Purification of monoclonal antibodies by aqueous two-phase systems

Abstract

Monoclonal antibodies (mAbs) are becoming increasingly important as biopharmaceutical product. The high specific affinity of mAbs is successfully used for the therapy of numerous severe diseases, such as autoimmune diseases, infections and many types of cancer. In order to be used as a drug, mAbs must be processed after biotechnological production by animal cell cultivation. For the benefit of patient safety, high quality and purity criteria apply, which must be met at the end of the manufacturing process. Due to the steadily increasing demand, in combination with improvements in the cell cultivation over the last decades, existing facilities and conventional purification methods are reaching their capacity limits. To meet these challenges, new, alternative methods such as aqueous two-phase extraction (ATPE) move into the focus of research. In the first part of this PhD thesis, the composition of the aqueous two-phase system (ATPS) was optimized with respect to yield and purity of the used mAb by means of a Design of Experiments (DoE) approach. Subsequently, a new methodology was developed to separate the mAb-containing product phase of the ATPS from the impurity-containing phase using a membrane-based approach. Thus, the clarification with the capture and first purification of the mAb was successfully integrated in one unit operation. In the second part of the thesis the integration of the ATPE into the subsequent downstream process was investigated. After optimization of the individual unit operations, the entire mAb production process based on ATPE was successfully executed with high yield, purity and constant product quality of the mAb. In the third part of the PhD thesis, the proof of concept for the application of ATPE to high cell density (HCD) processes was investigated, which allow process intensification and increased productivity of the mAb manufacturing. The influence of cell density was investigated using different ATPS. Clarification as well as mAb capture and purification were obtained for HCD cultivation. Thus the results of this work contribute to overcome existing bottlenecks in the production of mAbs, as important biopharmaceutical products.