Cambridge Healthtech Institute’s 12th Annual
Engineering Genes, Vectors, Constructs, and Clones
Exploring Tools and Developing Strategies to Expand the Protein Expression Toolbox
January 20-21, 2020
Part of the Biotherapeutic Expression & Production pipeline
The demand for high-quality biotherapeutic proteins has never been greater. Many variables still must be considered during the cell-line development process, including verification and sequence analysis of the gene or protein of interest, codon optimization,
vector construction, and clone/host selection – a time-consuming and expensive process. Additionally, protein expression scientists are now exploring new engineering tools, such as synthetic biology and systems engineering. Ultimately, these
tools must be weighed against traditional expression and production strategies to achieve the desired quantity and quality. Cambridge Healthtech Institute’s 12th Annual Engineering Genes, Vectors, Constructs, and Clones conference continues
the tradition of applying effective engineering strategies for protein expression and production research leading to functional biotherapeutic products. Learn from seasoned, savvy researchers as they share their real-world experiences, applications,
and results.
Final Agenda
Day 1 | Day 2 | Download Brochure
SUNDAY, JANUARY 19
4:00 - 6:00 pm Pre-Conference Registration (Sapphire West Foyer)
MONDAY, JANUARY 20
7:00 am Registration (Sapphire West Foyer) and Morning Coffee (Sapphire West & Aqua West Foyer)
9:00 Organizer’s Welcome Remarks
Mary Ann Brown, Executive Director, Conferences & Team Lead, PepTalk, Cambridge Healthtech Institute
9:05 Chairperson’s Opening Remarks
Harun Rashid, PhD, Senior Principal Scientist, Molecular Technology, Ambrx, Inc.
KEYNOTE PRESENTATION
9:10 High-Throughput Expression of Functional Proteins in a Microarray Format
Josh LaBaer, MD, PhD, Executive Director, Biodesign Institute, Arizona State University
The Nucleic Acid Programmable Protein Array uses printed cDNAs as templates to produce full-length proteins in situ, enabling high-throughput biochemical testing of thousands of well-folded proteins simultaneously. The
recombination-based vector system allows users to routinely execute the automated transfer of thousands of genes into useful protein expression vectors overnight. These methods are now extended to include decorating the proteins with post-translational
modifications, including glycosylation, acetylation, citrullination and others.
FEATURED PRESENTATION
9:50 Codon and Codon-Pair Usage Tables (CoCoPUTs): Facilitating Genetic Variation Analyses and Recombinant Gene Design
Chava Kimchi-Sarfaty, PhD, Deputy Associate Director for Research, Office of Tissues and Advanced Therapies, CBER, FDA
We have created for codon and codon-pair usage tables (CoCoPUTs), genome and tissue specific, a new and regularly updated website that encompasses the previously generated High-Performance Integrated Virtual Environment codon usage tables (HIVE-CUTs)
that has also been expanded to include codon-pair usage and dinucleotide statistics. The use of this tool will be discussed specifically for therapeutics’ design.
10:20 Networking Coffee Break (Sapphire West & Aqua West Foyer)
10:45 Cell-Free Protein Synthesis from Genomically Recoded Bacteria Enables Multisite Incorporation of Non-canonical Amino Acids
Antje Krüger, PhD, Postdoctoral Fellow, Michael Jewett Laboratory, Department of Chemical and Biological Engineering, Northwestern University
Cell-free protein synthesis has emerged as a powerful approach for expanding the range of genetically encoded chemistry into proteins. We recently established a bacterial cell-free protein synthesis platform based on genomically recoded Escherichia coli lacking release factor 1 that enables both high-yield protein synthesis and incorporation of multiple, identical non-canonical amino acids. In this talk, we will discuss the development of this platform and its use in engineering the translation
machinery.
11:15 Expression Optimization of an Antibody Fab Fragment in Escherichia coli with Non-Native Amino Acid (NNAA) Incorporated
Harun Rashid, PhD, Senior Principal Scientist, Molecular Technology, Ambrx, Inc.
In this study, expression of a “difficult-to-express” antibody Fab fragment with a non-native amino acid (NNAA) inserted was systematically optimized by expression vector engineering. After the various genetic elements on expression vector
were tested individually, the beneficial ones were then combined into a single expression vector, which resulted in significant improvement of Fab titer over the starting strain.
