The complexity of biotherapeutics and the increasing demands of “faster, better, more economical” resonate with recombinant protein expression and production researchers. To meet these goals, protein scientists are exploring new engineering
tools. However, many variables still must be considered during the engineering process, including verification and sequence analysis of the gene or protein, codon optimisation, vector construction and clone/host selection. Ultimately, these tools
must be weighed against traditional expression and production strategies to achieve the desired quantity and quality.
Final Agenda
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MONDAY 18 NOVEMBER
12:00 Conference Registration (Foyer A)
13:30 Organiser’s Welcome
Mary Ann Brown, Executive Director, Conferences, Cambridge Healthtech Institute
13:35 Chairperson’s Opening Remarks
Cecília Maria Arraiano, PhD, Investigador Coordenador, ITQB-Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa
13:45 Engineering Vector Components and Host Cells for Next-Generation Bioproducts
David James, PhD,
Professor, Bioprocess Engineering, Chemical and Biological Engineering, University of Sheffield
Engineering complex cellular performance characteristics is an unpredictable challenge made more difficult by the variability of CHO cell lines, protein products, and production processes. There is no one-size-fits-all solution. As a new paradigm for
cell line development we are developing a hyper-variable design space for mammalian cell factory engineering that utilises directed and synthetic variation of chemical, genetic, and cellular input components as a core strategy to optimize cell functional
performance beyond natural limits.
14:15 Synthetic Biology Applied to Modulate Heterologous Gene Expression Using Portable mRNA-Stabilizing 5’-UTR Sequences
Cecília Maria Arraiano, PhD, Investigador Coordenador, ITQB-Instituto de Tecnologia Química e Biológica,
Universidade Nova de Lisboa
Cell survival requires continuous degradation and synthesis of RNAs. Transcript stability is generally missed for manipulation in synthetic designs. By keeping transcription fixed and combining different 5´untranslated regions (5´UTRs) and
constant RBS, mRNA decay was made the limiting factor of gene expression flow. This work resulted in a collection of well-characterized mRNA-stabilizing sequences that can be used as modules in any construct following the assembly rules of the Standard
European Vector Architecture (SEVA) format.
14:45 Precise Genome Engineering of Hybridomas for Antibody Expression and Screening
Cristina Parola,
PhD, Postdoctoral Research Scientist, Biologics Research, Sanofi
By taking advantage of precision genome editing with CRISPR-Cas9, we have developed a novel mammalian cell platform for the expression of full-length antibodies in hybridoma cells. The Plug-and-(Dis)play (PnP) workflow included the initial generation
of a reporter, antibody-negative cell line; in the subsequent reprogramming step, a novel specificity is introduced by means of a synthetic antibody. Finally, we optimized HDR efficiency to render the system amenable to the expression and screening
of B cell repertoires: this feature allowed the de novo discovery of antibodies from immune libraries.
15:15 Challenges and Strategies in Expression of a Bi-Specific Antibody
Jay Sinha, PhD, Associate Director, Cell Culture Process Sciences, Global Biopharmaceutical Development, Incyte Corporation
A process has been developed for producing Phase I clinical study material. Several expression platforms were examined for expression of a model bispecific antibody. The product quality and titer were found to vary with changes to host-vector system.
The challenges and strategies used for cell line and process development will be discussed.
15:45 Networking Refreshment Break (Foyer D)
16:15 Moderator’s Opening Remarks
Kerry Chester, PhD,
Professor, Molecular Medicine, University College London Cancer Institute
16:20 Bispecific, Soluble TCR as the Next Therapeutic Platform
Bahija Jallal, PhD, CEO and Director of the Board,
Immunocore
Of the two adaptive immunity recognition motifs, only antibodies have been brought to patients. However, antibody therapeutics only recognize 10% of human proteome (surface-expressed). The other motif, T cell receptor (TCR), has potential to unlock 90%
of the human proteome, but requires converting a low-affinity, specificity membrane receptor into a soluble therapeutic. IMCgp100, a soluble, TCR bispecific-targeting melanoma, is the most advanced soluble TCR therapeutic in development.
17:20 Attacking Cancer Cell Surfaceomes with Recombinant Antibodies
James A. Wells, PhD, Professor, Departments of Pharmaceutical Chemistry and Cellular & Molecular Pharmacology, University of California, San Francisco
The cell surface proteome (surfaceome) is the primary hub for cells to communicate with the outside world. Oncogenes are known to cause huge changes in cells and we find this translates to significant remodeling of the surfaceome. We generate recombinant
antibodies to detect and then attack these cells by toxifying the antibodies or recruiting immune cells to kill. I’ll discuss the technologies for surface protein analysis, an industrialized platform for rapid antibody generation using phage
display, and using these tool reagents for target validation.
