Elevated endoplasmic reticulum pH is associated with high growth and
bisAb aggregation in CHO cells
Abstract
Chinese hamster ovary (CHO) bioprocesses, the dominant platform for
therapeutic protein production, are increasingly used to produce complex
multispecific proteins. Product quantity and quality are affected by
intracellular conditions, but these are challenging to measure and often
overlooked during process optimization studies. pH is known to impact
quality attributes like protein aggregation across upstream and
downstream processes, yet the effects of intracellular pH on cell
culture performance are largely unknown. Recently, advances in protein
biosensors have enabled investigations of intracellular environments
with high spatiotemporal resolution. In this study, we integrated a
fluorescent pH-sensitive biosensor into a bispecifc (bisAb)-producing
cell line to investigate changes in endoplasmic reticulum pH (pH
ER). We then investigated how changes in lactate
metabolism impacted pH ER, cellular redox, and product
quality in fed-batch and perfusion bioreactors. Our data show pH
ER rapidly increased during exponential growth to a
maximum of pH 7.7, followed by a sharp drop in stationary phase in all
perfusion and fed-batch conditions. pH ER decline in
stationary phase was driven by an apparent loss of cellular pH
regulation that occurred despite differences in redox profiles. Protein
aggregate levels correlated positively with pH ER,
indicating that aggregation is not solely driven by oxidative stress and
cellular protein load. Improved characterization of intracellular
changes in bioprocesses can help guide media optimizations, improve
bioprocess control strategies or provide new targets for cell
engineering.