11:45 Optimizing Gene Sequences for Improved Protein Expression in Industrial Microorganisms
Tomoshi Kameda, PhD, Senior Research Scientist, National Institute of Advanced Industrial Science and Technology
Codon optimization by synonymous substitution is widely used for recombinant protein expression. Recent studies have investigated sequence features for codon optimization based on large-scale expression analyses. However, these studies have been limited
to common host organisms such as Escherichia coli. Here, we develop a codon optimization method for Rhodococcus erythropolis, a gram-positive GC-rich actinobacterium attracting attention as an alternative host organism. We optimize
the coding sequences of 12 genes and confirm that 9 of them (75%) achieve increased expression levels compared with wild-type sequences. And, we also confirm our method is effective in other bacteria for industrial use.
12:15 pmPANEL DISCUSSION: Exploring Expression Tools
Moderator: Harun Rashid, PhD, Senior Principal Scientist, Molecular Technology, Ambrx, Inc.
Panelists: Speakers of the Session
12:45 Session Break
12:55 LUNCHEON PRESENTATION I: Machine Learning for Predictive Antibody Design and Humanization
Claes Gustafsson, PhD, Co-Founder, CEO, ATUM
Engineering of better antibodies, improved cell lines and higher production yields requires efficient tools to navigate biological high dimensional sequence-function space. We describe
how traditional humanization approaches that incorporate homology modeling and CDR grafting can be drastically improved by applying DoE and machine learning methodologies to generate a small number of humanized molecules with improved develop-ability
profiles. The results from humanization experiments from 12 different antibodies impart predictive design strategies for future antibody humanizations.
1:25 Luncheon Presentation II (Sponsorship Opportunity Available)
2:00 Chairperson’s Remarks
Jeffrey Barrick, PhD, Associate Professor, Molecular Biosciences, The University of Texas at Austin
2:05 Transposon-Encoded CRISPR–Cas Systems Direct RNA-Guided DNA Integration
Samuel Sternberg, PhD, Assistant Professor, Department of Biochemistry and Molecular Biophysics, Columbia University
Conventional CRISPR–Cas systems maintain genomic integrity by leveraging guide RNAs for the nuclease-dependent degradation of mobile genetic elements. We uncovered a remarkable inversion of this paradigm, in which bacterial transposons co-opt
nuclease-deficient CRISPR–Cas systems to catalyze RNA-guided integration of mobile genetic elements into the genome. This discovery of a fully programmable, RNA-guided integrase lays the foundation for genomic manipulations that obviate
the requirements for double-strand breaks and homology-directed repair.
2:35 Defending Engineered Bacteria against Evolutionary Failure
Jeffrey Barrick, PhD, Associate Professor, Molecular Biosciences, The University of Texas at Austin
Mutations in engineered DNA sequences can occur rapidly and infiltrate processes with ‘broken’ cells. Selfish genetic parasites–such as transposons and prophages–are a major source of inactivating mutations in bacterial genomes.
In this talk, I will describe the broad-host-range CRISPRi-ME system that can be added to a bacterial cell to repress these mobile elements and other strategies for creating chassis cells with lower-than-natural mutation rates to stabilize their
functions.
3:05 Find Your Table and Meet Your BuzZ Session Moderator
3:15 BuzZ Sessions with Refreshments
Join your peers and colleagues for interactive roundtable discussions.
Click here for more details
4:30 Generation of Improved Host Cell Lines for Biomanufacturing Using Vector and Cell Line Engineering Technologies
Dennis Pfaff, PhD, Investigator III, CLSD, NIBR Biologics Center, Novartis Pharma AG
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 to further reduce
timelines during cell line development.
5:00 Sequential Mechanism-Derived Heterogeneity in a Therapeutic Monoclonal Antibody
Matthew Schenauer, PhD, Senior Research Scientist II, Biologics Analytical Operations, Gilead Sciences
The observation, identification, and mechanistic basis for a unique product-related variant in a purified monoclonal antibody (mAb) therapeutic is described. It will be demonstrated that unlike typical mAb heterogeneity, the variant arose by via distinct
and sequential biochemical and molecular biological processes.
5:30 The Cellular Impact of Glycoengineering
Nathan Lewis, PhD, Associate Professor, Department of Pediatrics, University of California, San Diego
Many therapeutic proteins are glycosylated, and the glycans often significantly impact drug safety and efficacy. Thus, there has been an increased interest in engineering the host cell to provide tailored glycosylation. Here I will present our work
wherein we have deeply studied the impact of cell engineering on >180 clones, and how the engineering of glycosylation impacts cell phenotypes, their transcriptomes, and glycan patterns in expected and unexpected ways.
