As the pharmaceutical and biotech industries increasingly shift attention to biologics, much more attention is being paid to the prospect of membrane-bound proteins as drug targets for antibodies and other protein scaffolds. For the large GPCR and ion
channel target classes, biologics offer improved selectivity, an alternative for targets with known function that have not been amenable to small molecule drugs and the potential for using antibodies for the targeted delivery of therapeutics. However,
for the field to advance, fundamental challenges in optimizing antigen quality and presentation, discovery methodologies, protein engineering and target identification must be resolved.
The two-part Antibodies Against Membrane Protein Targets meeting provides a forum in which discovery biologists and protein engineers can come together to discuss next-generation strategies and technologies that will allow antibody- and
alternate-scaffold-based therapeutics directed against these target families to advance into the clinic and beyond.
The second conference in the set offers an examination of state-of-the-art approaches for the expression of high quality membrane protein antigens and antibody generation, then explores selection and screening strategies that can be applied to discover
binders with functional activity against GPCR and ion channel targets.
Final Agenda
Thursday, September 27
11:50 am Conference Registration Open (Foyer)
12:20 pm Plenary Keynote Program
2:00 Refreshment Break in the Exhibit Hall with Poster Viewing (Grand Ballroom)
2:45 Welcome Remarks
Kent Simmons, Senior Conference Director, Cambridge Healthtech Institute
2:50 Chairperson’s Opening Remarks
Iain D. G. Campuzano, Principal Scientist, Discovery Attribute Sciences, Amgen
2:55 Droplet-Microfluidics for the Discovery of Antibodies Binding or Modulating Receptors on Target Cells
Christoph Merten, PhD,
Group Leader, Microfluidics, European Molecular Biology Laboratory (EMBL), Germany
Droplet microfluidics has become a powerful tool for antibody screening. After giving a general overview on the technology, I will introduce our new 2-cell platform, facilitating assays involving two different cell types. This allows screening hybridoma
cells or even primary plasma cells (from mice and humans) for the secretion of antibodies binding or modulating membrane receptors on specific target cells. Approximately 100,000 antibodies can be screened in a single experiment in less than 24 hours.
3:25 Strategies for Antibody Discovery to Integral Membrane Proteins
Robert Pejchal, PhD, Associate Director, Antibody Discovery,
Adimab LLC
Discovery of antibodies that target integral membrane proteins at the cell surface represent a unique challenge. Using yeast presentation of large synthetic human antibody libraries, Adimab has developed methods for isolation of target specific binders
utilizing whole cells selection, NGS, and high-throughput screening. Considerations for utilization of solubilized protein are also discussed.
3:55 Integrating New Visualization and Analysis Tools for Large Antibody Datasets from Single-Cell Screening
Maia Smith, MSc, Data Visualization Lead, AbCellera
High throughput discovery technologies enable the rapid generation of large panels of antibody candidates. The resulting volume of data is complex and multidimensional, making it exceedingly difficult to analyze with conventional workflows in order
to select leads. We will show how AbCellera’s pipeline integrates a dynamic, interactive visualization software to easily explore large panels of antibodies and select lead candidates with specific properties for development.
4:25 Refreshment Break in the Exhibit Hall with Poster Viewing (Grand Ballroom)
5:00 Conformational Display Selection of Functional Antibodies against GPCRs
John Burg, PhD, Head, Biochemistry, Ab Initio Biotherapeutics
G protein-coupled receptors (GPCRs) are critical signaling mediators for both endogenous ligands and therapeutics. By combining next-generation synthetic antibody libraries with novel selection techniques, we have enabled the facile discovery of functional
antibodies against GPCRs. Our selections have yielded antibodies that recognize native extracellular GPCR epitopes, with the ability to modulate receptor signaling. Our technology opens the door to routine discovery in this important new class
of biologics.
5:30 Generation of Positive Allosteric Modulators of TRPV1 Using Affimer Reagents
Darren Tomlinson, PhD, Associate Professor, University of Leeds, United
Kingdom
Affimers are scaffolding proteins that constrain variable regions for molecular recognition and can be used as alternatives to antibodies in many applications. In this talk, I will focus on the isolation of Affimer reagents against TRPV1, an ion channel
implicated in pain disorders. The reagents act as positive allosteric modulators, and we have mimicked the interaction with small molecules through in silico drug design based on a co-crystal structure.
