(5f) Effect of Endoplasmic Reticulum Stress On Protein Productivity In Recombinant CHO Cells

Nearly 70% of the recombinant therapeutics used worldwide are produced as secretory proteins in mammalian cells. Productivity of such secretory proteins is a multifaceted phenomenon. Multiple pathways operate in coordination to secrete functionally active proteins out of the cells. The high levels of intracellular protein may disrupt endoplasmic reticulum (ER) homeostasis by increasing the protein folding load on the ER. In response to the high ER stress levels, the cell can respond in different ways: overall translation attenuation, feedback regulation of chaperones or apoptosis. The final choice depends on the interplay between the adaptive and apoptosis pathways.

In this work, we attempt to understand the effect of ER stress on productivity through a combination of experimental and modeling approach. From the available literature on regulation between different genes in the ER, a comprehensive model has been developed for protein secretion. The model accounts for three pathways - protein folding, degradation and unfolded protein response. The model is being simulated to study the effect of genetic alterations on the overall folding efficiency. An optimum chaperone concentration is predicted for enhanced folding efficiency. Depending on the kinetics of folding and degradation, increasing the chaperone concentration above this critical concentration may lead to decrease in folding efficiency. It has also been observed experimentally that over-expression of BiP, a key chaperone, leads to increase in productivity in some cell lines and decrease in others.

In parallel, we have also profiled the dynamics of more than 20 key genes involved in these pathways in different recombinant protein CHO cell lines exhibiting varying productivity levels of IgG. The data shows that the transcript levels of chaperones and UPR transducers vary with productivity levels, with a general increase in overall levels in the high producers. The resulting data is being further analyzed using multivariate analysis tools to identify the key genes influencing productivity. Such a systems level analysis of protein secretion pathway should provide a rational basis to identification of cell engineering targets for enhanced productivity.