Chem 21(12):2153–2163.  https://doi.org/10.1021/bc100261d CrossRefPubMedGoogle Scholar
  4. Chen P, Harcum S (2006) Effects of elevated ammonium on glycosylation gene expression in CHO cells. Metab Eng 8(2):123–132.  https://doi.org/10.1016/j.ymben.2005.10.002 CrossRefPubMedGoogle Scholar
  5. Cherlet M, Marc A (1999) Hybridoma cell behaviour in continuous culture under hyperosmotic stress. Cytotechnology 29(1):71–84.  https://doi.org/10.1023/a:1008014909474 CrossRefPubMedPubMedCentralGoogle Scholar
  6. deZengotita VM, Kimura R, Miller WM (1998) Effects of CO2 and osmolality on hybridoma cells: growth, metabolism and monoclonal antibody production. Cytotechnology 28(1):213–227.  https://doi.org/10.1023/a:1008010605287 CrossRefPubMedPubMedCentralGoogle Scholar
  7. deZengotita VM, Schmelzer AE, Miller WM (2002) Characterization of hybridoma cell responses to elevated pCO2 and osmolality: intracellular pH, cell size, apoptosis, and metabolism. Biotechnol Bioeng 77(4):369–380.  https://doi.org/10.1002/bit.10176 CrossRefPubMedGoogle Scholar
  8. Elgundi Z, Reslan M, Cruz E, Sifniotis V, Kayser V (2017) The state-of-play and future of antibody therapeutics. Adv Drug Deliv Rev 122:2–19.  https://doi.org/10.1016/j.addr.2016.11.004 CrossRefPubMedGoogle Scholar
  9. Franco R, Daniela G, Fabrizio M, Ilaria G, Detlev H (1999) Influence of osmolarity and pH increase to achieve a reduction of monoclonal antibodies aggregates in a production process. Cytotechnology 29(1):11–25.  https://doi.org/10.1023/a:1008075423609 CrossRefPubMedPubMedCentralGoogle Scholar
  10. Grilo AL, Mantalaris A (2018) The increasingly human and profitable monoclonal antibody market. Trends Biotechnol.  https://doi.org/10.1016/j.tibtech.2018.05.014
  11. Han YK, Kim YG, Kim JY, Lee GM (2010) Hyperosmotic stress induces autophagy and apoptosis in recombinant Chinese hamster ovary cell culture. Biotechnol Bioeng 105(6):1187–1192.  https://doi.org/10.1002/bit.22643 CrossRefPubMedGoogle Scholar
  12. Harris RJ, Kabakoff B, Macchi FD, Shen FJ, Kwong M, Andya JD, Shire SJ, Bjork N, Totpal K, Chen AB (2001) Identification of multiple sources of charge heterogeneity in a recombinant antibody. J Chromatogr B 752(2):233–245.  https://doi.org/10.1016/S0378-4347(00)00548-X CrossRefGoogle Scholar
  13. Heidemann R, Zhang C, Qi H, Larrick Rule J, Rozales C, Park S, Chuppa S, Ray M, Michaels J, Konstantinov K, Naveh D (2000) The use of peptones as medium additives for the production of a recombinant therapeutic protein in high density perfusion cultures of mammalian cells. Cytotechnology 32(2):157–167.  https://doi.org/10.1023/a:1008196521213 CrossRefPubMedPubMedCentralGoogle Scholar
  14. Hintersteiner B, Lingg N, Zhang P, Woen S, Hoi KM, Stranner S, Wiederkum S, Mutschlechner O, Schuster M, Loibner H, Jungbauer A (2016) Charge heterogeneity: basic antibody charge variants with increased binding to fc receptors. mAbs 8(8):1548–1560.  https://doi.org/10.1080/19420862.2016.1225642 CrossRefPubMedPubMedCentralGoogle Scholar
  15. Hodoniczky J, Zheng YZ, James DC (2005) Control of recombinant monoclonal antibody effector functions by Fc N-glycan remodeling in vitro. Biotechnol Prog 21(6):1644–1652.  https://doi.org/10.1021/bp050228w CrossRefPubMedGoogle Scholar
  16. Hong JK, Lee SM, Kim KY, Lee GM (2014) Effect of sodium butyrate on the assembly, charge variants, and galactosylation of antibody produced in recombinant Chinese hamster ovary cells. Appl Microbiol Biotechnol 98(12):5417–5425.  https://doi.org/10.1007/s00253-014-5596-8 CrossRefPubMedGoogle Scholar
  17. Hossler P, Khattak SF, Li ZJ (2009) Optimal and consistent protein glycosylation in mammalian cell culture. Glycobiology 19(9):936–949.  https://doi.org/10.1093/glycob/cwp079 CrossRefPubMedGoogle Scholar
  18. Jefferis R (2009) Glycosylation as a strategy to improve antibody-based therapeutics. Nat Rev Drug Discov 8(3):226–234.  https://doi.org/10.1038/nrd2804 CrossRefPubMedGoogle Scholar
