Cell culture medium

A culture medium, feed medium technology, applied in the field of cell culture and recombinant protein production

Active Publication Date: 2017-12-01
BOEHRINGER INGELHEIM INT GMBH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Embodiment 1

[0171] CHO2 (CHO-DG44) rituximab cells were cultured in RPMI-based basal medium, RPMI amino acid (AA) ratios versus optimal amino acid (AA) ratios with different total cumulative amino acid amounts. Medium 4 with RPMI AA ratios (Medium 4.0, 4.1, 4.2 and 4.3) and medium with optimized AA ratios ( The media compositions of media 5.0, 5.1 and 5.2) are shown in Table 1 and the corresponding amino acid ratios are shown in Table 2.

[0172] Small changes in total AA concentration are due to changes in molecular weight and minimal changes in the amino acid powder used. The purpose of this experiment was to demonstrate the effect of optimized amino acid ratios at different total cumulative amino acid levels. Experiments were performed in batch mode in duplicate (N=2).

[0173] Table 1: Composition of media 4.0, 4.1, 4.2 and 4.3 and media 5.0, 5.1 and 5.2

[0174]

[0175] *Gln, Ile and Cys were added with stock solution

[0176] **Gln and Ile were added with original solution ...

Embodiment 2

[0191] Based on the optimized amino acid ratios of basal medium 5 (RPMI-based), several amino acids varied + / - 20% and + / - 40% in their molar concentrations as single component methods (calculated based on optimized AA ratios mole percent). This performance was then compared to a control culture grown in medium 5.3 (same as medium 5.0, but with all amino acids added individually as stocks). All essential amino acids are provided in concentrated stock solutions to engineer different media compositions. Thus, the media had comparable total cumulative amino acid amounts of approximately 43-44 mM, but different amino acid ratios. In one experiment, +20% and -20% changes in single amino acid concentrations (L-arginine, L-asparagine, L-aspartate, L-histamine) relative to control medium 5.3 were tested. acid, L-leucine, L-lysine, L-methionine, L-phenylalanine, L-proline, L-serine, L-threonine, L-tryptophan, Single component approach for L-tyrosine, L-valine). For single amino aci...

Embodiment 3

[0204] Based on the optimized amino acid ratios and the amino acids identified in Example 2 in basal medium 5 (RPMI-based), the additional amino acids varied in their molar concentrations in different medium backgrounds due to the single component approach - 40%. This medium containing optimized AA ratios was further optimized for serum-free recombinant protein production and was chemically defined and superior to the modified RPMI medium used in previous experiments. In this experiment, single amino acids were reduced in batch mode to demonstrate the effect of optimal amino acid ratios on pH, dissolved oxygen (DO) and temperature in basal media under controlled bioreactor conditions.

[0205] Based on the optimized amino acid ratio in basal medium 6.2, the single amino acid L-lysine, L-methionine, L-proline, L-tryptophan or L-tyrosine in its molar concentration A 40% reduction, or a 20% or 40% reduction in both L-tyrosine and L-lysine. The resulting performance was compared...

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Abstract

The present invention provides a basal cell culture medium and a feed medium with novel amino acid ratios and/or ferrous cholate as iron carrier that result in improved performance of mammalian cell culture processes, such as CHO cultivation and protein production processes, in particular in increased product titer (e.g. of monoclonal antibodies). Also provided are methods for culturing mammalian cells and producing a protein of interest using said basal cell culture medium and optionally feed medium. The invention also provides for a medium platform that comprises (i) the basal cell culture medium and (ii) the feed medium.

Description

technical field [0001] The present invention relates to the fields of cell culture and recombinant protein production in mammalian cells. It specifically relates to optimal production (e.g. titers) and performance (e.g. cell growth) of products (e.g. (poly)peptides, antibodies, antibody fragments and antibody-derived molecules) in mammalian cell culture using recombinant mammalian host cells A new basal cell culture medium and a new feed medium. Background technique [0002] The development of mammalian cell culture processes for the large-scale industrial manufacture of therapeutic proteins such as monoclonal antibodies began about 25 years ago. Efficient bioprocessing for the production of biopharmaceuticals primarily requires (i) high-yielding, stable, and regulatory-accepted (usually mammalian) cell lines, (ii) optimal cell culture media to support production in different (usually mammalian) ) host cells and cell growth and production in different culture systems and p...

Claims

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Application Information

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IPC IPC(8): C12N5/071C12P21/00
CPCC12N2511/00C12N2500/24C12N2500/32C12N2500/02C12N5/0682C12P21/00C12N2500/90C12N5/0031C12N5/0037C07K16/2887C07K2317/24C12N2500/46
Inventor C·克莱门斯J·绍布M·林克P·朔恩T·舒尔茨
Owner BOEHRINGER INGELHEIM INT GMBH
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