Cambridge Healthtech Institute’s Tenth Annual

Engineering Genes and Hosts

Exploring Strategies in Systems Engineering and Synthetic Biology

January 8-9, 2018

 

The mandate of “faster, better, less expensive” resonates with recombinant protein expression and production researchers. Thus, protein expression scientists are exploring new engineering tools including synthetic biology and systems engineering. However, many variables still must be considered during the engineering process, including verification and sequence analysis of the gene or protein of interest, codon optimization, vector construction and clone/host selection. Ultimately, as with any new system, these tools must be weighed against traditional expression and production strategies to achieve the desired quantity and quality.

Cambridge Healthtech Institute’s Tenth Annual Engineering Genes and Hosts conference continues the tradition of applying effective engineering strategies for protein expression and production research leading to functional protein products. Learn from seasoned, savvy researchers as they share their real-world experiences, applications and results.

Final Agenda

SUNDAY, JANUARY 7

4:00 - 6:00 pm Pre-Conference Registration

MONDAY, JANUARY 8

7:00 am Registration and Morning Coffee

Synthetic Biology

9:00 Welcome by Conference Organizer

Mary Ann Brown, Executive Director, Conferences, Cambridge Healthtech Institute

9:05 Chairperson’s Opening Remarks

Mark Welch, Ph.D., Vice President, Research and Development, ATUM (formerly DNA2.0)

KEYNOTE PRESENTATION

9:10 Forward Genome Engineering

Ryan_GillRyan T. Gill, Ph.D., Slade Professor, Chemical and Biological Engineering, University of Colorado

I discuss new technologies and approaches that are enabling the engineering of biological systems at scales of 100,000s of designer mutations in parallel.

FEATURED PRESENTATION

9:50 A Semi-Synthetic Organism that Stores and Retrieves Increased Genetic Information

Floyd_RomesbergFloyd Romesberg, Ph.D., Professor, Chemistry, The Scripps Research Institute

Since the last common ancestor of all life on earth, biological diversity has been encoded in a four-letter, two-base-pair genetic alphabet. Expansion of the genetic alphabet to include a fifth and sixth letter for a third, unnatural base pair not only has immediate utility for a number of applications, such as site-specific oligonucleotide labeling, but also serves as the foundation for an organism with an expanded genetic code. Toward this goal, we have examined a large number of different unnatural nucleotides bearing mainly hydrophobic nucleobase analogs that pair based on packing and hydrophobic interactions rather than H-bonding. Optimization based on extensive structure-activity relationship studies and two screens resulted in the identification of a class of unnatural base pairs that are well recognized by DNA and RNA polymerases. More recently, we have engineered E. coli to import the requisite unnatural triphosphates and shown that DNA containing the unnatural base pair is efficiently replicated, transcribed, and translated within the cell, resulting in the first semi-synthetic organism that stores and retrieves increased information.

10:20 Networking Coffee Break

10:45 Codon and Codon Context Optimization in Synthetic Gene and Gene Library Design

Dimitris_PapamichailDimitris Papamichail, Ph.D., 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. I present computational methods to design genes with desired codon and codon context content, and low-cost gene variant libraries for high-throughput experimentation.

11:15 Synthetic Biology for Natural Product Biosynthesis

Christopher_BoddyChristopher Boddy, Ph.D., Professor & Director, Biochemistry Program, University of Ottawa

Synthetic biology approaches are dramatically impacting the scientific community’s ability to heterologous express complex natural product biosynthetic pathways, revolutionizing the discovery and characterization of these pathways as well as the development the natural products they encode. In this presentation we will highlight new strategies for heterologous expression of bacterial biosynthetic pathways, focusing on the violacein biosynthetic pathway, and examine methods to harness biosynthetic pathways for overproduction of natural products, using nonulosonic acid biosynthetic pathways.

11:45 Data-Driven Approaches for Rapid Scale-Up of Bioproducts

Derek Abbott, Ph.D., Director, Analytics, Amyris, Inc.

