Cambridge Healthtech Institute’s Inaugural
Protein Purification Technologies

Achieving Purity & Quality

2-3 November 2015

Protein purification is the most costly and time-consuming process in the manufacture of proteins. Challenges are multiplied when purifying complex molecules, such as membrane proteins, bispecifics and antibody-drug conjugates. The “Protein Purification Technologies” meeting explores the strategies and technologies employed to streamline steps and keep up with industry’s growing demands. The agenda examines current developments and issues of scale, along with purifying challenging proteins, such as membrane proteins, and innovating ‘traditional’ technologies such as Protein A and chromatography. Purifying proteins in different expression systems will also be addressed including mammalian, bacterial and insect cells.

Final Agenda

Day 1 | Day 2 | Speaker Biographies | Download Brochure  

: Unpublished Data | : Case Study

Monday, 2 November

12:00 Conference Registration

15:50 Refreshment Break in the Exhibit Hall with Poster Viewing

PURIFYING ANTIBODIES

16:30 Chairperson’s Remarks

Dirk Linke, Ph.D., Professor, Molecular Microbiology, Biosciences, University of Oslo

16:35 Magnetically-Driven Hybrid Technologies for Antibody Purification

Ana Cecília Afonos Roque, Ph.D., Assistant Professor, Chemistry, UCIBIO, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa

Two magnetically-driven hybrid technologies have been recently proposed by our group as novel alternatives for non-chromatographic antibody purification. One approach concerns with the incorporation of magnetic particles into macroporous structures to confer a magnetic response to the new composites. This magnetic response was explored for accelerated and efficient protein elution. In a second approach, the potential to combine aqueous two-phase extraction with magnetic separation was explored with success.

17:05 Purification of Common-Light-Chain Bispecific Antibodies

Juergen Nett, Ph.D., Associate Director, High Throughput Expression, Adimab LLC

A variety of bispecific constructs benefit from the use of a single variable light region pairing with multiple distinct variable heavy regions. This talk will demonstrate new techniques to purify these common-light-chain bispecific IgG molecules to homogeneity. A panel of bispecific constructs are then generated that bind to each target with high affinity and exhibit favorable biophysical properties similar to traditional therapeutic antibodies.

17:35 Aqueous Two-Phase Systems: A New Platform for Antibody Purification
Maria Raquel Aires-Barros, Ph.D., Full Professor, Institute for Bioengineering and Biosciences, Bioengineering Department, Instituto Superior Técnico, Universidade de Lisboa
Aqueous two-phase systems (ATPS) have proven to be a valuable option for the downstream processing of monoclonal antibodies (mAbs), combining a high biocompatibility and selectivity with an easy and reliable scale up and capability of continuous operation. A continuous process based in ATPS was developed by our group, for the capture of mAbs from mammalian cell culture supernatants. Furthermore, an ATPS microfluidic platform was designed as an effective tool to accelerate bioprocess design and optimization.

18:05 Welcome Reception in the Exhibit Hall with Poster Viewing

19:05 End of Day One

Day 1 | Day 2 | Speaker Biographies | Download Brochure  

Tuesday, 3 November

07:45 Registration and Morning Coffee

PURIFYING MEMBRANE PROTEINS

08:30 Chairperson’s Remarks

Karin Felderer, Ph.D., Associate Director, Protein Production, Protein Sciences, MorphoSys AG

08:40 Bacterial Membrane Separation, Fractionation and Solubilization as Essential Steps in Membrane Protein Purification

Dirk Linke, Ph.D., Professor, Molecular Microbiology, Biosciences, University of Oslo

We strive to compare and improve different protocols for the fractionation of proteins from Gram-negative bacteria into outer membrane, cytoplasmic membrane, periplasmic, and cytosolic fractions. I will compare 5 methods for membrane fractionation, and will discuss downstream processing and protein purification. I will also show recent data on fluorescence labeling of different membrane systems, that we use to track yield and purity of our membrane samples.

