Our increasing understanding of disease mechanisms coupled with technological advances has facilitated the generation of pharmaceutical proteins, which are able to address yet unmet medical needs. Diseases that were fatal in the past can now be treated with novel biological medications improving and prolonging life for many patients. Pharmaceutical protein production is, however, a complex undertaking, which is by no means problem-free. The demand for more complex proteins and the realization of the importance of post-translational modifications have led to an increasing use of mammalian cells for protein expression. Despite improvements in design and production, the costs required for the development of pharmaceutical proteins still are far greater than those for conventional, small molecule drugs. To render such treatments affordable for healthcare suppliers and assist in the implementation of precision medicine, further progress is needed. In five papers this thesis describes strategies and methods that can help to advance the development and manufacturing of pharmaceutical proteins. Two platforms for antibody engineering have been developed and evaluated, one of which allows for efficient screening of antibody libraries whilst the second enables the straightforward generation of bispecific antibodies. Moreover, a method for epitope mapping has been devised and applied to map the therapeutic antibody eculizumab’s epitope on its target protein. In a second step it was shown how this epitope information can be used to stratify patients and, thus, contribute to the realization of precision medicine. The fourth project focuses on the cell line development process during pharmaceutical protein production. A platform is described combining split-GFP and fluorescence-activated droplet sorting, which allows for the efficient selection of highly secreting cells from a heterogeneous cell pool. In an accompanying study, the split-GFP probe was improved to enable shorter assay times and increased sensitivity, desirable characteristics for high-throughput screening of cell pools. In summary, this thesis provides tools to improve design, development and production of future pharmaceutical proteins and as a result, it makes a contribution to the goal of implementing precision medicine through the generation of more cost-effective biopharmaceuticals for well-characterized patient groups. / <p>QC 20170828</p>

The BioProcessing Demonstration and Training Laboratory was established with a collaboration between Utah State University and Thermo Fisher- Scientific, Inc. This lab was developed into a fully functioning tissue culture facility. Demonstration of tissue culture procedures were necessary for the lab to be industry-fully qualified. The CHO 1-15 cell line protocols were optimized by establishing conditions for reproducibility in shaker flasks and bioreactors (2 to 250 L capacity). CHO 1-15 is the cell line of choice for protein production in the bio-manufacturing industry. Oxidative stress is a problem in the industry because it can cause a decrease in protein production. Mesobiliverdin IXα and biliverdin IXα possess antioxidant capabilities. The effects of the antioxidant nature of these compounds were tested on the CHO 1-15 cell line. Cell cultures from silkworm and spider silk producing cells were also pursued. Methods to produce a primary cell line from spider silk gland cells were developed. Cell lines from spider and silkworm silk producing glands appeared to have the capacity to secrete full-length native proteins ranging in size from 200 to 500 kDa, and possibly larger.