18:20 Welcome Reception in the Exhibit Hall with Poster Viewing (Rio Pavilion)
19:30 End of Day
TUESDAY 19 NOVEMBER
07:45 Registration (Foyer A) and Morning Coffee (Foyer D)
08:30 Chairperson’s Remarks
Thomas Rexer, PhD, Team Lead, Dynamics of Complex Technical Systems, Bioprocess Engineering, Max Planck Institute
08:35 Integrating Cell-Free Expression, Purification, and Bioconjugation
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Marco G. Casteleijn, PhD, Senior Researcher, Industrial Biotechnology, VTT Technical Research Institute of Finland
We aim to develop new tools for cell-free protein synthesis. For example, to integrate protein expression, purification, and bioconjugation in small volumes coupled with cell-free protein synthesis. We compared light triggered release with traditional
affinity chromatography. Moreover, we explored transferring a moiety from a captured peptide to the target protein without further purification steps and used time gated Raman spectroscopy to evaluate protein quality.
09:05 Development of a High-Yield Cell-Free Synthesis Platform from Pichia Pastoris
Karen Polizzi,
PhD, Reader in Biotechnology, Department of Chemical Engineering, Imperial College London
Pichia pastoris (syn Komagataella spp.) is a methylotrophic yeast used in recombinant protein manufacture because of its high volumetric productivity. We have developed a CFPS platform using P. pastoris via optimisation of reaction conditions and vector design and overexpression of global regulators of ribosome synthesis to increase overall yields. The result is a system that is suitable for prototyping vectors before strain development or manufacturing
of proteins directly.
09:35 Problem-Solving Breakout Discussions
TABLE 9: Evaluating Protein Quality During Production in Real-time
Moderator: Marco G. Casteleijn, PhD, Senior Researcher, Industrial Biotechnology, VTT Technical Research Institute of Finland
- Reporter genes - good feed-back or just rumors?
- Raman Spectroscopy - quality confirmed or just exited molecules?
- Terahertz spectroscopy - protein quantification or too complex?
- Optical sensors - nice close-up or a murky view?
- Functionality assays - in vivo or sampling takes time?
TABLE 10: What Are the Promises of Synthetic Biology for Bioproduction?
Moderator: Philippe H. Jais, MD, PhD, President and CSO, Eukarÿs SAS
- Which product has benefited the most from synthetic biology? And in the future?
- What can genome editing do for bioproduction?
- Is bioproduction in mammalian cells the new frontier?
- What can non-natural nucleotide and non-natural amino-acids bring to bioproduction?
TABLE 11: Cell-Free Protein Expression (CFPS) as a Useful Tool for Protein-Protein Interaction Analysis
Moderator: Shayli Varasteh Moradi, PhD, Research Associate, Science and Engineering School, Queensland University of Technology
- Advantages of CFPS system in synthetic biology
- Using CFPS to study protein-protein interaction
- The application of CFPS in antibody and drug screening
10:30 Coffee Break in the Exhibit Hall with Poster Viewing (Rio Pavilion)
11:15 Synthetic Platform for in vitro Glycoengineering of Proteins by a Cell-Free, Compartmentalized Multi-Enzyme Cascade
Thomas Rexer, PhD,
Team Lead, Synthetic Glycobiotechnology, Bioprocess Engineering, Max Planck Institute for Dynamics of Complex Technical Systems Magdeburg
N-linked glycans attached to proteins are involved in a wide range of processes such as biological recognition, protein stability, immunogenicity, and antigenicity. Therefore, the glycosylation of proteins is an important parameter to be considered
in the optimisation of animal cell culture-derived drugs including monoclonal antibodies. The presented cell-free system is an integral part of a synthetic platform for in vitro glycoengineering of proteins by
model-supported, cost-efficient and scalable biocatalytic processes being established by our group.
11:45 Cell-Free Based Approach for Rapid Screening of Antibody Fragment Libraries
Shayli Varasteh
Moradi, PhD, Research Associate, Science and Engineering School, Queensland University of Technology
Cell-free protein expression system (CFPS) allows the robust production of recombinant proteins in a multiplexed format. We developed a rapid method for antibody fragment libraries screening based on eukaryotic Leishmania tarentolae (LTE)
system in combination with AlphaLISA technology to study protein-protein interaction. The presented technique provides a powerful tool for rapid protein binders’ selection with high sensitivity and throughput.