  19. Kamachi Y, Omasa T (2018) Development of hyper osmotic resistant CHO host cells for enhanced antibody production. J Biosci Bioeng 125(4):470–478.  https://doi.org/10.1016/j.jbiosc.2017.11.002 CrossRefPubMedGoogle Scholar
  20. Kanda Y, Yamada T, Mori K, Okazaki A, Inoue M, Kitajima-Miyama K, Kuni-Kamochi R, Nakano R, Yano K, Kakita S, Shitara K, Satoh M (2007) Comparison of biological activity among nonfucosylated therapeutic IgG1 antibodies with three different N-linked fc oligosaccharides: the high-mannose, hybrid, and complex types. Glycobilogy 17(1):104–118.  https://doi.org/10.1093/glycob/cwl057 CrossRefGoogle Scholar
  21. Konno Y, Kobayashi Y, Takahashi K, Takahashi E, Sakae S, Wakitani M, Yamano K, Suzawa T, Yano K, Ohta T, Koike M, Wakamatsu K, Hosoi S (2012) Fucose content of monoclonal antibodies can be controlled by culture medium osmolality for high antibody-dependent cellular cytotoxicity. Cytotechnology 64(3):249–265.  https://doi.org/10.1007/s10616-011-9377-2 CrossRefPubMedGoogle Scholar
  22. Leblanc Y, Ramon C, Bihoreau N, Chevreux G (2017) Charge variants characterization of a monoclonal antibody by ion exchange chromatography coupled on-line to native mass spectrometry: case study after a long-term storage at +5 degrees C. J Chromatogr B 1048:130–139.  https://doi.org/10.1016/j.jchromb.2017.02.017 CrossRefGoogle Scholar
  23. Lee JH, Jeong YR, Kim YG, Lee GM (2017) Understanding of decreased sialylation of fc-fusion protein in hyperosmotic recombinant Chinese hamster ovary cell culture: N-glycosylation gene expression and N-linked glycan antennary profile. Biotechnol Bioeng 114:1721–1732CrossRefGoogle Scholar
  24. Mutsumi Takagi TM, Yoshida T (2001) Effects of shifts up and down in osmotic pressure on production of tissue plasminogen activator by Chinese hamster ovary cells in suspension. J Biosci Bioeng 91(5):509–514.  https://doi.org/10.1016/S1389-1723(01)80282-6 CrossRefGoogle Scholar
  25. Nasseri SS, Ghaffari N, Braasch K, Jardon MA, Butler M, Kennard M, Gopaluni B, Piret JM (2014) Increased CHO cell fed-batch monoclonal antibody production using the autophagy inhibitor 3-MA or gradually increasing osmolality. Biochem Eng J 91:37–45.  https://doi.org/10.1016/j.bej.2014.06.027 CrossRefGoogle Scholar
  26. Noh SM, Sathyamurthy M, Lee GM (2013) Development of recombinant Chinese hamster ovary cell lines for therapeutic protein production. Curr Opin Chem Eng 2(4):391–397.  https://doi.org/10.1016/j.coche.2013.08.002 CrossRefGoogle Scholar
  27. Oh SKW, Vig P, Chua F, Teo WK, Yap MGS (1993) Substantial overproduction of antibodies by applying osmotic pressure and sodium butyrate. Biotechnol Bioeng 42(5):601–610.  https://doi.org/10.1002/bit.260420508 CrossRefPubMedGoogle Scholar
  28. Parsons TB, Struwe WB, Gault J, Yamamoto K, Taylor TA, Raj R, Wals K, Mohammed S, Robinson CV, Benesch JL, Davis BG (2016) Optimal synthetic glycosylation of a therapeutic antibody. Angew Chem 55(7):2361–2367.  https://doi.org/10.1002/anie.201508723 CrossRefGoogle Scholar
  29. Pollock J, Ho SV, Farid SS (2013) Fed-batch and perfusion culture processes: economic, environmental, and operational feasibility under uncertainty. Biotechnol Bioeng 110(1):206–219.  https://doi.org/10.1002/bit.24608 CrossRefPubMedGoogle Scholar
  30. Shen D, Kiehl TR, Khattak SF, Li ZJ, He A, Kayne PS, Patel V, Neuhaus IM, Sharfstein ST (2010) Transcriptomic responses to sodium chloride-induced osmotic stress: a study of industrial fed-batch CHO cell cultures. Biotechnol Prog 26(4):1104–1115.  https://doi.org/10.1002/btpr.398 CrossRefPubMedGoogle Scholar
  31. Siganporia CC, Ghosh S, Daszkowski T, Papageorgiou LG, Farid SS (2014) Capacity planning for batch and perfusion bioprocesses across multiple biopharmaceutical facilities. Biotechnol Prog 30(3):594–606.  https://doi.org/10.1002/btpr.1860 CrossRefPubMedPubMedCentralGoogle Scholar
