In recent work we have identified design faults in commonly used bacterial expression plasmids. We have used modern methods for DNA assembly and directed evolution to correct these design faults. The new designs are a simple and effective way to increase protein production yields in bacterial cell factories. 

The increasing demands for protein reagents needed for drug discovery efforts drive the development of an efficient methodology for recombinant protein production. Within the BioMolecular Resources Department at Genentech, we have developed a platform for high-throughput construct generation and expression screening in baculovirus, E. coli and mammalian transient systems applicable to all protein types and localizations.

The recombinant adeno-associated virus (AAV) vector has emerged as a promising gene therapy platform due to its robust, sustained, and tissue-targeted gene delivery. One major hurdle for clinical AAV application is large-scale manufacturing. Here we present a scalable AAV vector production method using chemically defined, high-density insect cell-based expression system.

This presentation shows how to simplify the drug development route to the clinic, without compromising on deliverables, ensuring fast timelines and high titers ahead of industry standards.

• How to avoid royalties, milestones and licence fees during cell line development
• How utilizing ultra-high throughput productivity and scalability assessment ensures high titers can be achieved on a fast timeline
• Why using a CHO-K1 cell line ensures better stability, protein folding and processing

Strain selection is a critical step which often gets locked down at an early point during microbial process development, with an emphasis on product titer. However, this is accompanied by a certain amount of risk if product quality is not sufficiently evaluated. A new platform approach to accelerated strain selection, based on both product titer and quality evaluation, leveraging high through put tools and robotics, will be presented.

Transfections of large amounts of mRNA can be used to assess compatibility of cell line and recombinant protein while mimicking high productivity, circumventing the need to establish high producing clones early on in product development. By gradually increasing Epo-Fc mRNA load in CHO cells, bottlenecks in N-glycosylation were detected. Furthermore, we report how this system could verify optimization of N-glycan processing capacities by modulating the expression of key glycosyltransferases.

Using a tetracycline-inducible expression system (Tet-On) we show that expression of recombinant antibodies in CHO cells is proportional to their mRNA level. In these cells, induction leads to a systematic and extensive reprogramming of the transcriptome, shifting the total expression of constitutive CHO genes towards the production and secretion of the desired antibody. This multi-omic study provides insight into the impact of biotherapeutics production on CHO cell physiology.

Lipid metabolism plays a key role in cellular processes central to achieving high recombinant protein titres. Processes such as secretion, cell division and endoplasmic reticulum size and function are highly dependent on lipids and their metabolism. We have used genetic engineering to manipulate lipid metabolism in CHO cells, targeting SCD1 and SREBF1 expression, to expand the endoplasmic reticulum and ultimately enhance secretory recombinant protein titres between 1.5 and 9-fold.

The extensive post-translational processing of clusterin, coupled with its potent binding to any misfolded protein, have meant that its expression as a fully functional recombinant protein has been very difficult and this has limited structure-function studies. We developed a new rapid mammalian cell expression/purification system to accomplish this which has a yield of the order of 30-40 mg per litre of culture and can be completed in about one week.

Influenza viruses infect millions of people each year, resulting in significant morbidity and mortality in the human population. Herein, we describe the approach for developing stable transfected human cell lines for the expression of recombinant influenza virus hemagglutinin (HA) and recombinant influenza virus neuraminidase (NA) proteins for the purpose of in vitro and in vivo vaccine development. Our platform can be easily adapted for the production of other pathogen-related proteins.

Selecting the right CDMO partner is critical for any institution as the decision can impact product quality, cost and timeline.
This presentation will delve into the CDO Upstream Process at Samsung Biologics which helps customers achieve high quality products with accelerated timelines.