The 4th annual Protein Expression System Engineering conference examines the functioning of the cellular machinery harnessed during protein biosynthesis, and how to engineer hosts to efficiently express a protein of interest. The
intricate steps required to achieve properly folded protein will be discussed, including verification and sequence analysis of the gene, codon optimization, vector construction, selecting and optimizing a clone, and selecting a host system. In
addition, engineering host cells to sustain expression for longer time periods will be discussed, along with overcoming cellular stress response to produce and secrete functionally active recombinant proteins.
Final Agenda
SC12: Transient Protein Production in Mammalian Cells
Richard Altman, MS, Scientist, Protein Technologies, Amgen
Henry C. Chiou, PhD, Associate Director, Cell Biology, Life Science Solutions, Thermo Fisher Scientific
*Separate registration required.
THURSDAY, MAY 3
12:00 pm Registration (Commonwealth Hall)
12:35 Luncheon in the Exhibit Hall with Poster Viewing (Commonwealth Hall)
1:40 Chairperson’s Remarks
Shahram Misaghi, PhD, Senior Scientist, Early Stage Cell Culture, Genentech, Inc.
1:50 iML1515, A Computable Knowledge-Base of Escherichia coli Metabolism and its Structural Proteome
Colton Lloyd, PhD Candidate, Bioengineering, University of California, San Diego
This talk will introduce the latest genome-scale model of E. coli metabolism, iML1515, which models activity of 1,515 metabolic genes and provides a link to the 3D structure of each protein. In addition,
it will expand on how the model can be applied to extract knowledge from emerging big data types in biology and to analyze differences in protein structural characteristics within the E. coli species.
2:20 Next-Gen Methods for Optimizing Biological Systems
Eric Kelsic, PhD, Staff Scientist, Wyss Institute, Harvard University
I will present work optimizing codon usage and protein function using high-throughput synthesis and DNA sequencing. To understand the determinants of codon choice in E. coli, we generated 12,726
in situ codon mutants in the Escherichia coli essential gene infA and measured their fitness with MAGE-seq. Our results shed light on natural codon distributions and should improve
engineering of gene expression for synthetic biology applications. I will also share recent work optimizing AAV capsid variants for improved in vivo DNA delivery.
2:50 Multi-Omics Integration Accurately Predicts Cellular State in Unexplored Conditions for Escherichia Coli
Minseung Kim, PhD Candidate, Computer Science, UC Davis Genome Center, University of California, Davis
We develop semi-supervised normalization pipelines and perform experimental characterization to create Ecomics, a multi-omics compendium for Escherichia coli. We then use this resource to train a
multi-scale model that integrates four omics layers to predict genome-wide concentrations and growth dynamics. We demonstrate the predictive performance of the model for the various omics layers far exceeds various baselines. This work
provides an integrative framework of omics-driven predictive modelling that is broadly applicable to guide biological discovery.
3:20 INTERACTIVE PANEL DISCUSSION
Next-Gen Escherichia coli
Moderator: William Gillette, PhD, Principal Scientist, Deputy Director, Protein Expression Laboratory, Leidos Biomedical Research, Inc.
Panelists:
David Humphreys, PhD, Director and Head, Protein Sciences, UCB Pharma
Colton Lloyd, PhD Candidate, Bioengineering, University of California, San Diego
Minseung Kim, PhD Candidate, Computer Science, UC Davis Genome Center, University of California, Davis
3:50 - 4:20 Networking Refreshment Break (Harbor & Mezzanine Level)
Michael Jewett, PhD, Charles Deering McCormick Professor of Teaching Excellence, and Associate Professor,
Chemical and Biological Engineering, Northwestern University
Protein glycosylation is integrally involved in human biology and disease. Unfortunately, the study of glycans in native systems and the intentional manipulation of protein glycosylation patterns remain limited by the tools available
for biochemical characterization of glycosylation enzyme activities and the synthesis of defined glycoforms. To address these limitations, we here describe a novel cell-free glycoprotein synthesis technology. This makes possible
new application areas in the production of glycoprotein therapeutics.