  32. Sun Z, Zhou R, Liang S, McNeeley KM, Sharfstein ST (2004) Hyperosmotic stress in murine Hybridoma cells: effects on antibody transcription, translation, posttranslational processing, and the Cell Cycle. Biotechnol Prog 20(2):576–589.  https://doi.org/10.1021/bp0342203 CrossRefPubMedGoogle Scholar
  33. Sung YH, Song YJ, Lim SW, Chung JY, Lee GM (2004) Effect of sodium butyrate on the production, heterogeneity and biological activity of human thrombopoietin by recombinant Chinese hamster ovary cells. J Biotechnol 112(3):323–335.  https://doi.org/10.1016/j.jbiotec.2004.05.003 CrossRefPubMedGoogle Scholar
  34. Wang W (2005) Protein aggregation and its inhibition in biopharmaceutics. Int J Pharm 289(1–2):1–30.  https://doi.org/10.1016/j.ijpharm.2004.11.014 CrossRefPubMedGoogle Scholar
  35. Wang Z, Ma X, Zhao L, Fan L, Tan W-S (2012) Expression of anti-apoptotic 30Kc6 gene inhibiting hyperosmotic pressure-induced apoptosis in antibody-producing Chinese hamster ovary cells. Process Biochem 47(5):735–741.  https://doi.org/10.1016/j.procbio.2012.02.001 CrossRefGoogle Scholar
  36. Wang K, Zhang T, Chen J, Liu CX, Tang J, Xie Q (2018) The effect of culture temperature on the aggregation of recombinant TNFR-fc is regulated by the PERK-eIF2a pathway in CHO cells. Protein Pept Lett 25:570–579.  https://doi.org/10.2174/0929866525666180530121317 CrossRefPubMedGoogle Scholar
  37. Wu M-H, Dimopoulos G, Mantalaris A, Varley J (2004) The effect of hyperosmotic pressure on antibody production and gene expression in the GS-NS0 cell line. Biotechnol Appl Biochem 40(1):41–46.  https://doi.org/10.1042/BA20030170 CrossRefPubMedGoogle Scholar
  38. Xie P, Niu H, Chen X, Zhang X, Miao S, Deng X, Liu X, Tan WS, Zhou Y, Fan L (2016) Elucidating the effects of pH shift on IgG1 monoclonal antibody acidic charge variant levels in Chinese hamster ovary cell cultures. Appl Microbiol Biotechnol 100(24):10343–10353.  https://doi.org/10.1007/s00253-016-7749-4 CrossRefPubMedGoogle Scholar
  39. Zhang X, Sun YT, Tang H, Fan L, Hu D, Liu J, Liu X, Tan WS (2015) Culture temperature modulates monoclonal antibody charge variation distribution in Chinese hamster ovary cell cultures. Biotechnol Lett 37(11):2151–2157.  https://doi.org/10.1007/s10529-015-1904-3 CrossRefPubMedGoogle Scholar
  40. Zheng C, Zhuang C, Chen Y, Fu Q, Qian H, Wang Y, Qin J, Wu X, Qi N (2018a) Improved process robustness, product quality and biological efficacy of an anti-CD52 monoclonal antibody upon pH shift in Chinese hamster ovary cell perfusion culture. Process Biochem 65:123–129.  https://doi.org/10.1016/j.procbio.2017.11.013 CrossRefGoogle Scholar
  41. Zheng C, Zhuang C, Qin J, Chen Y, Fu Q, Qian H, Wu T, Wang Y, Wu X, Qi N (2018b) Combination of temperature shift and hydrolysate addition regulates anti-IgE monoclonal antibody charge heterogeneity in Chinese hamster ovary cell fed-batch culture. Cytotechnology 70(4):1121–1129.  https://doi.org/10.1007/s10616-018-0192-x CrossRefPubMedGoogle Scholar
  42. Zhou W, Chen C-C, Buckland B, Aunins J (1997) Fed-batch culture of recombinant NS0 myeloma cells with high monoclonal antibody production. Biotechnol Bioeng 55(5):783–792.  https://doi.org/10.1002/(SICI)1097-0290(19970905)55:5<783::AID-BIT8>3.0.CO;2-7 CrossRefPubMedGoogle Scholar
  43. Zhuang C, Zheng C, Chen Y, Huang Z, Wang Y, Fu Q, Zeng C, Wu T, Yang L, Qi N (2017) Different fermentation processes produced variants of an anti-CD52 monoclonal antibody that have divergent in vitro and in vivo characteristics. Appl Microbiol Biotechnol 101(15):5997–6006.  https://doi.org/10.1007/s00253-017-8312-7 CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.School of PharmacyWuhan UniversityWuhanChina
  2. 2.Shanghai Taiyin Biotech Co., Ltd.ShanghaiChina
  3. 3.School of PharmacyShanghai Jiao Tong UniversityShanghaiChina

Personalised recommendations