Combination of temperature shift and hydrolysate addition regulates anti-IgE monoclonal antibody charge heterogeneity in Chinese hamster ovary cell fed-batch culture
- 220 Downloads
Charge heterogeneity has been broadly studied as a critical quality attribute during monoclonal antibody (mAb) production that may subsequently affect product stability and biopotency. However, the charge variation distribution is poorly controlled, so methods of more effective control need to be explored. In this study, the combined effects of temperature shift (37–34, 37–32, or 37–30 °C) and hydrolysate addition (0.100 g/L) to culture feed on the charge heterogeneity of anti-IgE mAb were investigated. The results showed that the distribution of charge variation was significantly regulated by the combination of hydrolysate addition with a highly sub-physiological temperature (34 °C). In addition, under this condition, the main peak content significantly increased, and the acidic peak content significantly decreased. Furthermore, we explored Lys variant content, which is the major basic variant content, as well as its relationship with temperature shift and hydrolysate addition. Lys variant levels were positively related to the Lys and Arg concentrations in the medium and negatively related to carboxypeptidase B and carboxypeptidase H transcript levels. The combination of temperature shift and hydrolysate addition can thus effectively improve anti-IgE mAb charge heterogeneity and significantly increase main variant levels and decrease acidic variant levels.
KeywordsCarboxypeptidase Charge heterogeneity Lysine variants Temperature
This work was supported by the Science and Technology Commission of Shanghai Municipality (Nos. 14431903100, 17431905800).
- Heidemann R, Zhang C, Qi HS, 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 32:157–167. https://doi.org/10.1023/a:1008196521213 CrossRefPubMedPubMedCentralGoogle Scholar
- Khawli LA, Goswami S, Hutchinson R, Kwong ZW, Yang JH, Wang XD, Yao ZL, Sreedhara A, Cano T, Tesar D, Nijem I, Allison DE, Wong PY, Kao YH, Quan C, Joshi A, Harris RJ, Motchnik P (2010) Charge variants in IgG1 Isolation, characterization, in vitro binding properties and pharmacokinetics in rats. MAbs 2:613–624. https://doi.org/10.4161/mabs.2.6.13333 CrossRefPubMedPubMedCentralGoogle Scholar
- Logsdon SL, Oettgen HC (2015) Anti-IgE therapy: clinical utility and mechanistic insights. In: Lafaille JJ, DeLafaille MAC (eds) Ige antibodies: generation and function. Current topics in microbiology and immunology, vol 388. Springer, Berlin, pp 39–61Google Scholar
- Xie PP, Niu HJ, Chen XN, Zhang XT, Miao SW, Deng XC, Liu XP, 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 100:10343–10353. https://doi.org/10.1007/s00253-016-7749-4 CrossRefPubMedGoogle Scholar
- Yan BX, Steen S, Hambly D, Valliere-Douglass J, Bos TV, Smallwood S, Yates Z, Arroll T, Han YH, Gadgil H, Latypov RF, Wallace A, Lim A, Kleemann GR, Wang WC, Balland A (2009) Succinimide formation at Asn 55 in the complementarity determining region of a recombinant monoclonal antibody IgG1 heavy chain. J Pharm Sci 98:3509–3521. https://doi.org/10.1002/jps.21655 CrossRefPubMedGoogle Scholar