4:50 Combining Metabolic and Process Engineering Strategies to Improve Recombinant Glycoprotein Production and Quality
Olivier Henry, PhD, Associate Professor, Chemical Engineering, École Polytechnique de Montréal
The accumulation of lactate and ammonia remains a major factor limiting the performance of fed-batch cell culture processes. These by-products have detrimental effects on production yields and can also negatively impact product quality.
By combining process and cellular engineering strategies, we demonstrate that significant concomitant reductions in lactate and ammonia accumulation can be achieved in fed-batch cultures, leading to increased product titers without
impacting product quality.
5:20 End of Day
5:20 Registration for Dinner Short Courses (Commonwealth Hall)
SC12: Transient Protein Production in Mammalian Cells
Richard Altman, MS, Scientist, Protein Technologies, Amgen
Henry C. Chiou, PhD, Associate Director, Cell Biology, Life Science Solutions, Thermo Fisher Scientific
*Separate registration required.
FRIDAY, MAY 4
8:00 am Morning Coffee (Harbor Level Lobby)
8:30 Chairperson’s Remarks
Olivier Henry, PhD, Associate Professor, Chemical Engineering, École Polytechnique de Montréal
8:35 Engineering an Expression System for the Production of Biotherapeutics Mimicking the Endogenous Counterpart
Lars Stöckl, PhD, Senior Director, R&D, Glycotope GmbH
Some biopharmaceuticals, such as bispecific constructs or complex glycoproteins, remain very challenging in recombinant production. We present data from two case studies of host cell engineering, clone and upstream perfusion
development for products that were produced in the human GlycoExpress cell line, which overcomes productivity and quality limitations compared to rodent cell lines.
9:05 Mammalian Display: Antibody Discovery, Affinity Maturation and Developability Screening in IgG Format
Michael Dyson, PhD, CTO, Antibody Engineering, IONTAS, Ltd.
Using directed integration of antibody genes by CRISPR/Cas9 and TALE nucleases, we have constructed large libraries in mammalian cells containing a single antibody gene/cell. This has permitted construction of millions of monoclonal
stable cell lines displaying IgG antibodies on their surface from which antibodies have been selected by flow cytometry for specificity, binding affinity, species cross-reactivity and expression level. Expression in production
cell lines also enables high-throughput developability screening.
9:35 Protein Expression System Engineering
Jamie Freeman, PhD, Product manager, Horizon Discovery Group
Aside from single gene knockouts to allow for metabolic selection systems, the CHO host remains largely unchanged. I will present how we have used a combination of techniques including genome engineering approaches such as
CRISPR and rAAV to improve the biomanufacturing capacity of our GS knockout CHO K1 cell line.
10:05 Networking Coffee Break (Harbor & Mezzanine Level)
10:35 An Automated Metabolic Modeling and Analysis Pipeline for Chinese Hamster Ovary Clone Selection and Process Optimization
Tobias Grosskopf, PhD, Scientist, Cell Culture Research, Roche Diagnostics GmbH
An increase in fermentation capacity and analytical capabilities has led to an ever growing amount of data to characterize antibody producing CHO cell lines. We have generated an automated pipeline to analyze all relevant process data by an integrated central model and generate a predictive score, which allows for selection of the lead clone among several candidate clones. The central model will get better over time and hence allow for the accumulation of biological and process knowledge.
11:05 Engineering of Protein Secretion Using Systems Biology Models
Nathan Lewis, PhD, Assistant Professor, Pediatrics and Bioengineering, Systems Biology & Cell Engineering,
University of California, San Diego
The complexity of biotherapeutics and their host cells pose unique challenges to drug production. To address this, we are mapping out the complex cellular pathways controlling protein synthesis and secretion in CHO cells.
