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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: Modifying CHO using ‘Omics’ and Ideas From Novel Platforms

University of Manchester

Jun 18
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7:00am - 8:15am 75 mins
8:15am - 8:45am 30 mins
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: Modifying CHO using ‘Omics’ and Ideas From Novel Platforms
  • Alan Dickson, PhD - Director of Centre of Excellence in Biopharmaceuticals (COEBP), Professor of Biotechnology, University of Manchester
9:15am - 9:45am 30 mins
Horizon Discovery Spotlight Presentation
9:45am - 10:30am 45 mins
Networking Refreshment Break
10:30am - 11:00am 30 mins
Generation of Improved Host Cell Lines for Biomanufacturing using Vector and Cell Line Engineering Technologies
  • Dennis Pfaff - Investigator, Novartis

A toolbox of vector elements and novel engineered CHO cell lines were developed, which resulted in an increase of titer and improved product quality. By integrating these vector and cell line engineering technologies, we are aiming for further reducing time lines during cell line development.

11:00am - 11:30am 30 mins
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
CHO Informatics: A Case Study from Pfizer
  • Wei Wei - Principal Scientist, Pfizer
12:00pm - 12:30pm 30 mins
Sartorius Spotlight Presentation
12:30pm - 1:45pm 75 mins
1:45pm - 2:15pm 30 mins
Genome engineering to reduce viral particle release by CHO cells
  • Nicolas Mermod, Ph.D. - 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
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
Examining role of PKM1 in Lactogenic Behaviour and Metabolic Shift in CHO Cells, A Case Study
  • Shahram Misaghi, PhD - Senior Scientist, Genentech, Inc

Our studies revealed that pyruvate kinase muscle-1 (PKM1) expression correlates with lactogenic behaviour in CHO cells and deletion of PKM1 or PKM gene in CHO cells reduces lactate production and results in a metabolic shift in these cells.

For certain deletion configurations, this resulted in an increase productivity, which is likely due to the observed metabolic shift.

3:15pm - 4:00pm 45 mins
Networking Break
4:00pm - 4:30pm 30 mins
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
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