Decisions made by researchers during the microliter, benchtop development of biotherapeutics made from monoclonal antibodies (mAb) and recombinant proteins often don’t scale to larger stirred tank bioreactors. Identifying reliable operational parameters as early as possible in the development process can significantly streamline scaleup to trial and commercialization quantities.

Scale-up challenges are common among biologics manufacturers. Protein production decreases as cells age, and the bioreactor’s hydrodynamic characteristics can affect process conditions and allow byproducts to accumulate, which can cause cells to lose viability prematurely.

To minimize these challenges, Olivier Henry, PhD, professor, Polytechnique Montreal, and colleagues there and at the National Research Council Canada, advise even bench scientists to consider manufacturing needs. Specifically, they recommend identifying “adequate critical process parameters that are more representative of large-scale stirred tank bioreactor production.”

In a recent paper, Henry’s team detailed the aeration and feeding strategies they used as they scaled production of SARS-CoV-2 spike protein from a 0.75 L bioreactor to a 1 L multisystem bioreactor. Those methods improved process cells’ longevity by five days, increased the integral of viable cell concentration by 1.25-fold, and increased the yield of SARS-CoV-2 spike protein by 2.53-fold in an inducible Chinese hamster ovary (CHO) stable cell pool.

Ensure adequate aeration

Setting appropriate dissolved oxygen setpoints was “important in terms of increasing endpoint viability and protein production yield,” they report. Additionally, setting the sparging air caps appropriately affected cell culture longevity and lactate accumulation. “Finding the adequate aeration conditions is paramount for a scalable process,” the scientists add.

Furthermore, they report, “Increased feeding was only beneficial when coupled with slow pump addition.” Their feed/pump parameters reduced stress on the cells, possibly by minimizing nutrient fluctuations and exposing the cells to greater osmolarity. This approach also enabled more consistent lactate consumption. Additionally, dynamic feed strategies increased titer.

To implement the team’s strategies, the scientists advise incorporating automation to monitor and react to changing process conditions. For monitoring, they suggest focusing on bio-capacitance and the oxygen transfer rate if dissolved oxygen levels are measured accurately, because of their direct correlation to viable cell volume. If dissolved oxygen levels cannot be measured accurately, however, they recommend using standard respiration tests to estimate the oxygen uptake rate.

Finally, they point out, “Feeding strategies and processing parameters must be optimized in tandem since the positive impact of dosage and increased feeding can be undone by adverse aeration conditions and inappropriate do setpoints.”

Future work will evaluate the correlation between process-related parameters and quality profiles of the SARS-CoV-2 spike protein.