Here, I demonstrate how this information provides insights into the protein-production capacity of CHO cells, and how metabolic needs differ across products. Furthermore, these resources allow us to control the production
of toxic by-products and thereby improve bioprocess phenotypes.
11:35 PKM1 Expression Appears to Drive Lactogenic Behavior in CHO Cell Lines, Triggering Lower Viability and Productivity; A Case Study
Shahram Misaghi, PhD, Senior Scientist, Early Stage Cell Culture, Genentech, Inc.
Lactogenic behavior displayed by some CHO cell lines during manufacturing may result in a decline in viability, productivity, and alterations in product quality. We identified two lactogenic cell lines expressing different
antibody molecules during the cell line development (CLD) process. These lactogenic behaviors were differentially mitigated through process development by optimizing either nutrient feeds or culture pH, depending on
the cell line. CRISPR/Cas9 mediated knockout of the PKM-1 isoform abolished lactate accumulation even under lactogenic conditions.
12:05 pm Mitochondrial-Derived Small RNAs as Powerful Tools to Boost CHO Cell Productivity
Lisa Alexandra Pieper, PhD, Postdoctoral Researcher, Early Stage Bioprocess
Development, Boehringer Ingelheim Pharma GmbH & Co. KG
In an effort to improve the performance of a manufacturing clone expressing a complex therapeutic protein, we introduced the human mitochondrial genome-encoded small RNA-1978, resulting in a vastly increased specific productivity.
Notably, by applying the respective small RNA in combination with directed modulations of the cell culture process we successfully maximized the final product titer, proving the superiority of integrated cell line engineering
and process optimization.
12:35 Luncheon Presentation (Sponsorship Opportunity Available) or Enjoy Lunch on your Own
1:05 Networking Refreshment Break (Harbor & Mezzanine Level)
1:35 Chairperson’s Remarks
Lars Stöckl, PhD, Senior Director, R&D, Glycotope GmbH
1:40 Development and Applications of Universal Platforms for Genetic Code Expansion
Abhishek Chatterjee, PhD, Assistant Professor, Chemistry, Boston College
The ability to site specifically incorporate unnatural amino acids (UAAs) into proteins in living cells has emerged as a powerful method to probe and manipulate protein structure and function. We are expanding the scope
of this technology by establishing platforms that enable facile introduction of previously inaccessible chemical functionalities into the genetic code of both bacteria and eukaryotes.
2:10 Development of DNA-Encoded Chemical Libraries at Pfizer
Anokha S. Ratnayake, PhD, Principal Scientist, Pfizer Global R&D Groton Labs
The design and development of successful DNA-encoded libraries (DELs) require implementation of reliable analytical techniques and assays for on-DNA reaction monitoring, validation of on-DNA chemistries and assuring
library quality (QC). This presentation will focus on the background on DNA-encoded library technology (DELT), elements of library design and the details of on-DNA chemistry validation, highlighting the associated
analytical development processes.
2:40 Codon and Codon Pair Optimization in Synthetic Gene Design
Dimitris Papamichail, PhD, Assistant Professor, Computer Science, The
College of New Jersey
Advances in de novo synthesis of DNA and computational gene design methods make possible the customization of genes and gene libraries by direct manipulation of features such as
codon and codon context bias. In this talk, I will present computational methods to design genes with desired codon and codon context content, and tools that allow for the direct manipulation of protein-coding
genes.
3:10 Quantity or Quality? Enhancing Co-Translational Protein Folding with Sub-“Optimal” Synonymous Codons
Patricia L. Clark, PhD, Rev. John Cardinal O’Hara C.S.C. Professor, Biochemistry,
University of Notre Dame
Historically, “optimizing” a gene for heterologous expression consisted of substituting rare codons with synonymous common codons. This strategy can increase the amount of protein produced but may adversely
affect the yield of native, active protein. This talk will focus on rare codon distribution in coding sequences and rational strategies for rare codon placement to enhance folding yield.
3:40 End of Conference