Antibody purification and purity monitoring

摘要

Processes for producing and purifying recombinant proteins are disclosed. In particular, the present disclosure provides processes of producing and purifying multi-subunit proteins expressed in yeast or filamentous fungal cells. The production and/or purification of such proteins are monitored for impurities, preferably using lectin binding assays, such that one or more process parameters may be adjusted to maximize the amount of desired recombinant protein and minimize the amount of glycosylated impurities. The processes can also be monitored for other undesired product-associated impurities, such as aggregates and nucleic acids. In exemplary embodiments, the recombinant proteins are multi-subunit proteins, such as antibodies, the host cell is a yeast, such as Pichia pastoris, and the glycosylated impurity is a glycovariant of the desired recombinant polypeptide, such as an N-linked and/or O-linked glycovariant.

主权项

1. A fermentation process for producing a desired recombinant antibody polypeptide and purifying the desired recombinant polypeptide from the fermentation medium, wherein the process includes:
(i) conducting one or more fermentation processes or runs which each comprise culturing yeast cells under conditions that result in the expression and secretion of a desired recombinant polypeptide and one or more impurities into the fermentation medium; (ii) periodically obtaining one or more samples of the fermentation medium as each fermentation process proceeds or after different fermentation runs are conducted; (iii) detecting the amount and/or type of glycosylated impurities in the sample(s) using a lectin that binds to said glycosylated impurities, and (iv) modifying one or more of the operating parameters or conditions of one or more of the fermentation processes or runs based on the amount of detected glycosylated impurities in the sample(s), wherein said operating parameters or conditions are selected from the group consisting of temperature, pH, gas constituent, feed constituent, agitation, aeration, antifoam and duration, while said fermentation processes or runs are being conducted, and (v) pooling different samples, eluates or fractions thereof containing the desired recombinant antibody polypeptide from the same or different fermentation processes or runs as the fermentation processes or runs are being conducted based on the amount and/or type of detected glycosylated impurity relative to the amount of the recombinant antibody polypeptide in said different samples, eluates or fractions thereof.