Mitigation of therapeutic protein fragmentation during cell culture harvest processes
AssigneeAmgen Inc.
InventorsGang Xiao, Hyo Seon Chung, Quanzhou Luo, Tamer Eris, Joseph Edward Basconi, Xiaoyang Zhao, Pavel Bondarenko, Krista Marie Petty, Oliver Kaltenbrunner
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Abstract
Disclosed herein are methods for harvesting a recombinant protein comprising an asparaginyl endopeptidase (AEP) cleavage site, as well as methods for inhibiting cleavage of a recombinant protein comprising an AEP cleavage site, during a biomanufacturing process.
Claims22
A method for harvesting a recombinant protein during a biomanufacturing process comprising:
- establishing a cell culture by inoculating a bioreactor with mammalian cells expressing the recombinant protein;
- maintaining the cell culture during a growth phase and a production phase, wherein an exogenous protease inhibitor is optionally admixed with the cell culture during the growth phase and/or during or after the production phase;
- cooling the cell culture, wherein a dissolved oxygen level of about 64 mmHg to about 128 mmHg is optionally maintained in the cell culture during the cooling;
- admixing an acidic solution with the cell culture to obtain an acidified cell culture with a pH of about 4.6 to about 5.3;
- and
- incubating the acidified cell culture to induce agglomeration of one or more cell culture impurities,
- wherein the recombinant protein comprises an asparaginyl endopeptidase (AEP) cleavage site.
A method for inhibiting cleavage of a recombinant protein comprising an asparaginyl endopeptidase (AEP) cleavage site during a biomanufacturing process comprising an acid precipitation, the method comprising:
- measuring a quantity or activity of AEP and/or a quantity of one or more fragments of the recombinant protein in a sample isolated from a cell culture, harvest, or purification operation of the biomanufacturing process;
- and
- adjusting the acid precipitation pH to about 4.6 to about 5.3 if the quantity or activity of AEP and/or the quantity of the one or more fragments is above a threshold.
Dependencies are inferred from the claim text.