Optimization of microbial production of any product requires repeated iterations of the design-build-test-analyze engineering cycle to achieve economic viability. The presentation covers details of the automated platforms that enable Amyris scientists to rapidly cycle through a data-driven strain improvement process. Additionally, we present our vision for how the industry can leverage these platforms to positively impact the bioeconomy by dramatically reducing the cost and time to market for a variety of molecules.

12:15 pm SELECTED POSTER PRESENTATION: A High Throughput Approach to Construct Generation and Expression Screening for Recombinant Protein Production

Christine Kugel, Scientific Researcher, Biomolecular Resources, Genentech, Inc.

12:45 Session Break

ATUM 1:00 Luncheon Presentation: A Systematic Approach to Address Development and Production Challenges for Complex Biologics

Claes_GustafssonClaes Gustafsson, Ph.D., CCO & Founder, ATUM (formerly DNA2.0)

By using design of experiment (DoE) methodologies coupled with machine learning, transient transfection has been optimized to yield milligram levels of purified proteins in just a few days. Stable cell lines can be created in weeks to generate proteins at the gram scale. Case studies showing how these methods have been applied to multiple targets including soluble secreted proteins and integral membrane proteins will be presented.

 

Genome Engineering

2:00 Chairperson’s Remarks

Bjørn Voldborg, Director, CHO Cell Line Development, The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark

2:05 CRISPR/Cas Tools for Host Cell Improvement in the Baculovirus-Insect Cell System

Donald_JarvisDonald L. Jarvis, Ph.D., Professor, Molecular Biology, University of Wyoming

For the past 30 years, my group has undertaken efforts to improve insect cell hosts used in the baculovirus-insect cell system (BICS). One of our major efforts has focused on engineering protein glycosylation pathways in the BICS to create new systems capable of producing “humanized” recombinant glycoproteins. In this presentation, we report development of novel CRISPR/Cas9 tools for site-specific genome editing in the baculovirus-insect cell system. We then describe the use of these new tools to enhance our glycoengineering efforts by targeting an endogenous Spodoptera frugiperda (Sf) glycogene, which antagonizes human-type glycan elongation.

2:35 New Strategies of CRISPR/Cas9-Based High-Throughput Functional Genomic Screening

Dongxin_ZhaoDongxin Zhao, Ph.D., Postdoctoral Fellow, Prashant Mali Lab, Department of Bioengineering, University of California, San Diego

Cancer is a complex disease of which targetable vulnerabilities are the consequence of reprogramming of genetic architecture driven by various genetic mutations. Dissection of genetic interactions in a systematic way would provide unprecedented insights in drug discovery. Applying the new powerful CRISPR/Cas9 technology, we developed a combinatorial screening platform which allows for both high-throughput mapping of synthetic lethality and quantification of cancer cell type-specific interactions in metabolic circus.

3:05 Transition to BuzZ Sessions

3:15 BuzZ Sessions with Refreshments

Join your peers and colleagues for interactive roundtable discussions.

4:30 Mammalian Display: Antibody Discovery, Affinity Maturation and Developability Screening in IgG Format

Mike_DysonMike Dyson, Ph.D., CTO, 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.

5:00 RNA Structural Determinants of Optimal Codons Revealed by MAGE-Seq

Eric_KelsicEric Kelsic, Ph.D., Staff Scientist, George Church Laboratory, Wyss Institute, Harvard Medical School

To understand the determinants of codon choice across a gene, we generated 12,726 in situ codon mutants in the Escherichia coli essential gene infA and measured their fitness with MAGE-seq. Correlating predicted 5’ RNA structure with fitness revealed that codons even far from the start of the gene are deleterious if they disrupt the native 5’ RNA conformation. Our results shed light on natural codon distributions and should improve engineering of gene expression for synthetic biology applications.