6:00 - 7:15 Welcome Reception in the Exhibit Hall with Poster Viewing (Sapphire Ballroom)
7:15 Close of Day
Day 1 | Day 2 | Download Brochure
TUESDAY, JANUARY 21
8:15 am Registration (Sapphire West Foyer) and Morning Coffee (Sapphire West & Aqua West Foyer)
8:45 Chairperson’s Remarks
Nathan Lewis, PhD, Associate Professor, Department of Pediatrics, University of California, San Diego
8:50 Modeling Chinese Hamster Ovary Cell Metabolism: A Systematic Look at Model Parameters and Risk of Overfitting
Matthew Schinn, PhD, Postdoctoral Researcher, Department of Pediatrics, University of California, San Diego
Metabolic network models provide mechanistic understandings of cell metabolism, and therefore could guide the rational design of cell lines and culture processes. However, such models are liable to overfit due to their high degrees of freedom. Here
we systematically evaluate a wide range of model parameters important to describing CHO fedbatch culture performance.
9:20 Model-Driven Process Development for Enhanced Bioprocessing
Meiyappan Lakshmanan, PhD, Research Scientist & Group Leader, Systems Biology, Bioprocessing Technology Institute, A*STAR
Chinese hamster ovary (CHO) cells are the preferred choice for biotherapeutic protein production. However, ensuring consistent high product quality remains a major challenge. The availability of the CHO genome sequence has enabled the development
of genome-scale models (GEMs) to examine the metabolic signatures of CHO cells upon varying bioprocess conditions. This talk will show how the genome-scale models can help process development by characterizing key bottlenecks in media formulations
and propose targets for media/feed optimization.
9:50 Coffee Break in the Exhibit Hall with Poster Viewing (Sapphire Ballroom)
11:00 Democratizing Cell-Free Protein Synthesis: Improving Access for Broad Bioengineering Applications
Javin Oza, PhD, Assistant Professor, Chemistry & Biochemistry, California Polytechnic State University, SLO
Cell-free protein synthesis (CFPS) is a platform technology that leverages cell extracts for the on-demand production of proteins. CFPS supports discovery by obviating the barriers presented by the cell, enabling scientists and engineers to rapidly
characterize genes, constructs, and clones associated with functional genomics, ‘difficult’ proteins, and metabolic engineering. I will discuss our efforts to reduce the barriers to implementing CFPS to enable broad applications in
bioengineering.
11:30 Using Human Blood Extracts as a Renewable Resource to Produce Recombinant Proteins
David Burgenson, PhD Candidate, Center for Advanced Sensor Technology (CAST), University of Maryland Baltimore County
One of the early in vitro translation cell-free protein expression systems used rabbit reticulocytes (immature red blood cells) as a source of cells to produce active cell-free extract. Using this system as a source of
inspiration, our group began looking for cells present in human blood that could be used to produce translationally active cell-free extract.
12:00 pm Engineering of a Viral Free SURE Host Cell Line for Biologic Production
Pierre-Olivier Duroy, PhD, Genome Editing Director, SELEXIS SA
CHO cells have budding type-C endogenous retroviruses (ERVs) embedded in their genome and release viral particles in the culture supernatant. The presence of ERVs has raised safety concerns. We systematically characterized the type-C ERV elements at the genome, transcriptome and viral particle level. Using genome editing methods, we disrupted the expressed ERVs without compromising cell growth, size or recombinant protein production. We will present a novel strategy to mitigate CHO endogenous retroviruses during biopharmaceutical manufacturing.
12:30 Session Break
12:40 Luncheon Presentation: GlycoExpress® - An Alternative Host for Difficult to Express Proteins
Lars Stockl, PhD, Senior Director, R&D, Glycotope GmbH
The era of classical IgG molecules in bio-pharma development is shifting rapidly to more challenging complex biopharmaceuticals. With CHO being a good production cell line for IgG molecules, they might fail to produce more challenging candidates.
The GlycoExpress® (GEX®) system represents an ideal alternative for the production these difficult to express protein molecules and will provide case studies which demonstrate the superiority in productivity and product quality vs CHO
cell expression.
1:10 Close of Engineering Genes, Vectors, Constructs, and Clones Conference
5:45 - 8:45 Recommended Dinner Short Course*
SC6: Assembling an Effective Toolbox of Expression Systems to Support Your Drug Discovery Efforts - Detailed Agenda
Instructors:
Richard Altman, Field Application Scientist, Protein Expression, Biosciences Division, Life Sciences Solutions Group, Thermo Fisher Scientific
Henry Chiou, PhD, Director, Cell Biology, Life Science Solutions, Thermo Fisher Scientific
Dominic Esposito, PhD, Director, Protein Expression Laboratory, Frederick National Laboratory for Cancer Research
*Separate registration required
Day 1 | Day 2 | Download Brochure