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Cambridge Healthtech Institute’s Second Annual
Protein Aggregates and Particles

Tools and Techniques for Effective Prediction and Analysis of Aggregates and Particles

4-5 November 2015


Aggregation not only impacts a product’s developability, but can also affect its stability, formulation, and immunogenicity. Aggregation can occur at any stage from development to manufacturing, thus the detection, characterization and monitoring of all aggregates, visible, sub-visible and submicron particles are becoming a major task in drug development. At the Protein Aggregates & Particles conference, we aim to uncover the underlying mechanism of aggregation, from which to develop tools to detect and characterize aggregates and particles, and ultimately to design novel biotherapeutics with reduced aggregation.

Final Agenda 

Day 1 | Day 2 | Speaker Biographies | Download Brochure  

Unpublished Data Icon: Unpublished Data | Case Study Icon: Case Study

 

 

Wednesday, 4 November

07:45 Registration and Morning Coffee

 

ENGINEERING PROTEIN THERAPEUTICS FOR REDUCED AGGREGATION

08:30 Chairperson’s Opening Remarks

Salvador Ventura, Ph.D., Professor, Biochemistry and Molecular Biology, Institute of Biotechnology and Biomedicine, University of Barcelona

 

KEYNOTE PRESENTATION

08:35 Understanding (and Controlling) Aggregation of Antibody-Drug Conjugates

Fred JacobsonFred Jacobson, Ph.D., Staff Scientist, Kadcyla™ Technical Development Leader, Protein Analytical Chemistry, Genentech, Inc.

 

09:20 Engineering Antibodies for Improved Developability Properties

Chris Lloyd, Ph.D., Senior Scientist, Antibody Discovery and Protein Engineering, MedImmune

09:50 Stable Human Antibody Therapeutics and Phage Display Libraries through Engineering of Variable Domains

Daniel ChristDaniel Christ, Ph.D., Associate Professor of Medicine; Head, Antibody Therapeutics, Immunology Program, Garvan Institute of Medical Research

Human antibody variable domains often display poor biophysical properties and a propensity to aggregate. We have identified aggregation hotspots in the CDR regions of antibody variable VH and VL domains, and have developed generally applicable strategies to overcome these limitations. Here we outline the application of the technology to human antibody therapeutics and antibody phage display libraries.

10:20 Innovative Technologies to Improve the Characterization of Protein Aggregates

Matthew Brown, Ph.D., Scientist, Malvern Instruments Ltd.

Understanding the process of protein aggregation is a key component of QbD approaches during biotherapeutic development or deviation resolution of legacy products. Advanced characterization technologies, now available to the biopharmaceutical industry, offer detailed insights into protein behavior to improve product stability and process knowledge and understanding.

10:50 Coffee Break in the Exhibit Hall with Poster Viewing

 

IMPACT OF AGGREGATION ON FILLING AND FORMULATION

11:30 Challenges and Considerations Associated with Aggregation of Biopharmaceuticals during Fill Finish Process Development, Transfer and Commercialisation

Francis Carroll, Development Scientist, Technical Development, Genzyme Ireland Ltd.

Fill Finish processing of biopharmaceuticals presents numerous challenges for the inhibition of aggregation. During filling operations, degradation induced by shear stress, chemical and photo exposure, manifests itself in elevated levels of HMWS. Furthermore, control of excipient state during lyophilisation is critical for maintaining a protein in its native state, which can inhibit increases in aggregation during a product’s shelf life.

Case Study Icon12:00 Formulation Development Based on Sub-Visible Particle Morphology Using Micro-Flow Imaging

Malin Persson, Ph.D., Senior Research Scientist, Biopharm Formulation Development, Novo Nordisk A/S

12:30 Sponsored Presentation (Opportunity Available)

13:00 Luncheon Presentation (Sponsorship Opportunity Available) or Lunch on Your Own

13:30 Session Break

 

MECHANISM OF PROTEIN AGGREGATION

14:00 Chairperson’s Remarks

Jennifer McManus, Ph.D., Lecturer, Department of Chemistry, National University of Ireland Maynooth

14:05 Understanding Protein Aggregation in Pharmaceutical Products

Jun LiuJun Liu, Ph.D., Senior Group Leader and Principle Scientist, Genentech, Inc.

