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Key Sessions

Nathan Lewis, PhD

KEYNOTE: A Systems Approach To Engineering Protein Production in Mammalian Cells

University of California, San Diego

Alan Dickson, PhD

KEYNOTE: CHO Cells - Too Good To Be Wrong

University of Manchester

Jun 18
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7:00am - 8:15am 75 mins
Registration
8:15am - 8:45am 30 mins
Info
KEYNOTE: A Systems Approach To Engineering Protein Production in Mammalian Cells
  • Nathan Lewis, PhD - Associate Professor, Department of Pediatrics & Bioengineering, University of California, San Diego

The majority of biotherapeutics are produced in mammalian cells. To facilitate cell line development, we are mapping the pathways involved in mammalian cell growth and protein production. We have used these with genome editing techniques to develop cell lines with improved traits for protein production.


8:45am - 9:15am 30 mins
KEYNOTE: CHO Cells - Too Good To Be Wrong
  • Alan Dickson, PhD - Director of Centre of Excellence in Biopharmaceuticals (COEBP), Professor of Biotechnology, University of Manchester
9:15am - 9:45am 30 mins
Info
Improving the CHO Cell Host Using Different Genome Editing Approaches
  • Mario Pereira - Field Application Scientist, Horizon Discovery

Horizon Discovery has optimised cell-based assays which will be combined with our expertise in CRISPR screening to identify novel genetic targets that will improve the CHO cell host for biomanufacturing. We will describe the outcomes of our proof of concept work and how we will translate these findings into new commercially available cell lines.

9:45am - 11:00am 75 mins
Networking Refreshment Break
11:00am - 11:30am 30 mins
Info
CRISPR/Cas9 Platform Development for Drug Target Discovery and Validation
  • Benjamin Haley, PhD - Senior Scientist, Genentech, Inc

Adaptation of CRISPR/Cas9, a bacterial genome defense system, for eukaryotic molecular genetics has ushered in a new phase of the genomics revolution. Here, I will present the framework of a CRISPR/Cas9-based platform for drug target discovery and validation and how this platform can be exploited for single gene to genome-scale experimentation.

11:30am - 12:00pm 30 mins
Info
Development of CHO Informatics To Support CHO Cell Line Characterization
  • Wei Wei, PhD - Principal Scientist, Pfizer

Omics-based approaches have provided key tools to understand the underlying biological processes critical to bioprocessing. Developing effective bioinformatics tools for the analysis and integration of complex omics is essential.  Case studies will be presented to highlight how we develop sophisticated and state-of-the-art pipelines for the in-depth characterization of recombinant CHO cell lines expressing biotherapeutic targets.

12:00pm - 12:30pm 30 mins
Info
Small scale perfusion-mimic processing in ambr 15 cell culture micro bioreactor system; improving screening capability for intensified processes
  • Alison Rees-Manley - ambr 15 Product Manager, Sartorius Stedim Biotech

With the growing trend in the Biopharma market towards development of intensified processes, high throughput tools offer many advantages to speeding up development timelines.  Identifying key parameters for optimisation and selecting the best media and feeds for intensified processes, utilizing small scale models, cannot only save considerable time, but also presents a cost-effective way of understanding which conditions should be implemented at larger scale. 

Current tools are insufficient to effectively screen intensified process parameters at the micro bioreactor scale in an automated fashion. The newly launched capabilities of the ambr 15 offer many advantages for screening conditions, media and feeds, due to automation, parallel operation and low working volumes.  In this talk the capabilities of the ambr 15 system will be discussed along with their benefits to modelling intensified processes including data illustrating different approaches to running perfusion cultivations in ambr. 

12:30pm - 1:45pm 75 mins
Luncheon
1:45pm - 2:15pm 30 mins
Info
Genome engineering to reduce viral particle release by CHO cells
  • Nicolas Mermod, PhD - Professor, Director, Institute of Biotechnology, University of Lausanne, Switzerland

CHO cells are known to express endogenous viral elements embedded in their genome, and to release retroviral-like particles in the culture supernatant. This complicates the detection of potential contamination by viral adventitious agents, and, despite the lack of evidence of infectivity of these particles, raises safety and regulatory concerns.

Using Next generation sequencing approaches, we characterized several families of endogenous retroviral elements (ERVs) present in CHO-K1 cell genome.

We focused on one highly conserved ERV group of the Gammaretrovirus gender, as it was potentially functional, giving rise to viral-llike particle containing viral genomic RNA. Transcriptome and viral particle analysis validated the functionality of ERVs from this group, and it further indicated that the mRNA and viral genome are expressed from few (approximately 4) ERV sequences.

