Cambridge Healthtech Institute’s 6th Annual
Optimizing Expression Platforms
Tools for Effective Expression, Production, and Purification
January 17-18, 2019
The utilization of engineered therapeutic proteins for basic research, clinical diagnostics, and therapy continues to expand. Consequently, protein expression laboratory managers and researchers face challenges for efficient expression, production, and
purification even while improving quantity and quality and minimizing time and cost. Transient protein production (TPP) has the advantage of speed and limiting risk while stable transfection – the longer and more complex process – has
the advantage of producing long-term expression of the biotherapeutic of interest. The rapidly increasing need for recombinant proteins necessitates further improvements in both technologies.
Cambridge Healthtech Institute’s 6th Annual Optimizing Expression Platforms conference convenes protein expression specialists who share their experiences with process and platform differences, tradeoffs, and improvements for producing recombinant
proteins in their expression and production laboratories.
Final Agenda
THURSDAY, JANUARY 17
7:45 am Registration and Morning Coffee (Sapphire West Foyer)
8:10 Organizer’s Welcome Remarks
Mary Ann Brown, Executive Director, Conferences, Cambridge Healthtech Institute
8:15 Chairperson’s Opening Remarks
Howard R.G. Clarke, PhD, Principal Scientist, Cell Sciences, Seattle Genetics
KEYNOTE PRESENTATION
8:20 Chromosome Stability Approach: Lengthening of High-Yield Production Levels of IgG-Producing CHO Cells by Downregulation
of Breast Cancer 1
Takeshi Omasa, PhD, Professor, Department of Material and Life Science, Graduate School of Engineering, Osaka University
The effects of breast cancer 1 (BRCA1) downregulation on gene amplification efficiency and long-term productivity were investigated in CHO cells. Our results suggest that high-producing cells, which maintain their productivity long term, were efficiently
established by BRCA1 downregulation. In this presentation, I would like to introduce the chromosome stability and effect of BRCA1 downregulation.
9:00 Antibody Expression Stability in CHO Clonally Derived Cell Lines and Their Subclones
Howard R.G. Clarke, PhD, Principal
Scientist, Cell Sciences, Seattle Genetics
Cell line development involves lengthy screening to identify a stable line having consistent growth, productivity and product quality. To investigate production stability in CHO cells we analyzed primary clones and their respective subclones. Cell lines
derived from single cell progenitors grow into populations of cells with phenotypic heterogeneity. Here I present the genetic and epigenetic characterization of these heterogeneous cell line populations.
9:30 Three Aspects for Your Cell Line Success: Titer, Speed & Quality
Bernd Rehberger, Lead Scientist, Cell Line Development, Sartorius Stedim Cellca GmbH
Ensuring biologics are expressed with a high titer as well as the required quality is becoming more important. The CellcaCHO™ expression platform delivers increased productivities, easier expression of complex proteins & shortened timelines
to clinic. The timeline of 4 months until delivery of the RCB, including an upstream process, was achieved by the optimization of the expression vector system, implementation of state-of-the art down-scale models & integration of fast screening
methods for quality assessment.
10:00 Coffee Break in the Exhibit Hall with Poster Viewing (Sapphire Ballroom)
11:00 NEW: SELECTED POSTER PRESENTATION: Implementing an Automated Platform for Cell Line Development
Xiaoyan Tang, MD, Senior Scientist, Cell Line Development, Merck Research Labs
11:30 Engineering a Stable CHO Cell Line for the Expression of a MERS-Coronavirus Vaccine Antigen
Wen-Hsiang Chen, PhD, Assistant Professor, Department of Pediatric, Section of Tropical Medicine, Baylor College of Medicine
Human vaccine against MERS-CoV is not available. We have developed a stably transfected adherent CHO cell line for the production of the MERS-CoV protein subunit, S377-588 (Fc tagged). The adjuvanted protein vaccine expressed in adherent CHO could protect
transgenic animal model from infection with live MERS-CoV. We also have developed a suspension monoclonal CHO cell line able to express S377-588-Fc in serum-free media, which is ready for scaled-up production.
12:00 pm Session Break
12:10 Luncheon Presentation I: New Tools for Screening & Harvesting Solutions for CHO & HEK293 Cells, for Both Transient and Stable Cells
Samuel Ellis, Vice President,
Biochemist,Thomson Instrument Company
Evaluation of different transfection tools, product quality, and titer for both CHO and HEK293 cell lines. Data will be presented on techniques and technology that mimic large-scale bioreactors in non-controlled devices from 1mL-3L. Technologies presented
include well plates and culture tube systems with incorporated filtration methodology. A new direct harvesting technique will also be introduced that eliminates centrifugation while maintaining 0.2um sterile filtration. All of these tools will be
presented with case studies from scientists.