09:10 GPCR Structural Biology: A Case Study with a Novel Fluorescent Fusion Partner

Saurabh Sen, Ph.D., Principal Scientist, Immune Modulation and NBE Discovery, Boehringer Ingelheim Pharmaceuticals, Inc.

Structural biology of membrane proteins and GPCRs has seen major advances over the past few years, but overcoming the challenges of functional GPCR expression and structural stabilization are still time-consuming and expensive. The talk will focus on an approach that combines a powerful cloning strategy with novel transmembrane guides and a novel fluorescent reporter which significantly increases functional receptor expression in E. coli along with other downstream processing benefits.

10:40 Coffee Break in the Exhibit Hall with Poster Viewing

AFFINITY PURIFICATION

11:20 Recombinant Production and Utilization of Affinity of Proteins

Sophia Hober, Ph.D., Professor, Protein Technology, Biotechnology, KTH Royal Institute of Technology

Strategies for optimization of protein production and purification will be presented as well as a miniaturized high throughput method for protein verification. Fusion tags for improved selectivity of protein purification processes including protein domains with dual affinity will be presented. The dual affinity design can be of advantage for protein purification as well as other applications where affinity to more than one target molecule can be of value. Moreover, a novel, affinity-based method, that renders site-specific labeling of antibodies possible will be discussed.

11:50 Application of a Novel Fragment Complementation-Based Affinity System for Improved Protein Purification in a Single Step – from Highly Soluble to Membrane Proteins and Therapeutic Binder Molecules

David O’ Connell, Ph.D., Lecturer & Director, MSc Programmes in Biotherapeutics, Biomolecular & Biomedical Research, University College Dublin

In this presentation the performance of a novel EF hand-based fragment complementation system will be highlighted for the purification of GFP, carbonic anhydrase, GPCR and DARPin binder proteins. The application of a single-step purification rationale will be compared to tandem affinity purification approaches and downstream assays will be described.

12:20 Enjoy Lunch on Your Own

14:00 Dessert Break in the Exhibit Hall with Poster Viewing

OVERCOMING PURIFICATION CHALLENGES

14:30 Chairperson’s Remarks

David O’ Connell, Ph.D., Lecturer & Director, MSc Programmes in Biotherapeutics, Biomolecular & Biomedical Research, University College Dublin

14:35 Case Studies: IDPs “Intrinsically Disordered Proteins,” Difficult to Produce, but with a Significant Role in Cellular Pathways

Mario Lebendiker, Ph.D., Head, Protein Purification Facility, Wolfson Centre for Applied Structural Biology, Institute of Chemistry, The Hebrew University of Jerusalem

Disordered domains in IDPs “Intrinsically disordered proteins” or IDRs “Intrinsically disordered regions” play a significant role in cellular pathways, with an enormous potential as medical targets. Because of their lack of stable tertiary structure and their dynamic interchanging extended and flexible conformations, these types of proteins are prone to aggregate, and constitute an extraordinary challenge to produce. We’ll briefly illustrate here our strategy to produce enough amounts of these type of targets for biophysical and biochemical studies that allows us to show the way by which such domains act.

15:05 High Yield Expression of Protein Kinases in Insect Cells

Svend Kjaer, Ph.D., Head, Protein Purification, Structural Biology Science Technology Platform, Francis Crick Institute

Baculovirus mediated infection of insect cells is a well-established technology for production of proteins with many end-point applications. In this presentation, I’ll present data on the expression and purification of a kinase target and demonstrate the importance of co-expression strategies for downstream success. Moreover, data on a generic method, which significantly improves yields by infection of insect cell at high cell densities will be presented.

