Use of the Ambr 250 to enable rapid clone selection and process development for large scale manufacturing processes

Currently, widely used bench scale bioreactor systems require much user manipulation, a large amount of raw materials, and have a long turnaround time for reactor cleaning and rebuilding. New technologies such as robotic disposable bioreactor systems provide a solution that is miniaturized, high throughput, and substantially automated. The Ambr® 250 offers such a solution, with 24x250mL bioreactors controlled independently. Although this new technology is rapidly being adopted by several groups as a way to increase efficiency and speed within upstream development, it remains to be proven that these systems are complete models for process characterization. We have shown that Ambr 250 is a good scale down model for multiple cell line systems. The aim of this work is to further characterize the engineering environment of the Ambr® 250 with a view of defining its role in industrial cell culture process development and scale-up. CFD modeling of the Ambr 250 mammalian vessel with validation via Particle Image Velocimetry (PIV) was conducted to simulate the hydrodynamic environment in the vessel. These findings were evaluated against current benchtop models and manufacturing scales. Cultures were run utilizing different engineering parameters (vvm, P/V, kLa) to assess the scalability of the current system. Cell growth, production, and product quality were compared across to recommend operating conditions for the Ambr® 250 that best match manufacturing scale reactors. Multiple CHO host cell lines were examined in order to find optimal operating conditions for the Ambr® 250 system

Understanding the impact of high gas entrance velocity on CHO cell culture processes to improve process scale up

Please click Additional Files below to see the full abstract

High yielding large-scale transient expression of Biotherapeutics in CHO: Beyond preclinical development

Please click Additional Files below to see the full abstract

Next generation perfusion process development for production of biologics

Please click Additional Files below to see the full abstract

Fucosylation inhibitor development for producing afucosylated antibodies

Please click Additional Files below to see the full abstract

A Master Cell Bank (MCB) banking troubleshooting case study: Challenges and process improvements with comprehensive root cause analysis

Mammalian cells, especially Chinese hamster ovary (CHO) cells, are routinely used in the biopharmaceutical industry for production of recombinant therapeutic proteins. Master Cell banking is one of the key step during drug development, which ensures preservation of cells at low temperatures for an extended period of time for GMP drug substance manufacturing. CHO cells can show significant variation in growth characteristics during cell line development. This variation necessitates the need for a robust Master Cell Bank (MCB) manufacturing process to ensure consistent MCB thaw and growth. Numerous efforts have been done to understand the cryopreservation mechanism as well as techniques to improve the robustness of banking processes. However, failure of MCB releasing still happens across the industry. A case study will be presented highlighting experiments carried out to identify root cause of MCB thaw and expansion variability. In this study, the health of the cells was examined using an Apoptosis assay and Transmission Electron Microscopy (TEM) analysis to gain a better understanding of the cell bank health. Process improvements that included further passaging of the cell line for improved robustness of the MCB manufacturing process will be discussed

Development of an N-1 perfusion process and optimized scale-down models for implementation in a platform CHO cell culture manufacturing process

The use of N-1 perfusion, coupled with high-inoculum fed batch in CHO cell culture manufacturing processes, has been shown to increase volumetric productivity and shorten the duration of the fed-batch production phase. Implementation of N-1 perfusion as part of a platform process requires the ability to screen multiple clones and to optimize media and process parameters in a high-throughput manner. We have developed an N-1 perfusion process, along with a series of scale-down models for N-1 perfusion using shake flasks, cell culture tubes, and deep-well plates. Process parameters for scale-down models were optimized to maximize comparability of growth profiles and cell culture performance relative to 5L N-1 perfusion bioreactors. Scale-down models were used to inoculate fed-batch experiments in Ambr15 micro-bioreactors at high seeding density, in order to compare growth and productivity profiles to those observed in 5L bench scale bioreactors. Multiple cell lines derived from different CHO hosts were evaluated in order to verify the robustness of the scale-down models. Results demonstrated that cell growth and viability in the optimized scale-down models were comparable to those observed in 5L N-1 perfusion bioreactors. Furthermore, growth, productivity, and product quality profiles from high-inoculum fed-batch experiments were comparable regardless of inoculum source. Optimized scale down models of N-1 perfusion, coupled with Ambr15 fed-batch production micro-bioreactors, have now been integrated into a high-throughput and robust workflow to enable DOE and screening experiments for clone selection, media development and parameter optimization in a platform N-1 perfusion process for monoclonal antibody manufacturing

Enabling pandemic speed to clinic for SARS-CoV2 neutralizing antibodies

Please click Additional Files below to see the full abstract

Upstream control strategy development for afucosylated species in mAb biomanufacturing

Please click Additional Files below to see the full abstract

Identification of Cell Culture Factors Influencing Afucosylation Levels in Monoclonal Antibodies by Partial Least-Squares Regression and Variable Importance Metrics

Retrospective analysis of historic data for cell culture processes is a powerful tool to develop further process understanding. In particular, deploying retrospective analyses can identify important cell culture process parameters for controlling critical quality attributes, e.g., afucosylation, for the production of monoclonal antibodies (mAbs). However, a challenge of analyzing large cell culture data is the high correlation between regressors (particularly media composition), which makes traditional analyses, such as analysis of variance and multivariate linear regression, inappropriate. Instead, partial least-squares regression (PLSR) models, in combination with machine learning techniques such as variable importance metrics, are an orthogonal or alternative approach to identifying important regressors and overcoming the challenge of a highly covariant data structure. A specific workflow for the retrospective analysis of cell culture data is proposed that covers data curation, PLS regression, model analysis, and further steps. In this study, the proposed workflow was applied to data from four mAb products in an industrial cell culture process to identify significant process parameters that influence the afucosylation levels. The PLSR workflow successfully identified several significant parameters, such as temperature and media composition, to enhance process understanding of the relationship between cell culture processes and afucosylation levels