文献类型：期刊文献

英文题名：Highly sialylated recombinant human erythropoietin production in large-scale perfusion bioreactor utilizing CHO-gmt4 (JW152) with restored GnT I function

作者：Goh, John S. Y.[1];Liu, Yingwei[2];Liu, Haifeng[2,3];Chan, Kah Fai[1];Wan, Corrine[1];Teo, Gavin[1];Zhou, Xiangshan[2];Xie, Fusheng[3,4];Zhang, Peiqing[1];Zhang, Yuanxing[2];Song, Zhiwei[1]

机构：[1]ASTAR, Bioproc Technol Inst, Singapore 138668, Singapore;[2]E China Univ Sci & Technol, State Key Lab Bioreactor Engn, Shanghai 200237, Peoples R China;[3]Shandong Dong E E Jiao Co Lte, Shandong, Peoples R China;[4]Shandong Ehua Biopharmaceut Co Ltd, Shandong, Peoples R China

年份：2014

卷号：9

期号：1

起止页码：100

外文期刊名：BIOTECHNOLOGY JOURNAL

收录：;EI(收录号：20140317210921);WOS:【SCI-EXPANDED(收录号:WOS:000337540100010)】；

基金：We would like to thank Dr. Natasha Pereira for critical review of the manuscript. This work was jointly funded by the Agency for Science, Technology and Research (A*STAR), Singapore, and the Open Funding Project of the State Key Laboratory of Bioreactor Engineering (SKLBE), East China University of Science and Technology, Shanghai, China.

语种：英文

外文关键词：CHO cells; Erythropoietin (EPO); Glycosylation; N-Acetylglucosaminyltransferase-I (GnT I); Sialylation

摘要：Therapeutic glycoprotein drugs require a high degree of sialylation of their N-glycans for a better circulatory half-life that results in greater efficacy. It has been demonstrated that Chinese hamster ovary (CHO) glycosylation mutants lacking N-acetylglucosaminyltransferase I (GnT I), when restored by introduction of a functional GnT I, produced highly sialylated erythropoietin (EPO). We have now further engineered one of such mutants, JW152, by inactivating the dihydrofolate reductase (DHFR) gene to allow for the amplification of the EPO gene with methotrexate (MTX). Several MTX-amplified clones maintained the ability to produce highly sialylated EPO and one was selected for culture in a perfusion bioreactor that is used in an existing industrial EPO-production bioprocess. Extensive characterization of the EPO produced was performed using total sialic quantification, HPAEC-PAD and MALDI-TOF MS analyses. Our results demonstrated that the EPO produced by the mutant line exhibits superior sialylation compared to the commercially used EPO-producing CHO clone cultured under the same conditions. Therefore, this mutant has the industrial potential for producing highly sialylated recombinant EPO and potentially other recombinant glycoprotein therapeutics.