(683a) Constrained Optimisation of Cell Culture Feeding Strategy and Temperature Shift Duration to Enhance Monoclonal Antibody Titre and Purity

Recombinant proteins have been extensively studied for their wide therapeutic and research applications. The main therapeutic product category is that of monoclonal antibodies, which have been approved to treat a variety of chronic and life-threatening diseases. Increasing production titre has been achieved mainly by cell culture medium improvement and genetic engineering. However, mathematical modeling of bioprocess dynamics is a valuable tool to improve industrial production at a much faster rate and lower cost in comparison to statistical design of experiment approaches. A holistic insight to comprehend the interplay between crucial processes variables can be gained through the application of process system engineering tools. In this study, a single stage population balance model has been built to capture mammalian cells behaviour in fed-batch bioreactors. The model represents two operating modes; the first at constant physiological temperature and the second with a shift to mild hypothermic conditions (32^oC) from the fifth day onwards, which represents common industrial practice.

The model considers the dynamic profile of substrates and metabolites, product titre and one of the main cell-derived impurities, host cell proteins (HCPs), which can impact product immunogenicity and integrity. Certain HCPs are naturally secreted, while others are released as a result of cell death and lysis. Our model considers these three sources of HCPs to determine their overall concentration at harvest. Culture osmolality is also considered as a determining factor for cell growth rate and size distribution.

The model parameters have been estimated with high confidence using a recently published dataset for antibody-producing Chinese hamster ovary (CHO) fed-batch cell cultures for physiological temperature and with a shift to mild hypothermia in mid-exponential growth phase. The model was then used to optimise feeding volume and frequency as well as duration of temperature downshift subject to maintaining culture viability above 80%, as is current industrial practice. and this has been set as a constraint in all our model optimisation runs. Specifically, we looked at three scenarios: (a) increased feeding to sustain high viability for longer, which maintained titre but reduced cell death; (b) a 50% reduction in feed volume to avoid overfeeding, which led to a slightly higher titre compared to the control; (c) a shift in culture temperature on day 6 which gives the highest product/ HCP ratio.

In general, higher product titres and prolonged culture viability can be attained at the expense of higher feeding volume. However, when a constraint on HCP concentration is also applied model-based optimisation results in shorter culture duration and, in turn, overall lower antibody titre. This study shows the usefulness of mathematical modeling for exploring trade-offs in bioprocess performance. It is also a viable tool to systematically accelerate the duration of processes development and optimisation. Integrating this model with a downstream purification model to evaluate the cost of removing these fractions of impurities, can help determine what concentration of HCPs can be economically tolerated in the cell culture supernatant and aid whole bioprocess design.