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Key Sessions

Nicole Borth

Genome Scale Science For CHO: From Chasing The High Productivity Miracle Gene To Exquisite Phenotype Control

BOKU University and ACIB

Nicolas Mermod

Engineering Strategies to Suppress Viral Particle Release from CHO Cells

University of Lausanne, Switzerland

08:20 - 08:30 10 mins
Chairperson's Opening Remarks
08:30 - 09:00 30 mins
Info
New Technologies: Cell Line and Host Cell Engineering
Genome Scale Science For CHO: From Chasing The High Productivity Miracle Gene To Exquisite Phenotype Control
  • Nicole Borth - Professor, BOKU University and ACIB

Traditionally in cell line development and engineering, there was a quest for individual genes which, if overexpressed or knocked out, would enhance performance, with very limited success. However, high level performance is more likely to be the result of the perfect combination of expression level of multiple genes. Today, with detailed knowledge of genomes, transcriptomes and the regulatory mechanisms that define the later, and with new tools available to manipulate these, the exquisite control of phenotypes starts to become a realistic target.

09:00 - 09:30 30 mins
Info
New Technologies: Cell Line and Host Cell Engineering
Technology Toolbox for Cell Line Development
  • Holger Laux - Fellow, Novartis Cell Line Develeopment

Chinese Hamster Ovary cells are widely used for large-scale production of recombinant biopharmaceuticals. We will present that applying transcriptomics derived approaches supported the identification of the root cause of cell growth inhibition and low productivity of a difficult to express therapeutic protein and how state of the art cell line engineering tools enabled the high expression of this therapeutic protein. Especially the combination of the recently published CHO genome with screening methods and cell line engineering tools has enabled the development of superior CHO cell lines.

09:30 - 10:00 30 mins
New Technologies: Cell Line and Host Cell Engineering
Spotlight Presentation
10:00 - 10:45 45 mins
New Technologies: Cell Line and Host Cell Engineering
Morning Coffee & Poster Tour 1
10:45 - 11:15 30 mins
Info
New Technologies: Cell Line and Host Cell Engineering
Structural and epitranscriptomic manipulation of RNA to enhance transgene expression
  • Niall Barron, PhD - Professor of Biochemical Engineering, University College Dublin and Principle Investigator, NIBRT

The translation of circular mRNA represents an interesting possibility for improving recombinant protein production. Initiation and termination are two rate limiting steps of translation therefore by encoding a gene lacking a stop codon on a circular RNA molecule we generated an infinite EPO open reading frame and demonstrate the production of secreted protein from a circular mRNA in mammalian cells. In addition the use of an epitranscriptomic approach to improving gene expression will be presented.

11:15 - 11:45 30 mins
Info
CRISPR and Genome Editing Technologies
Application on CRISPR Cas9 to Accelerate Cell Line Development and Engineering

If you are interested in presenting on this topic please contact: catherine.marshall@knect365.com

11:45 - 12:00 15 mins
CRISPR and Genome Editing Technologies
Spotlight Presentation
  • A Representative from Polyplus - -, Polyplus
12:00 - 12:15 15 mins
CRISPR and Genome Editing Technologies
Novel Technologies for Cell Line Development and Engineering
  • A Representative from Selexis - -, Selexis
12:15 - 13:30 75 mins
CRISPR and Genome Editing Technologies
Lunch and Live Labs
13:30 - 14:00 30 mins
Info
CRISPR and Genome Editing Technologies
Engineering Strategies to Suppress Viral Particle Release from CHO Cells
  • Nicolas Mermod - 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. Transcriptome and viral particle analysis validated the functionality of ERVs from this group, and it further indicated that the mRNA and viral genome may be expressed from few (approximately 3) 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 viral genome-loaded particles below detection limits.

14:00 - 14:30 30 mins
Info
Omics and Big Data Integration in Cell Line Development and Engineering
Characterizing Clone Performance: A Union of Structure and Sequencing
  • Steven Huhn - Senior Scientist, Merck
  • Higher order chromatin territories and chromatin features within CHOK1 chromosomes
  • High resolution 3D FISH combined with nuclear architecture to characterize transgenic loci
  • High resolution chromatin architecture interfaced with recombinant DNA
  • Epigenetic characterization of transgenes
  • Genetic characterization of clones using next-generation sequencing platforms
  • Differences in rDNA integration between linear and integrase treated

The generation of a recombinant CHO lineage relies on the integration of a transgene into the donor genome, an approach which results in transgenic heterogeneity. Requirements for predictable transgene expression demands higher level knowledge of the interplay behaviour of recombinant DNA and the CHO genome.  Here, we map higher level chromatin features to the CHO genome and develop a contiguous, CHO chromosome specific epigenome map. We also utilize super-resolution confocal microscopy and FISH to characterize the interplay of chromatin and chromatin territories with recombinant DNA. We utilize this data in conjunction with whole genome and transcriptome sequencing to predict clone performance.

14:30 - 15:00 30 mins
Info
Systems and Synthetic Biology Applications
Putting The Horse Before The Cart: Designing Gene Expression Constructs For CHO Cell Engineering
  • Adam Brown, Ph.D. - Lecturer of DNA Engineering, University of Sheffield

Manufacture of non-natural, engineered protein formats will require purpose-built designer cell factories. While a great deal of effort has been devoted to identifying target genes for CHO cell engineering, relatively little has been spent figuring out to optimally express them. This presentation discusses design rules for creating plasmid vectors encoding complex gene combinations, describing how genetic component compositions can be designed and configured in order to optimize the performance of engineered cell factories.

15:00 - 15:30 30 mins
Systems and Synthetic Biology Applications
Spotlight Presentation
  • A Representative from Sartorius - -, Sartorius
15:30 - 16:00 30 mins
Systems and Synthetic Biology Applications
Afternoon Coffee and Poster Tour 2
16:00 - 16:30 30 mins
Info
Systems and Synthetic Biology Applications
The Relevance Of Cell Size In A CHO Fed Batch Process: Metabolic And Transcriptomic Characterization
  • Dirk E. Martens - Associate Professor, Bioprocess Engineering, Wageningen University

In a fed-batch process, using a commercially available media system a switch is observed from a cell proliferation phase to a phase where cell division is arrested and cell growth continues in the form of a threefold increase in cell size and dry weight. Metabolic flux and transcriptome analysis is applied to better understand the biological mechanisms associated with this switch.

16:30 - 17:00 30 mins
Info
Systems and Synthetic Biology Applications
Panel Discussion: Integrating New Engineering and Cell Line Development Technologies into Established Workflows
  • What new technologies have been adopted by industry and used routinely in cell line development?
  • Success rates and realities of engineering tools? e.g. genomics, genome editing, CRISPR, NGS, targeted integration
  • What is the industry status on CRISPR adoption?
  • Are titres and timelines better with new technologies compared to standard approaches?
  • Specific site integration vs. random integration: What is the impact on expression level and expression stability?
  • What are the best, efficient and smart ways to use cell line engineering in a workflow?
  • Where and how are industry incorporating new technologies and engineering for clinical and pipeline products when timelines are critical?
  • Feasibility of cell line engineering: Are people doing customised cell line engineering for individual projects and is this feasible?
  • Industry opinions on engineering: Engineer the host or look for replacements?
17:00 - 17:05 5 mins
End of Cell Line Development & Engineering 2019
08:20 - 08:30

Chairperson's Opening Remarks

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17:00 - 17:05
End of Cell Line Development & Engineering 2019

End of Cell Line Development & Engineering 2019

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