Using CRISPR-Cas9-mediated CHO genome engineering, we mutagenized the conserved ERV sequence group. Comparison of genomic and viral particle sequences allowed the identification of one ERV that encodes the viral genome of corresponding retroviral particles. We show that particular mutations within this ERV suffice to decrease the release of genome-loaded viral particles by several orders of magnitude.

2:15pm - 2:45pm 30 mins
Info
Glycoengineering CHO Cells For The Production Of Therapeutic Glycoproteins
  • Bjørn Voldborg - Director CHO Cell Line Development, The Novo Nordisk Foundation Center for Biosustainability

At CFB we have used our high throughput cell engineering pipeline to generate a panel of engineered CHO cells with improved properties for the production of recombinant therapeutic proteins.

We now have a large collection of cell lines generating tailored glycoprofiles, and we have used these to produce a therapeutic protein with a defined N-glycan profile, matching the current plasmas derived product.

2:45pm - 3:15pm 30 mins
Info
Expression of highly glycosylated viral antigens in mammalian cells
  • Sara O'Rourke, PhD - Project Scientist, The University of California at Santa Cruz

The significance and difficulties associated with the expression and purification of appropriately glycosylated viral antigens has only became clear in recent years. Combining high efficiency electroporation using the MaxCyte STX and Robotic selection using a Molecular Devices ClonePix2, we achieved high level expression of a series of highly glycosylated HIV-1 Envelope glycoproteins both in transient and stable platforms.  The methodology was readily transferable on the transient level, to other viral antigens and immune molecules.  New stable lines and potential therapeutics are in the early stages of generation.

3:15pm - 4:00pm 45 mins
Networking Break
4:00pm - 4:30pm 30 mins
Info
Optimization of Expression Vector Design to Streamline the Transition from Research to Clinical Cell Line Development
  • Yizhou Zhou, PhD - Scientist II, Cell Culture Development, Biogen

Expression vector design plays a key role for efficient recombinant protein expression in Chinese Hamster Ovary cell lines. Here we evaluated vector design strategies including signal peptide and codon editing choices as well as vector construction approaches that accelerated the transition of lead molecules from Research and optimized clinical cell line development.

4:30pm - 5:00pm 30 mins
Info
Glycoengineering of The Human Embryonic Kidney FreeStyle 293-F Cell Line Towards Prolongend Bioavailablity of Recombinant Coagulation Factor VII
  • Rico Uhler - PhD Candidate, Mannheim University of Applied Sciences & Octapharma Biopharmaceuticals

Cell line engineering techniques like CRISPR/Cas9 can efficiently alter the glycosylation of recombinant proteins by modifying the gene expression of glycosyltransferases, glycosidases or proteins involved in transport and metabolism of glycosylation precursors in producer cell lines. N‑glycosylation is particularly important for the pharmacokinetics of recombinant glycoprotein therapeutics and can prolong dosing intervals to the benefit for patients, especially when administration is performed intravenously.

Recombinant glycoproteins produced in non-human cells may carry glycostructures antigenic to humans. Using the human embryonic kidney FreeStyle 293-F (HEK 293-F) cell line as expression platform offers the advantage of human-type glycosylation patterns. Therapeutic coagulation factor VII (FVII) can prevent and treat bleeding episodes in patients with congenital FVII deficiency, Hemophilia A or B with inhibitors to Factor VIII or Factor IX and acquired hemophilia. However, FVII expressed in the HEK 293-F cell line carries considerable levels of terminal N-acetylgalactosamine (GalNAc) on its N-glycans.

In order to lower binding of FVII to the hepatic asialoglycoprotein receptor (ASGP-R), and thus reduce FVII clearance, two GalNAc transferases were knocked-out in the HEK 293-F cell line by CRISPR/Cas9. Effects of B4GalNT3 and B4GalNT4 single or double knock-outs on released N-glycan level of purified FVII were analyzed by hydrophilic interaction liquid chromatography coupled with mass spectrometry. Our results revealed a successful reduction of terminal GalNAc accompanied by an increase in terminal galactosylation and, beneficially, N-glycan sialylationl. N-glycan profiles were correlated with ASGP‑R binding measured by surface plasmon resonance and pharmacokinetics assayed in a rat model. Reduced GalNAc levels and increased sialylation led to lower ASGP-R binding and improved FVII recovery in vivo by more than 50 %. Using the example of FVII expressed in a modified HEK 293-F cell line, we demonstrated that glycoengineering via cell line design is a straightforward and rational approach to optimize the glycosylation profile of protein therapeutics using state-of-the-art gene-editing tools.

5:00pm - 6:00pm 60 mins
Networking Reception in Poster & Exhibit Hall