Process analytical technology is expected to deliver consistent product quality. The refolding operation is affected by critical process parameters such as composition and pH of the solubilization and refolding buffers, and these need to be monitored to ensure consistent product quality. For manufacturing processes based on bacterial systems, refolding is a critical unit operation which determines the economic feasibility and quality of the final product in manufacturing processes involving inclusion bodies. Conventionally, the success of refolding is quantified after the completion of the process by testing refolded samples using reversed-phase high performance liquid chromatography. Researchers have explored the possibility of controlling the refold process using measurements of dissolved oxygen tension, fluorescence, infrared spectroscopy or light scattering. However, these approaches are based on the kinetics and intermediates of refolding reactions, and minimum importance is given to manufacturing facility relevant variables such as buffer preparation and component addition during solubilisation and refolding.<br/>This work proposes use of statistical process control limits for monitoring protein refolding, from buffer preparation to refold quenching for manufacturing of therapeutic proteins in <i>E. coli</i>-based systems. Our approach involves use of online redox potential, temperature, and pH to develop an MVDA-based model. Time profiles of ORP, pH and temperature during refolding about a set point are treated as batch evolution variables and analysed using chemometrics. The refolding unit operation is treated as a semi-batch evolution process which is to be maintained within operating ranges at each point in time. A PAT tool is developed to leverage statistical process control (SPC) charts for detecting deviations from normal operation, identifying the process step causing the deviation, allowing unrecoverable batches to be disposed of early, and recording a signature for each run for future reference when CQAs are not reached. The model has been integrated with LabVIEW to allow us to monitor refolding processes in real time. The proposed strategy was able to successfully identify process deviations and was implemented for obtaining consistent product quality.<br/>

Process analytical technology (PAT) based control of protein refolding: Granulocyte colony stimulating factor (GCSF) as a case study

2019 Spring National Meeting

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