Changes in Media Composition due to Expression of IgG in HEK293 and CHO DG44 Cells

Quality of recombinant/therapeutic proteins produced by engineered cell lines is a concern shared by both manufacturers and regulatory agencies. New assays are needed to facilitate earlier and more rapid analysis of therapeutic protein quality. Cell culture supernatant contains secreted proteins that are indicative of the cellular condition and thus are potential indicators of recombinant protein quality. This study identified changes in secreted host cell proteins (HCP) from cell pools exhibiting varying levels of recombinant antibody expression as determined by quantitative mass spectrometry. CHO DG44 cell pools were stably transfected with IgG expressing vectors and control with blank vectors. High, medium and low IgG-producing cell pools were regulated with methotrexate. Secreted HCP were analyzed in cell supernatants by mass spectrometry and a total of 450 proteins were identified. Protein functions included secreted proteins, regulatory proteins, chaperones and proteins associated with cell growth. Protein quantification revealed distinct patterns of secreted HCP concentrations that distinguished high, medium, low and non IgG-producing cell pools. A subset of 50 proteins was selected that showed the most dramatic concentration changes and the greatest ability to differentiate variation in IgG production. These proteins established a fingerprint of secreted HCP correlating to IgG expression.

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Peter G. Slade, Staff Scientist, Life Tech
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