PMID- 30157452 OWN - NLM STAT- MEDLINE DCOM- 20181211 LR - 20181211 IS - 1873-4863 (Electronic) IS - 0168-1656 (Linking) VI - 285 DP - 2018 Nov 10 TI - Differential gene expression of a feed-spiked super-producing CHO cell line. PG - 23-37 LID - S0168-1656(18)30612-6 [pii] LID - 10.1016/j.jbiotec.2018.08.013 [doi] AB - Feed supplements are concentrated cell culture media that contain a variety of nutrients, which can be added during a bioprocess. During fed-batch cultivation, feed media are typically added to a growing cell culture to maximize cell and product concentrations. In this study, only a single shot of feed medium was added on day 0 to a basal cell culture medium and compared to non-supplemented basal medium (feed-spiked at day 0 versus control experiments) by cultivation of a recombinant mAb expressing CHO cell line in batch mode under controlled conditions in a bioreactor. Since the feed-spike at day 0 was based on existing medium components without introducing additional supplements, a desirable process with decreased complexity was generated. Unlike cells in basal medium, feed-spiked cultures reached almost 2x higher peak cell concentrations (10 x 10(6) c/mL vs. 18 x 10(6) c/mL) and 3x higher antibody concentrations (0.8 g/L vs. 2.4 g/L). Batch process time and the integral over the viable cell count were similar for both process types. Constantly high cell-specific production rates in feed-spiked cultures (70 pg/cell/day) compared to continuously declining rates in basal medium (from 70 to 10 pg/cell/day) were responsible for an overall 70% higher cell-specific production rate and the higher product concentrations. To associate gene expression patterns to different process proceedings, transcriptome analysis was performed using microarrays. Several transcripts that are involved with glutamine de novo synthesis and citric acid cycle were significantly upregulated on several days in feed-spiked cultures. The top identified gene ontology (GO) terms related well to cell cycle and primary metabolism, cellular division as well as nucleobase formation or regulation, which indicated a more active proliferative state for feed-spiked cultures. KEGG biochemical pathway analysis and Gene set enrichment analysis (GSEA) further confirmed these findings from a complementary perspective. Moreover, several interesting gene targets, which have not yet been associated with recombinant protein expression, were identified that related to a higher proliferative state, growth, protein synthesis, cell-size control, metabolism, cell survival as well as genes that are associated with the control of the mammalian target of rapamycin (mTOR) in feed-spiked cultures. Analysis of critical product quality attributes (i.e. glycosylation, charge variants and size distribution) showed that feed-spiking did not change antibody quality. CI - Copyright (c) 2018 The Authors. Published by Elsevier B.V. All rights reserved. FAU - Reinhart, David AU - Reinhart D AD - Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna, Muthgasse 11, 1190 Vienna, Austria. Electronic address: david.reinhart@boku.ac.at. FAU - Damjanovic, Lukas AU - Damjanovic L AD - Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna, Muthgasse 11, 1190 Vienna, Austria. Electronic address: lukas.damjanovic@boku.ac.at. FAU - Castan, Andreas AU - Castan A AD - GE Healthcare Life Sciences AB, Bjorkgatan 30, 75184, Uppsala, Sweden. Electronic address: andreas.castan@ge.com. FAU - Ernst, Wolfgang AU - Ernst W AD - Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna, Muthgasse 11, 1190 Vienna, Austria. Electronic address: wolfgang.ernst@boku.ac.at. FAU - Kunert, Renate AU - Kunert R AD - Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna, Muthgasse 11, 1190 Vienna, Austria. Electronic address: renate.kunert@boku.ac.at. LA - eng PT - Journal Article DEP - 20180826 PL - Netherlands TA - J Biotechnol JT - Journal of biotechnology JID - 8411927 RN - 0 (Antibodies, Monoclonal) RN - 0 (Culture Media) RN - 0 (Recombinant Proteins) SB - IM MH - Animals MH - Antibodies, Monoclonal/*biosynthesis MH - Bioreactors MH - CHO Cells MH - Cell Culture Techniques/*methods MH - Cricetulus MH - Culture Media/*pharmacology MH - Gene Expression/*drug effects MH - Recombinant Proteins/biosynthesis OTO - NOTNLM OT - Bioprocess development OT - Chinese hamster ovary cells OT - Differential gene expression OT - Microarray OT - Monoclonal antibody OT - Product quality OT - Transcriptome EDAT- 2018/08/30 06:00 MHDA- 2018/12/12 06:00 CRDT- 2018/08/30 06:00 PHST- 2018/05/31 00:00 [received] PHST- 2018/08/15 00:00 [revised] PHST- 2018/08/25 00:00 [accepted] PHST- 2018/08/30 06:00 [pubmed] PHST- 2018/12/12 06:00 [medline] PHST- 2018/08/30 06:00 [entrez] AID - S0168-1656(18)30612-6 [pii] AID - 10.1016/j.jbiotec.2018.08.013 [doi] PST - ppublish SO - J Biotechnol. 2018 Nov 10;285:23-37. doi: 10.1016/j.jbiotec.2018.08.013. Epub 2018 Aug 26.