Sixth Annual Protein Expression
Part One: Enhancing Expression and Achieving Higher Throughput
Day 1 | Day 2
Part Two | Download Brochure
Tuesday, 6 November
7:30 Registration and Morning Coffee
8:30 Chairperson’s Remarks
M. Raafat El-Gewely, Ph.D. Professor, Institute of Medical Biology, University of Tromsø
» Keynote PRESENTATION
8:35 Protein Expression in Drug Discovery – New Challenges, New Solutions
Lorenz M. Mayr, Ph.D., Vice President, Biological Reagents & Assay Development, AstraZeneca R & D
Whereas protein expression has long been viewed as a mature science with no need for further improvement, current trends in drug discovery show an increased demand for fast & efficient production systems for recombinant proteins and protein complexes to cope with the demands for protein in sufficient amounts needed for modern hit discovery (HTS, FBS, structure) and lead optimization in discovery research.
9:05 FreEcoli™: A New Series of Escherichia coli Strains for Endotoxin-Free Production of Recombinant Proteins and Plasmid DNA
Uwe Mamat, Ph.D., Research Scientist, Structural Biochemistry, Research Center Borstel
We present here the FreE coli™ set of non-conditional E. coli derivatives that lack all outer membrane agonists for hTLR4/MD-2 activation. The FreE coli™ strains entirely lack LPS, yet remain viable despite exclusively elaborating the tetra-acylated, endotoxically inactive lipid A precursor lipid IVA. Consistent with the results that activation of hTLR4/MD-2 signaling by FreE coli™ cells was of several orders of magnitude lower compared with cells of the E. coli wild-type strain, heterologous proteins and monomeric, supercoiled plasmid DNAs prepared from different FreE coli™ strains were free of endotoxic activity.
9:35 Host Cell and Expression Engineering for Development of an E. coli Ketoreductase Catalyst: Enhancement of Formate Dehydrogenase Activity for Regeneration of NADH
Regina Kratzer, Ph.D., Researcher, Institute of Biotechnology and Biochemical Engineering, Graz University of Technology
An E. coli co-expressing Candida tenuis xylose reductase and Candida boidiniiformate dehydrogenase (FDH) was developed. Single expression of the FDH gave an enzyme activity of 400 units/gCDW. Co-expression, however, resulted in an 80% decline in FDH activity. Combined effects from increase in FDH gene copy number, supply of rare tRNAs and dampened expression of the ketoreductase brought up the FDH activity 3-fold.
10:05 Does Metabolism Limit Recombinant Protein Production?
Lars Blank, Ph.D., Chair of Applied Microbiology, RWTH Aachen University
The synthesis of proteins is one of the most energy consuming processes in the cell, with the result that cellular energy supply may become critical. We therefore quantified the impact of recombinant protein production on microbial metabolism. The insights into the operation of metabolism during recombinant protein production might guide the optimization of microbial hosts and fermentation conditions
10:35 Coffee Break in the Exhibit Hall with Poster Viewing
11:15 microRNAs in CHO Cell Culture Technology
Matthias Hackl, Ph.D., Research Assistant, Department of Biotechnology, University of Natural Resources and Applied Life Sciences, Vienna
The main impact of the now published genome, transcriptome and miRNome of CHO cells is that it enables researchers to begin to understand the molecular mechanisms of how these cells perform their tasks of efficient growth, high productivity and safe product quality. The focus of this presentation is how this information has improved our ability to use microRNAs for engineering of CHO cells.
11:45 Recombinant IgAs Expressed in CHO Cells: Bottlenecks of Recombinant Cell Lines and Improved Production Strategies
Renate Kunert, Ph.D., Professor, Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna, Austria
Immunoglobulins of subtype A (IgA) mediate a key role in mucosal immunity and are promising new immunotherapeutic candidates. We established recombinant Chinese hamster ovary (CHO) cells which stably expressed two different IgA1 antibodies under serum-free conditions. In this study, we extensively characterized the low and the high producing cell lines in long-term culture and identified bottlenecks in polypeptide expression and assembly.
