title: Improvement and simplification of fed-batch bioprocesses with a highly soluble phosphotyrosine sodium salt.
abstract: Fed-batch culture bioprocesses are currently used predominantly for the production of recombinant proteins, especially monoclonal antibodies. In these cultures, concentrated feeds are added during cultivation to prevent nutrient depletion, thus extending the cellular growth phase and increasing product concentrations. One limitation in these bioprocesses arises from the low solubility or stability of some compounds at high concentrations, in particular amino acids. This study describes the synthesis and evaluation of a phosphotyrosine disodium salt as a tyrosine source in fed-batch processes. This molecule is highly soluble in concentrated feeds at neutral pH. Mechanistic studies demonstrated that the molecule is cleaved in the cell culture supernatant after processing by released phosphatases, leading to phosphate and free L-tyrosine which can be taken up by the cells. No intact phosphotyrosine was detected intracellularly or incorporated into the sequence of the monoclonal antibody. The use of this new molecule allows the simplification of fed-batch processes in large scale manufacturing via the implementation of neutral pH, highly concentrated feeds.
authors.affiliation: Merck Millipore, Pharm Chemical Solutions, Upstream Cell culture media R&D, Germany. Electronic address: aline.zimmer@merckgroup.com.
authors.forename: Aline
authors.initials: A
authors.lastname: Zimmer
authors.affiliation: Merck Millipore, Pharm Chemical Solutions, Upstream Cell culture media R&D, Germany.
authors.forename: Ronja
authors.initials: R
authors.lastname: Mueller
authors.affiliation: Merck Millipore, Pharm Chemical Solutions, Upstream Cell culture media R&D, Germany.
authors.forename: Maria
authors.initials: M
authors.lastname: Wehsling
authors.affiliation: Merck Millipore, Pharm Chemical Solutions, Upstream Cell culture media R&D, Germany.
authors.forename: Alisa
authors.initials: A
authors.lastname: Schnellbaecher
authors.affiliation: Merck Millipore, Pharm Chemical Solutions, Upstream Cell culture media R&D, Germany.
authors.forename: Joerg
authors.initials: J
authors.lastname: von Hagen
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000378:metabolism*; D019149:Bioreactors; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D003470:Culture Media / Q000737:chemistry* / Q000378:metabolism; D019000:Phosphotyrosine / Q000737:chemistry* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism*; D012964:Sodium / Q000737:chemistry* / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D003470:Culture Media; D011994:Recombinant Proteins; D019000:Phosphotyrosine; D012964:Sodium
keywords: Fed-batch; Monoclonal antibody; Productivity; Single feed; Tyrosine solubility