Specific monoclonal antibody productivity and the cell cycle-comparisons of batch, continuous and perfusion cultures

Abstract

A selection of mouse hybridoma cell lines showed a variation of approximately two orders of magnitude in intracellular monoclonal antibody contents. The different levels directly influenced apparent specific monoclonal antibody productivity during the death phase but not during the growth phase of a batch culture. The pattern of changes in specific productivity during culture remained basically similar even though at different levels for all cell lines tested. Arresting the cells in the G1 phase using thymidine increased the specific productivity, cell volume and intracellular antibody content but at the same time led to decreased viability. In continuous culture DNA synthesis decreased with decreasing dilution rate though without an accompanying change in cell cycle and cell size distributions. The data shows both the decrease in viability and intracellular antibody content to be important factors which influence the negative association between specific antibody productivity and growth rate. In high cell density perfusion culture, when the cell cycle was prolonged by slow growth, viability was low and dead, but not lysed, cells were retained in the system, the specific antibody productivity was nearly two fold higher than that obtained in either batch or continuous cultures. The results imply that the prolongation of G1 phase and the increase in death rate of cells storing a large amount of antibody together cause an apparent increase in specific antibody productivity.

Proliferation control strategies to improve productivity and survival during CHO based production culture

“…Protein production is dependent on the phase of the cell-cycle and several genes such as those involved in ribosome biogenesis and protein translation are expressed highly in the G1 phase (Al-Rubeai and Emery 1990;Al-Rubeai et al 1992;Moore et al 1997;Fussenegger et al 1998Fussenegger et al , 2000Kaufman et al 1999Kaufman et al , 2001Carvalhal et al 2003;Ibarra et al 2003;Yoon et al 2003a, b;Fogolin et al 2004;Bi et al 2004;Trummer et al 2006). Cells arrested at the end of G1-phase of cell cycle are metabolically more active and bigger in size than non-arrested cells (Carvalhal et al 2003;Bi et al 2004).…”

“…Cells arrested at the end of G1-phase of cell cycle are metabolically more active and bigger in size than non-arrested cells (Carvalhal et al 2003;Bi et al 2004). For these reasons, the G1-phase of the cell cycle is considered the ideal time for increased production of recombinant proteins and G1 arrest has been used to increase the productivity in a number of commercially relevant cell lines such as hybridomas and CHO (Al-Rubeai and Emery 1990;Al-Rubeai et al 1992;Moore et al 1997;Fussenegger et al 1998Fussenegger et al , 2000Kaufman et al 1999Kaufman et al , 2001Ibarra et al 2003;Yoon et al 2003a, b;Fogolin et al 2004;Trummer et al 2006). Some studies have reported the S phase as the optimal production phase (Lloyd et al 2000;Fox et al 2005), an example being the increased production of human interferon-c (IFN-c) upon increasing the percentage of CHO in S phase (Fox et al 2005).…”

“…Quinidine and thymidine inhibit potassium channels in cells (Al-Rubeai et al 1992;Wang et al 1998;Andersen et al 2000). The level of potassium channel activity has been shown to be different in proliferating and quiescent stage of cell cycle and reduced potassium channel activity is hypothesized to result in cell cycle arrest (Wang et al 1998).…”

Cell Culture Processes for the Production of Viral Vectors for Gene Therapy Purposes

“…The resulting presence of large numbers of dead cells limits culture performance in two ways. Firstly, there is a need for high rates of cell proliferation to replace lost cells, which will reduce the specific productivity (Al-Rubeai et al 1992). Secondly, the release of proteases, DNA and other cellular components will lead to product degradation and will complicate downstream processing (Fassnacht et al 1999).…”

A Cyclin Distributed Cell Cycle Model in GS-NS0

“…The key underlying source of heterogeneity is cell cycle segregation [5][6][7], which is at the centre of cellular growth, death, and productivity, all of which vary during the different cell cycle phases. Specifically, the cell cycle phase can influence the mAb productivity, both of which have been reported to be cell cycle-, cell line-and promoter-dependent [8,9]. Therefore, a better understanding and knowledge of the cell cycle timing, transitions, and associated production profiles can aid the development (modelling, control, and optimisation) of these industrially-relevant systems [10].…”