15:35 How to Get the High Hanging Fruits – Tools for Generation of Difficult-to-Express Antigens

Karin Felderer, Ph.D., Associate Director, Protein Production, Protein Sciences, MorphoSys AG

A critical but often unaccounted factor in successful discovery and development of therapeutic antibodies is the availability of high quality antigens and tool proteins in sufficient amounts. The presentation will focus on the development of a novel lysozyme-based expression/purification tag system allowing us to produce difficult-to-express proteins of high quality as soluble monomers. Diverse case studies on expression of human CD19, Fc receptors and other proteins will be presented.

16:05 Native and Stabilized Membrane Proteins for Drug Discovery

Anass Jawhari, Ph.D., CSO, CALIXAR

CALIXAR has developed patented technology based on new chemistry approach to safely isolate membrane proteins (GPCR, ion channels, transporters, enzymes and viral proteins) for drug discovery (antibody development, ligand screening, SBDD and vaccine application). The presentation will illustrate some case studies including Adenosine receptor A2A that was solubilized and purified without any single mutation, truncation or fusion for antibody development purposes and crystallization.

16:35 Refreshment Break in the Exhibit Hall with Poster Viewing

INNOVATING PURIFICATION PROCESSES

17:15 Robust Design of Continuous Purification of Adenovirus Vectors by Two-Column, Simulated Moving-Bed, Size-Exclusion Chromatography

José Paulo Mota, Ph.D., Professor, Chemical and Biochemical Engineering, Animal Cell Technology, iBET & FCT-Universidade Nova de Lisboa

A two-column simulated moving-bed (2CSMB) process for continuous purification of adenoviral vectors by size-exclusion chromatography is presented and validated experimentally, and a general procedure for its robust design under parameter uncertainty is described. The pilot-scale runs yield a virus recovery of 86% and impurity clearance of 90% without any fine-tuning of the operating parameters. This yield is 60% better and the productivity is increased by 6-fold with respect to single-column batch chromatography for the same volume of resin.

17:45 Estimating Protein Phase Behavior Based on Osmotic Virial Coefficients – A Thermodynamics-Based Approach

Christian Kress, Dipl.-Ing, Scientist, Biochemical and Chemical Engineering, Laboratory of Thermodynamics, Technical University of Berlin

Currently product capturing within protein production is often based on Protein-A chromatography, limited in capacity and scalability, which leads to a demand for different workup strategies (e.g. extraction by ATPS or precipitation). In order to enable estimations on the applicability of these workup strategies, models to calculate partition coefficients or protein solubility are required. Therefore a new estimation strategy based on the second osmotic virial coefficients B22 and B23 is used.

18:15 Purification of Therapeutic Proteins under Cost Constraints

Stefan Schmidt, Ph.D., Vice President, Process Science & Production, Rentschler Biotechnologie GmbH

Biologicals and biosimilars represent the fastest growing segment in the drug pipeline. This puts a lot of pressure on economical manufacturing, primarily solving efficiency issues in downstream processing. Here I demonstrate how to solve challenges by applying disposables, using modular concepts, replacing costly affinity resins and reducing the number of steps in platform processes. The case studies with examples from various molecule classes highlight successful process design, optimization strategies and critical manufacturing parameters.

18:45 End of Protein Purification Technologies

Day 1 | Day 2 | Speaker Biographies | Download Brochure  

: Unpublished Data | : Case Study

Conference Programs

• Display of Biologics
• Engineering Antibodies
• Machine Learning - Part 2

• Antibody-Based Therapies
• Emerging Targets & Approaches
• Membrane Protein Targets

• Safety & Efficacy
• Advancing Bispecifics
• Engineering Bispecifics

• Modulating the TME
• Innovative CAR T Therapy
• Next-Gen Immunotherapies

• Optimisation & Developability
• Analytical Characterisation
• Protein Stability & Formulation

• Data Science & Engineering
• Optimising Expression
• Process Development

• Intro to Machine Learning
• Machine Learning - Part 1
• Machine Learning - Part 2

• Antibody-based Therapies
• Engineering Conjugates
• Next-Gen Immunotherapies