5:30 PANEL DISCUSSION: CRISPR/Cas Genome Editing for Protein Expression

CRISPR/Cas has emerged as a powerful tool for engineering the genome in diverse organisms. However, there are also financial and legal considerations in using this tool. Hear this panel of experts discuss the pros and cons of CRISPR/Cas genome editing and its role in enhancing desired protein expression.

Moderator:

Bjørn Voldborg, Director, CHO Cell Line Development, The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark


Panelists:

Mike Dyson, Ph.D., CTO, IONTAS, Ltd.

Donald L. Jarvis, Ph.D., Professor, Molecular Biology, University of Wyoming

Eric Kelsic, Ph.D., Staff Scientist, George Church Laboratory, Wyss Institute, Harvard Medical School

Nathan E. Lewis, Ph.D., Assistant Professor, Department of Pediatrics, University of California, San Diego

Dongxin Zhao, Ph.D., Postdoctoral Fellow, Prashant Mali Lab, Department of Bioengineering, University of California, San Diego

 Nanotemper Technologies 6: 00 - 7:15 Welcome Reception in the Exhibit Hall with Poster Viewing

7:15 Close of Day

TUESDAY, JANUARY 9

8:00 am Registration and Morning Coffee

Case Studies in Enhancing Expression Systems

8:30 Chairperson’s Remarks

Henry C. Chiou, Ph.D., Associate Director, Cell Biology, Life Science Solutions, Thermo Fisher Scientific

FEATURED PRESENTATION

8:35 Engineering Cell Factories for Protein Production

Bjorn_VoldborgBjørn Voldborg, Director, CHO Cell Line Development, The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark

We have engineered cell-based factories for protein production, using state-of-the-art technologies combined with high-throughput genome engineering, in silico modeling, deep -omics analysis and big data analysis. Our main focus is CHO cells, where we have generated a panel of CHO cell lines, with optimized phenotypes generating tailormade PTMs, improved bioprocess and product quality, etc.

9:05 SKIK Tag Increasing the Expression of Hard-to-Express Proteins and Its Application to Antibody Screening from Single B Cells

Teruyo_Ojima-KatoTeruyo Ojima-Kato, Ph.D., Researcher, Meijo University

A novel tag sequence SKIK can drastically increase the expression of hard-to-express proteins in Eschericha coli in vivo and in vitro protein synthesis systems without affecting its activity in most cases. We used the peptide for a novel antibody production from single B cells by RT-PCR, PCR and cell-free protein synthesis, because the amount of protein expressed can be highly improved and normalized for better evaluation of Fab synthesized.

 Horizon no tagline 9:35 Engineering a CHO K1 Toolbox for Reliable High Titer Protein Expression

Maria_PerieriaMario Pereira, Ph.D., Senior Scientist, Horizon Discovery

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.

9:50 Coffee Break in the Exhibit Hall with Poster Viewing

11:30 Establishing Protein Production without Reinventing the Wheel: ProteinData.Cloud

Peter_NollertPeter Nollert, Ph.D., Business Director, Research and Development, Bio Data Bridges

Establishing viable paths for recombinant protein sample production is typically inefficient due to the commonly applied laborious trial-and-error approach. ProteinData.Cloud is a platform to access and share otherwise invisible experimental recombinant protein production information. Researchers find engineered vector sequences, positive and negative recombinant expression trials and purification detail, apply and improve recombinant protein production strategies to their own targets to improve protein production yield and purity.
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VTU Technology GmbH  12:00 pm Unlock Pichia – Novel Strategies and Molecular Tools to Enhance Protein Expression

Iskander_DibIskandar Dib, Ph.D., Principal Scientist DSP & Analytics, VTU Technology GmbH

Pichia pastoris is an established, safe and highly competitive expression host with strong and effective secretory capacities often resulting in double-digit g/l levels of recombinant protein. Recent research has brought about new exciting technologies increasing the potential of this already powerful yeast production host. An extended molecular toolbox addresses challenges in protein folding and secretion and facilitates the expression of more complex proteins, thus maximizing yields without compromising product quality.

 


12:30 Close of Engineering Genes and Hosts Conference


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