6:00 Interrogation of Surface Ig-Bearing and Ig-Secreting Cells using Fluorescence Activated Cell Sorting
John S. Kenney, PhD, President, Antibody Solutions
Surface immunoglobulin (SIg)-bearing B cells and antibody secreting cells (ASCs) e.g., plasmablasts or plasmacytes, are two populations within the germinal center for isolating high-affinity, antigen specific antibodies. Using Fluorescence-activated
Cell Sorting (FACS), SIg cells may be identified by Ag-staining of SIg, whereas, ASCs require capture of antibody secreted by the cell prior to Ag-staining. I’ll demonstrate the utility of a FACS-based approach for isolating cells from either
population.
6:30 Dinner Short Course Registration (Foyer)
9:30 Close of Day
Friday, September 28
7:00 am Registration Open (Foyer)
7:30 Interactive Breakfast Breakout Discussion Groups
Room: Back Bay B
Grab a cup of coffee and join a breakout discussion group. These are informal, moderated discussions with brainstorming and interactive problem solving, allowing participants from diverse backgrounds to exchange ideas and experiences and develop future
collaborations around a focused topic.
Table 21: Characterization of Antibodies Against Membrane Proteins
Moderator: Joseph Rucker, PhD, Vice President, Research and Development, Integral Molecular, Inc.
- Affinity and Kinetics: Useful approaches for characterizing antibody binding
- Epitopes: Different techniques for epitope mapping; binning versus mapping; why do epitopes matter?
- Cell Function: Integrating functional assays into antibody discovery and development
Table 22: How Can we Optimize Therapeutic Antibody Discovery for Multi-Spanning Membrane Proteins (GPCRs, Ion Channels and Transporters)?
Moderator: Trevor Wilkinson, PhD, Associate Director, Antibody Discovery and Protein
Engineering, MedImmune Ltd., United Kingdom
- What are the most efficient methods to isolate antibodies that modulate function?
- What are the most efficient screening methods?
- What are the preferred formats for antigens?
- Can we establish rules for how to address specific target classes?
Table 23: Production Challenges for Membrane Proteins
Moderator: Nicola Burgess-Brown, PhD, Principal Investigator, Structural Genomics Consortium, University of Oxford, United Kingdom
- Construct design: preferred tags, proteases, how many constructs to screen?
- Challenges of expressing integral membrane proteins: which expression host?
- Problems of scale of production for structural biology: resource and cost issues
- Strategies to improve purification and stability of membrane proteins: detergents, lipids, affinity reagents, liposomes, nanodiscs
Table 24: Droplet Microfluidics for Antibody Discovery
Moderator: Christoph Merten, PhD, Group Leader, Microfluidics, European Molecular Biology Laboratory (EMBL), Germany
- What kind of screens can be done on primary plasma cells from mice and humans?
- What are the specific advantages compared to other screening formats?
- How to get access to the technology?
8:30 Chairperson’s Remarks
Christopher Murawsky, PhD, Principal Scientist, Amgen
8:35 Tag-on-Demand – Exploiting ‘Switchable’ Expression Technology for the Enrichment of High-Expressing Membrane Protein Cell
Zachary T. Britton, PhD, Scientist, Antibody Discovery
and Protein Engineering, MedImmune, LLC
Poor expression and detection of membrane protein therapeutic targets have hampered drug discovery and screening efforts. To address this issue, we have developed a “Tag-on-Demand” approach to exploit ‘switchable’ expression
of ‘tagged’ membrane proteins for detection and selection steps that can be turned off for ‘untagged’ expression of native proteins and used directly in whole-cell drug discovery efforts. Validation of this approach
using model membrane proteins will be presented.
9:05 Antibody Generation for Membrane Proteins and Complex Epitopes
Christopher Murawsky, PhD, Principal Scientist, Amgen
Many of the most therapeutically interesting targets are multi-pass or multi-subunit membrane proteins that have nominal surface area exposed that can serve as epitopes for antibody binding. The extracellular regions may interact to form complex,
non-linear epitopes that are required for activity and are the targets of function-modifying antibodies. We present case studies describing strategies to discover antibodies to these targets, focusing on antigen engineering, generation of large
and diverse antibody repertoires and high throughput screening.