12:40 NEW: Luncheon Presentation II: Difficult to Express Proteins: Novel Plasmid Technologies to Increase Product Yields
Sagrario Arias Rivas, PhD, Program Manager & Scientific Liason, Batavia Biosciences
Mammalian platform for high yields of difficult-to-express proteins - Precise sugar controlled microbial protein expression system - Accelerated process development.
1:10 Ice Cream Break in the Exhibit Hall with Poster Viewing (Sapphire Ballroom)
2:15 Chairperson’s Remarks
Masamichi Kamihira, PhD, Professor, Faculty of Engineering, Department of Chemical Engineering, Kyushu University
2:20 Scaling Up a High-Titer HEK293 Transient Transfection Process
Tia
Arena, MSc, Engineer I, Department of Cell Culture, Genentech
HEK293 transient expression systems are often used to quickly generate protein for research and preclinical studies. Here we describe engineering a HEK293 cell line that is more resistant to apoptosis and shear stress. After process optimization for seed
train (35 L) and transient transfections (up to 25 L), this robust cell line-enabled expression of antibodies and non-antibody proteins up to 800 mg/L in 7 days.
2:50 Recombinant Production of the Toxic Anti-Cancer Lectin Viscumin in Tobacco Plants and Microbial Cells: A Comparative Analysis of Yield, Process Costs and Toxicity
Johannes Buyel, Dr. rer. nat., Dr.-Ing., MSc, Head, Integrated
Production Platforms, Fraunhofer Institute for Molecular Biology and Applied Ecology IME
Viscumin is a potential anti-cancer protein that cannot be produced in mammalian cells due to its inherent toxicity. Manufacturing in microbial systems is cumbersome due to the formation of inclusion bodies that require a complex process, which provides
only low recoveries. Instead, plants can be used as a cost-effective alternative expression system that simplifies production and yields a more active product.
3:20 Production of Respiratory Syncytial Virus-F Protein Using High Cell Density Mammalian Expression System
Puneet Khandelwal, PhD, Senior Scientist and Group Leader, Specialty Bioanalytics, Teva Pharmaceuticals
Respiratory Syncytial Virus (RSV) is a leading cause of infant hospitalization, and life-threatening for elderly and immunosuppressed individuals. The RSV surface fusion glycoprotein (RSV-F) is well conserved and a suitable candidate for vaccine or prophylactic
therapeutic antibodies. Here, we described optimized high cell density transient expression including vector modifications for producing recombinant RSV-F protein with native-like trimers, with high affinity to neutralizing antibodies. Further, we
optimized conditions to maintain r RSV-F in native-like form using low molarity formulation buffer.
3:35 Networking Refreshment Break (Sapphire & Aqua West Foyer)
4:00 Manipulating Glycan Profile in a Transient Expression System and Application of a High-Throughput Capillary Western Method Using Lectins for Detection
Silvino Sousa, MSc, Senior Scientist, Global Protein Sciences,
AbbVie Bioresearch Center, AbbVie
I discuss approaches for modulating N-linked glycosylation of recombinant therapeutic proteins by manipulating media, process and/or genetics of the host cell factory. What is also needed is a rapid, simple, yet protein- and titer-agnostic method for
deriving detailed glycan signature directly and simultaneously from multiple samples of cell culture conditioned medium. I review methods that we have implemented for rapidly screening for glycan signatures directly from cell culture supernatants.
4:30 Accumulative Transgene Integration into a Predetermined Chromosomal Site of CHO Cells
Masamichi Kamihira, PhD, Professor, Faculty of Engineering, Department of Chemical Engineering, Kyushu University
An accumulative site-specific gene integration system (AGIS) based on the Cre-recombinase/loxP system, using mutated loxP sites (Kameyama et al., 2010, Biotechnol. Bioeng., 105, 1106–1114) has been applied for the generation of recombinant CHO
cells for producing antibodies (Wang et al., 2016, J. Biosci. Bioeng., 124, 583–590). AGIS can provide an efficient tool for repeated integration of transgenes into a predetermined chromosomal locus.
5:00 PANEL DISCUSSION: Transient, Stable or Both?
Speed, limiting risk and protein quality are often cited as advantages of transient protein production (TPP), while stable transfection – the longer and more complex process – has the advantage of producing long-term expression of the
biotherapeutic of interest. The rapidly increasing need for recombinant proteins necessitates further improvements in both technologies.