12:15 Enjoy Lunch on Your Own
13:45 Dessert Break in the Exhibit Hall with Poster Viewing (Rio Pavilion)
14:15 Chairperson’s Remarks
Bjørn Voldborg, MSc, Director, CHO Cell Line Development, The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark
14:20 KEYNOTE PRESENTATION: Unique Engineering Targets for Antibody Production Cell Lines: Selection, Cloning, Glycan Modifications, and Chromatin Readers
Volker Sandig, PhD, CSO, ProBioGen AG
Omics approaches are often applied to determine holistic strategies to improve key cell line attributes: yield, stability, robustness, and product quality. Instead, we have selected important junctions in known pathways to enhance cell line performance.
We will show how transgene cassettes embedded into transposons can be directed to most active genomic loci taking benefit of natural chromatin reader domains, discuss the impact for bispecific antibodies, and look into pathway deflection to set
specific glycan features.
14:50 Development of a Pre-Glycoengineered CHO-K1 Host Cell Line for the Expression of Antibodies with Enhanced Fc Mediated Effector Function
Oliver Popp, Dr. rer.
nat., Principle Scientist, pRED, Large Molecule Research, Roche Diagnostics GmbH, Roche Innovation Center Munich
Here, we present the development of a glycoengineered CHO-K1 host cell line, stably expressing β1,4-N-Acetylglucoseaminyltransferase III and α-mannosidase II, for the expression of a-fucosylated antibodies with enhanced Fc-mediated effector
function.
15:20 Expanding the CHO Cell Line Development Toolbox to Enable Fast-Track Development of Innovative Biotherapeutics
Valerie Schmieder, PhD, Postdoctoral Researcher, Cell Line Development, Bioprocess Development Biologicals, Boehringer Ingelheim Pharma GmbH & Co. KG
The increasing demand for novel biotherapeutics is driving the generation and implementation of innovative as well as disruptive tools for cell line development (CLD) in CHO. Additionally, more and more complex molecules, such as multi-specific antibodies,
are further challenging the production of therapeutic proteins from CHO. Here, we present our recent achievements in the use of state-of-the-art technologies to overcome current and future challenges in CLD.
15:50
Presentation to be Announced
16:20 Refreshment Break in the Exhibit Hall with Poster Viewing (Rio Pavilion)
17:00 Combining a CRISPR Library with Phenotypic Enrichment to Identify Gene Engineering Targets in CHO Cells
Niamh Keogh, Research
Scientist, Niall Barron Laboratory, Chemical and Bioprocess Engineering Department, National Institute for Bioprocessing Research & Training
CRISPR Technology has the ability to fundamentally change the capabilities of genetic engineering. My work focuses on using CRISPR/CAS 9 to generate individual knock outs of genes as well as using a CRISPR Library approach to create genome-wide loss
of gene function studies with the overall aim of discovering potentially beneficial gene targets for CHO cell line engineering.
17:30 Studying the Impact of Genetic Alterations Using a Targeted Integration CHO Host Cell Line
Mark Trautwein, Dr. rer. nat., Senior Scientist, Biologics Research, Bayer AG
Both the chromosomal environment of the integration site as well as the genetic elements of a transgene expression cassette contribute to the degree of high and stable transgene expression. We have used targeted integration host cell lines for evaluation
of different genetic elements of the transgene construct as well as for Crispr/Cas9-based implementation of targeted (epi)genetic modifications. This approach facilitates optimization of product-specific expression configurations.
18:00 Rethinking Gene Expression Using the Synthetic C3P3 Transcription System
Philippe H. Jais, MD, PhD, President and CSO, Eukarÿs SAS
Eukarÿs has developed the first ever artificial expression system by synthetic biology that is named C3P3 (cytoplasmic chimeric capping prone-phage polymerase). This enzymatic system, currently in its 3rd generation, synthesizes in vivo high amounts of mature messenger RNA and, consequently, protein of interest in mammalian cells. Besides its uses for therapeutics, the C3P3 system is used as a potent tool for the bioproduction of viruses and proteins.
18:30 Cell- and Vector-Engineering for Manufacturing Gene Therapy Lentiviral Vectors from Constitutive Producer Cell Lines
Ana Filipa Rodrigues, PhD, Research Scientist, Animal Cell Technology Unit, Cell Line Development and Molecular Biotechnology Laboratory, iBET - Instituto de Biologia Experimental e Tecnológica
Lentiviral vectors (LV) are currently the first choice for cell and gene therapy applications when long-term expression is required. To support robust LV manufacture, we present three novel approaches: i) LentiPro26, stable and constitutive packaging
cells, based on a mutated lentiviral gag-pro-pol and a stringent selection method; ii) high-titer chimeric envelops for three different pseudotypes, based on protein engineering; and iii) producer cell genome editing and engineering for increased
robustness to vector component’s cytotoxicity.
19:00 End of Cell Lines and Systems Engineering