12:15 Luncheon Presentation I
“Open Sourcing” - Increasing Options to Meet the Challenges of Drug Development and ManufacturingSimon Boa, Ph.D., Director, Business Development & Marketing, Merck Millipore
We’ll present a case study how a “open-sourcing” solutions based approach enabled the development of a clinical manufacturing process for a new drug in parallel with the construction of a new clinical manufacturing facility.
13:00 Luncheon Presentation II or Lunch on Your Own(Opportunity available, please contact Carol Dinerstein, Dinerstein@healthtech.com)
14:15 Chairperson’s Remarks
14:20 Improving Transient Gene Expression in Mammalian Cells: The Case of a Human Lysosomal Enzyme
José Luis Corchero, Ph.D., CIBER de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Barcelona
Transient gene expression (TGE) is a useful, fast method to produce recombinant proteins. This work describes TGE-based production of a human enzyme. Several parameters (cells, vectors, and others influencing cell metabolism) were studied and optimized. The proposed protocol allows producing up to several mg/L of active enzyme (or engineered versions) to be tested in vitro or in pre-clinical trials.
14:50 CHO Genome Characterization and Engineering to Improve Protein Synthesis and Secretion
Nicolas Mermod, Ph.D., Professor, Laboratory of Molecular Biotechnology, University of Lausanne
We have determined the genome sequence of a CHO cell line used for pharmaceutical production and of derived producer clones. This allowed the characterization of the alterations of the genomic and transgene sequence of candidate cell lines for screening purposes as well as cell engineering for increased protein secretion and modifications. This presentation will illustrate how a systematic and multi-level approach can be used to improve the expression of pharmaceutical proteins, including difficult-to-express ones.
15:20 Rational Engineering of Escherichia coli Strains for Plasmid Biopharmaceutical Manufacturing
Geisa Gonçalves, Researcher, Department of Bioengineering, Instituto Superior Técnico (IST), Lisbon, Portugal
15:50 Refreshment Break in the Exhibit Hall with Poster Viewing
16:30 Rapid Identification of High Affinity Monoclonal Antibodies Using the ForteBio Octet RED384 Instrument
Christian Frisch, Ph.D., Director, Research & Development, AbD Serotec, A division of MorphoSys AG
We present an off-rate screening method of crude E. coli lysates containing monovalent Fab fragments obtained after phage display of the HuCAL® antibody library. After antibody selection and ELISA-based primary screening, antibodies are ranked according to their kinetic off-rates on the ForteBio Octet RED384 instrument. The data show good correlation of kinetic parameters determined in lysates and in purified samples, thus enabling off-rate based ranking of unpurified antibodies at the screening stage.
17:00 Problem Solving Roundtable Discussions
Table 7: Expressing Protein Complexes for Drug Discovery
Moderator: Lorenz M. Mayr, Ph.D., Executive Director, Unit Head Biology,Protease Platform, Novartis Pharma
Table 8: Solving Expression Problems in CHO Cells
Moderator: Renate Kunert, Ph.D., Professor, Department of Biotechnology,University of Natural Resources and Life Sciences, Vienna, Austria
Table 9: Troubleshooting Transient Gene Expression in Mammalian Cell Lines
Moderator: José Luis Corchero, Ph.D., CIBER de Bioingeniería, Biomateriales yNanomedicina (CIBER-BBN), Barcelona
Table 10: Genome Engineering to Improve Expression
Moderator: Nicolas Mermod, Ph.D., Professor, Laboratory of Molecular Biotechnology, University of Lausanne
Table 11: Cell Free or Not Cell Free: Making the Decision/Transition
Moderator to be Announced
18:00-19:00 Welcome Reception in the Exhibit Hall with Poster Viewing
Day 1 | Day 2
Part Two | Download Brochure