Moderator:
Masamichi Kamihira, PhD, Professor, Faculty of Engineering, Department of Chemical Engineering, Kyushu University
Panelists:
Tia Arena, MSc, Engineer I, Department of Cell Culture, Genentech
Johannes Buyel, Dr. rer. nat., Dr.-Ing., MSc, Head, Integrated Production Platforms, Fraunhofer Institute for Molecular Biology and Applied Ecology IME
Howard R.G. Clarke, PhD, Principal Scientist, Cell Sciences, Seattle Genetics
Puneet Khandelwal, PhD, Senior Scientist and Group Leader, Specialty Bioanalytics, Teva Pharmaceuticals
Silvino Sousa, MSc, Senior Scientist, Global Protein Sciences, AbbVie Bioresearch Center, AbbVie
5:30 Close of Day
FRIDAY, JANUARY 18
8:00 am Registration (Sapphire West Foyer)
8:00 BuzZ Sessions with Continental Breakfast (Sapphire Foyer)
Protein therapeutics is a fast-growing global market. As the science improves, so does the complexity of the R&D organization. Ensuring product quality plus speed to market requires insights from stakeholders working across the stages of protein
science R&D. Join experts representing this PepTalk pipeline, peers, and colleagues for an interactive roundtable discussion. Topics include highlights from the week’s presentations, new technologies and strategies, challenges, and future
trends.
Click here for more details
Moderator: Richard Altman, MS, Scientist, Protein Technologies, Amgen
9:00 Chairperson’s Remarks
Richard Altman, MS, Scientist, Protein Technologies, Amgen
9:05 Managing a Collaborative Multidisciplinary Laboratory: Challenges, Strategies, and Benefits
Challise Sullivan, Life Scientist III, Advanced
Solutions Group, Leidos
The advantages of a laboratory staffed with skilled, versatile personnel and equipped with systems applicable to numerous applications can be vast. Integrating scientific and engineering expertise, cutting-edge technology, and a collaborative team
enables innovations spanning a wide range of disciplines and applications beyond those typically attained by conventional academic or industrial practices. This talk encompasses approaches to effectively manage multidisciplinary laboratory teams
along with the challenges and benefits of doing so.
9:35 High-Throughput Cloning for Biomarker Discovery and Functional Genomics
Vel Murugan, PhD, MBA, Research Scientist, Virginia G. Piper Center
for Personalized Diagnostics, The Biodesign Institute, Arizona State University
We generate and distribute expression clones around the world. DNASU is a central repository for plasmid clones and collections (DNASU.org). Currently we store and distribute over 300,000 plasmids including 75,000 human and mouse plasmids, full genome
collections, the protein expression plasmids from the Protein Structure Initiative as the PSI: Biology Material Repository (PSI : Biology-MR), and both small and large collections from individual researchers. We are also a founding member and
distributor of the ORFeome Collaboration plasmid collection. We discuss HT cloning methods that we employ for generating expression clones.
10:05 Recombinant Protein Production: Harmonizing the Process from Construct Generation through Protein Characterization
Richard Altman, MS, Scientist, Protein Technologies, Amgen
A robust, flexible protein production facility provides critical support to drug discovery efforts. We review the ongoing evolution of our protein production endeavors focusing on two critical components. The first is the strategic assembly of mammalian
expression “tools” that gives us a toolbox capable of expressing diverse and challenging candidate proteins. The second is the harmonization of the entire protein production process thereby reducing turnaround times and increasing
throughput.
10:35 Networking Coffee Break (Sapphire & Aqua West Foyer)
11:00 Making More Proteins: How to Get the Work Done and How to Avoid It
Peter Schmidt, PhD, Director, Recombinant Technologies
Research, CSL Behring
The increasing number of projects in early R&D combined with the need to characterize and evaluate new approaches and ideas result in a continuously increasing number of requests to purify and QC proteins. In order to meet these demands, it
is necessary to find a good balance between available resources and goals that can be realistically achieved.
11:30 CLOSING PANEL DISCUSSION: Protein Production Lab Challenges: Methodologies, Strategies, and the Art of Managing Multiple Projects
There are many challenges in operating protein production labs. This panel focuses on the following topics: initiating projects, basic expression and purification systems, pros and cons for each system, screening platforms, troubleshooting and
how much time should be spent on each system before moving to the next option. On top of “hands on” tips, we touch upon strategies on how to manage multiple “top priority” projects.
Moderator:
Richard Altman, MS, Scientist, Protein Technologies, Amgen
Panelists:
Vel Murugan, PhD, MBA, Research Scientist, Virginia G. Piper Center for Personalized Diagnostics, The Biodesign Institute, Arizona State University
Peter Schmidt, PhD, Director, Recombinant Technologies Research, CSL Behring
Challise Sullivan, Life Scientist III, Advanced Solutions Group, Leidos
Silvino Sousa, MSc, Senior Scientist, Global Protein Sciences, AbbVie Bioresearch Center, AbbVie
Bjørn Voldborg, MSc, Director, CHO Cell Line Development, The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark
12:30 pm Close of Conference