With omics data being generated at an unprecedented rate, genome-scale modelling has become pivotal in its organisation and analysis. However, machine learning methods have been gaining ground in cases where knowledge is insufficient to represent the mechanisms underlying such data or as a means for data curation prior to attempting mechanistic modelling. We discuss the latest advances in genome-scale modelling and the development of optimisation algorithms for network and error reduction, intracellular constraining and applications to strain design. We further review applications of supervised and unsupervised machine learning methods to omics datasets from microbial and mammalian cell systems and present efforts to harness the potential of both modelling approaches through hybrid modelling.

The era of big data: Genome-scale modelling meets machine learning.

Enzyme capacity-based genome scale modelling of CHO cells

A key aspect of large-scale production of biotherapeutics is a well-designed and consistently-executed upstream cell culture process. Process analytical technology tools provide enhanced monitoring and control capabilities to support consistent process execution, and also have potential to aid in maintenance of product quality at desired levels. One such tool, Raman spectroscopy, has matured as a useful technique to achieve real-time monitoring and control of key cell culture process attributes. We developed a Raman spectroscopy-based nutrient control strategy to enable dual control of lactate and glucose levels for a fed-batch CHO cell culture process for monoclonal antibody (mAb) production. To achieve this, partial least squares-based chemometric models for real-time prediction of glucose and lactate concentrations were developed and deployed in feedback control loops. In particular, feeding of lactic acid post-metabolic shift was investigated based on previous work that has shown the impact of lactate levels on ammonium as well as mAb product quality. Three feeding strategies were assessed for impact on cell metabolism, productivity, and product quality: bolus-fed glucose, glucose control at 4 g/L, or simultaneous glucose control at 4 g/L and lactate control at 2 g/L. The third feeding strategy resulted in a significant reduction in ammonium levels (68%) while increasing mAb galactosylation levels by approximately 50%. This work demonstrated that when deployed in a cell culture process, Raman spectroscopy is an effective technique for simultaneous control of multiple nutrient feeds, and that lactic acid feeding can have a positive impact on both cell metabolism and mAb product quality.

Tuning monoclonal antibody galactosylation using Raman spectroscopy-controlled lactic acid feeding

Concomitant consumption of glucose and lactate: A novel batch production process for CHO cells

The increasing demand for biopharmaceuticals produced in mammalian cells has driven the industry to enhance the productivity of bioprocesses through intensification of culture process. Fed-batch and perfusion culturing strategies are considered the most attractive choices, but the application of these processes requires the availability of reliable online measuring systems for the estimation of cell density and metabolic activity. This manuscript reviews the methods (and the devices used) for monitoring of the oxygen consumption, also known as oxygen uptake rate (OUR), since it is a straightforward parameter to estimate viable cell density and the physiological state of cells. Furthermore, as oxygen plays an important role in the cell metabolism, OUR has also been very useful to estimate nutrient consumption, especially the carbon (glucose and glutamine) and nitrogen (glutamine) sources. Three different methods for the measurement of OUR have been developed up to date, being the dynamic method the golden standard, even though DO and pH perturbations generated in the culture during each measurement. For this, many efforts have been focused in developing non-invasive methods, such as global mass balance or stationary liquid mass balance. The low oxygen consumption rates by the cells and the high accuracy required for oxygen concentration measurement in the gas streams (inlet and outlet) have limited the applicability of the global mass balance methodology in mammalian cell cultures. In contrast, stationary liquid mass balance has successfully been implemented showing very similar OUR profiles compared with those obtained with the dynamic method. The huge amount of studies published in the last years evidence that OUR have become a reliable alternative for the monitoring and control of high cell density culturing strategies with very high productivities.

New developments in online OUR monitoring and its application to animal cell cultures

The increasing demand for biopharmaceuticals produced in mammalian cells has driven the industry to enhance productivity of bioprocesses through different strategies. This is why fed-batch and perfusion cultures are considered more attractive choices than batch processes. In this context, the availability of reliable online measuring systems for cell density and metabolic activity estimation will help the application of these processes. The present work focuses on the comparison of two different monitoring tools for indirect estimation of biomass concentration in a HEK293 cell cultures producing IFN-γ: on one side, the oxygen uptake rate (O.U.R.) determination, by means of application of the dynamic method measurement which is already a widely used tool and, on the other side, a new robust online monitoring tool based on the alkali buffer addition used to maintain the pH set point. Both strategies allow a proper monitoring of cell growth and metabolic activity, with precise identification of the balanced cell growth and the most important action in the process, as is the media feeding. The application of these monitoring systems in fed-batch processes allows extending the growth of HEK293 cells, which in turn results in higher final cell concentrations compared with Batch strategy (7 · 106 cells mL−1), achieving 14 · 106 cells mL−1 for the fed-batch based on O.U.R. and 19 · 106 cells mL−1 for the fed-batch based on the alkali addition. Product titter is also increased in respect of the batch strategy (3.70 mg L−1), resulting in 8.27 mg L−1 when fed-batch was based on O.U.R. and 11.49 mg L−1 when it was based on the alkali buffer strategy. Results prove that fed-batch strategy based on the alkali buffer addition is a robust online monitoring method that has shown its great potential to optimize the feeding strategy in HEK293 fed-batch cultures.

J. Triquell

Papers

A new strategy for fed-batch process control of HEK293 cell cultures based on alkali buffer addition monitoring: comparison with O.U.R. dynamic method

Concomitant consumption of glucose and lactate: A novel batch production process for CHO cells