9:35 Improving Antibody Discovery against Challenging Targets
Rajesh Vij, Senior Scientific Researcher, Antibody Engineering,
Genentech
The generation of large and diverse panels of monoclonal antibodies against multi-transmembrane or integral membrane proteins has proven challenging, thus hindering the discovery of rare functional clones. Focusing on a case study, we will discuss
how antigen format, immunization scheme, and downstream expression and screening strategies can be optimized to identify antibodies with novel activities.
10:05 Coffee Break in the Exhibit Hall with Poster Viewing and Poster Competition Winner Announced (Grand Ballroom)
10:45 Cell Membrane Preparation to Enable Immunization, Antigen-Specific B-Cell Sorting and Screening for Antibodies against Cell Surface Targets
Habtom Hapte, PhD, Principal Scientist, Biotherapeutics Discovery, Boehringer Ingelheim
Here we report a strategy that resulted in identification of antigen-specific B cells using membrane prep as bait and a membrane prep based Meso-Scale Discovery (MSD) screening. Single B-cell sorting using membrane prep as bait generated 118 target-1-specific
hits. This strategy will enable us to generate immune responses against targets that are difficult to express and purify using membrane prep based immunization, sort and screen.
11:15 So Long Llamas: Rapid Discovery of Synthetic GPCR-Targeted Nanobodies
Conor McMahon, PhD, Postdoctoral Fellow, Biological Chemistry
and Molecular Pharmacology, Harvard Medical School
Single-domain camelid antibody fragments (‘nanobodies’) are broadly used as tools for biological research and have therapeutic potential. However, nanobody discovery typically requires animal immunization, presenting a host of barriers
to their production. We developed a fully synthetic yeast displayed nanobody library, which circumvents animal immunization and also allows rapid isolation of conformation-specific binders. From this library, we generated a range of functional
nanobodies targeting GPCRs, including conformational stabilizers and ligands.
11:45 Late Breaking Presentation
12:15 Enjoy Lunch on Your Own
1:15 Refreshment Break in the Exhibit Hall with Poster Viewing (Grand Ballroom)
1:55 Chairperson’s Remarks
Pawel Dominik, PhD, Senior Scientist, Cancer Immunology Discovery, Pfizer
2:00 Advances in Target Expression and Display for Discovery of Biological Therapeutics
Ivan Correia, MBA, PhD, Research Fellow, Head, Global Protein
Sciences, AbbVie Bioresearch Center
Generation of specific immune response to a target protein is dependent on correct fold and conformation state of the antigen and must represent its natural state in vivo. Other important considerations include post-translational
modifications and genetic polymorphism. Progress is continuously being made, and in this presentation, we introduce new perspectives and advances. We highlight many options available and successfully applied to a large number of target proteins.
2:30 Beyond Detergents: Nanoencapsulation of Membrane Proteins for Drug Discovery
Tim Dafforn, PhD, Professor, Biotechnology, University of
Birmingham, United Kingdom
For more than 40 years, the only way to extract membrane proteins from cells was to use a detergent. However, this approach was far from successful, producing samples with low stability and activity. The recent emergence of nanoencapsulation
systems like MSP and SMALP now provide membrane protein samples with high stability. In this talk, I will summarize these approaches and new developments that have occurred in the SMALP field.
3:00 Expression and Screening of Human Integral Membrane Protein Targets
Nicola Burgess-Brown, PhD, Principal
Investigator, Structural Genomics Consortium, University of Oxford, United Kingdom
The SGC promotes research advancement through our open access policy, and in the absence of IP. Globally, we have solved more than 2000 human protein structures and 10 novel integral membrane proteins (IMPs). Although we have made a significant
contribution to structural biology and protein production for functional studies, IMPs and protein-protein complexes still remain a challenge to produce. Here, I present our established approaches for eukaryotic expression and screening
IMPs using baculovirus/insect cells and BacMam technology.
3:30 Emerging Strategies for Membrane Protein Presentation in Antibody Discovery
Pawel Dominik, PhD, Senior Scientist, Cancer Immunology
Discovery, Pfizer
Phage display enables the discovery of high affinity antibodies to a variety of antigens. While discovering antibodies has become routine for most proteins, it is still challenging for transmembrane proteins. Nanodiscs have emerged as powerful
tools to study membrane proteins in a more natural lipid environment. By combining phage display and nanodiscs, we improved our ability to discover antibodies to multi-pass transmembrane proteins. This approach expands the toolbox for
the rapid discovery of therapeutic antibodies.
4:00 Close of Conference