Cell cycle phase dependent productivity of a recombinant Chinese hamster ovary cell line |
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Authors: | Roshni L Dutton Jeno Scharer Murray Moo-Young |
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Institution: | (1) BioProcess Assist (BPA) Ltd, 8 Maple Leaf Lane , Aberfoyle, ON, Canada, N1H 6H9;(2) Department of Chemical Engineering, University of Waterloo, Waterloo, ON, Canada |
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Abstract: | A Chinese Hamster Ovary cell line, CHO1-15500, producing recombinant human tissue type plasminogen activator (tPA) via the dihydrofolate reductase (DHFR) amplification
system, was studied in batch culture. In this system both DHFR and tPA are under the control of the strong constitutive viral
SV40 early promoter. Employing the cumulative viable cell-hour approach, the specific productivity of tPA had maxima in the
lag (0.065 pg cell−1 h−1) and early decline (0.040 pg cell−1 h−1) population growth phases. The viable population was assigned into four subpopulations (G1, S, G2/M phase, and Apoptotic
cells) using flow cytometric analysis. As expected, intracellular DHFR was maximally expressed during the S cell cycle phase.
The production of tPA, however, was found to be a direct linear function of the G1 phase, with a subpopulation specific productivity
of 0.080 pg c-h−1. Productivity maxima in the lag and early decline corroborate the flow cytometric data, indicative that this recombinant
tPA production occurs primarily in the G1 cell cycle phase, not the S phase. This suggests that endogenous regulatory mechanisms
are important controlling influences on the production of recombinant tPA in this ovarian cell line. Productivity from recombinant
cell lines cannot be inferred from either the plasmid construct or the host cell alone. Elucidation of the relationship between
expression of recombinant protein and the cell cycle phases of the host cell is a major component of the characterization
of the animal cell production system. This information facilitates rational process design, including operating mode, modelling
and control, and medium formulation. |
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Keywords: | Cell cycle associated protein productivity Chinese hamster ovary (CHO) DHFR Regulation of expression Specific productivity Tissue plasminogen activator (tPA) |
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