Protein aggregates are common degradation products for therapeutic proteins. Due to the complex nature of protein aggregation, the underline mechanisms and their potential biological impacts are not always well understood. In this presentation, we will give an overview of protein aggregation phenomenon for a few pharmaceutical products. The mechanism and implication of these protein aggregates will be reviewed and discussed.

14:35 Aggregation Analysis at High and Low Protein Concentrations

Jennifer McManusJennifer McManus, Ph.D., Lecturer, Department of Chemistry, National University of Ireland Maynooth

Aggregation of proteins may occur by a number of different mechanisms, which can lead to a range of aggregate types. Using a range of analytical techniques the formation of protein aggregates by various mechanisms has been assessed at low and where possible, at moderate to high protein concentrations. The effect of sugars on protein stability will also be discussed.

Case Study and Unpublished Data Icon 15:05 Effects of Protein Aggregation on Uptake, Processing and Presentation by Dendritic Cells – A Case Study

Anja Langenkamp, Ph.D., Principal Scientist, Immunopathology, Pharmaceutical Sciences, Roche Pharmaceutical Research & Early Development, Roche Innovation Center Basel

Studies show that tolerance can be broken in transgenic mouse models by harshly stressed protein therapeutics. However, the underlying mechanisms and relevance for humans remain unclear. Thus, we studied the influence of aggregation on the uptake, presentation and activation of dendritic cells - the key regulators for adaptive immunity. First results will be presented that provide mechanistic insights into the properties of monomeric and aggregated variants of a therapeutic monoclonal antibody.

15:35 Refreshment Break in the Exhibit Hall with Poster Viewing

 

MODELING AND PREDICTION OF AGGREGATION PROPENSITY

16:15 Tuning the Aggregation Propensity of Protein Structures

Salvador VenturaSalvador Ventura, Ph.D., Professor, Biochemistry and Molecular Biology, Institute of Biotechnology and Biomedicine, University of Barcelona

This talk presents a method to overcome the current limitations of predicting aggregation by sequencing. The AGGRESCAN3D (A3D) server overcomes the limitations by taking into account the protein structure and the experimental aggregation propensity scale. The identified aggregation-prone residues can be virtually mutated to design variants with increased solubility. Additionally, the A3D server takes into account the dynamic fluctuations of protein structure in solution, which may influence aggregation propensity.

16:45 Rational Design of Protein Solubility

Michele VendruscoloMichele Vendruscolo, Ph.D., Professor, Department of Chemistry, University of Cambridge

I will discuss the extent to which the solubility and aggregation of proteins are related to the physico-chemical properties of their amino acid sequences. Based on these properties, I will present methods for the prediction of the solubility and aggregation of proteins and illustrate how these methods can be of practical interest and importance.

17:15 Determinants and Impact of Antibody Aggregation on Production and Application

Joost SchymkowitzJoost Schymkowitz, Ph.D., Professor, VIB Switch Lab, Department of Cellular and Molecular Medicine, KULeuven

As most proteins, antibodies have a propensity to aggregate that is determined by their primary sequence and aggregation acts as a bottleneck on both production and application. Accurate prediction of antibody quality is currently lacking but would be of value to help identify good antibodies. I will discuss a number of key determinants and how to employ them for this purpose.

17:45 Standards, Measurements, and Analysis for Protein Particle Characterization

Richard CavicchiRichard Cavicchi, Ph.D., Scientist, Bioprocess Measurements Group, National Institute of Standards and Technology

Concentration measurements of protein aggregates obtained on orthogonal instrument types often differ significantly. NIST is developing a protein aggregate standard that simulates aggregate properties for sizes from 1 µm to visible particles. In addition, we are using a custom microfluidic device that compares orthogonal measurements on single particles to analyze microfabricated particles of defined dimensions and protein aggregates to discern the effects of shape and porosity.

18:15 Networking Reception in the Exhibit Hall with Poster Viewing

19:15 End of Day

Day 1 | Day 2 | Speaker Biographies | Download Brochure  

 

 

Thursday, 5 November

08:00 Morning Coffee

 

METHODS FOR DETECTING, IDENTIFYING AND CHARACTERISING AGGREGATES & PARTICLES

08:30 Chairperson’s Remarks

Antonio Ribeiro, Ph.D., Professor, Pharmaceutical Technology, University of Coimbra Azinhaga de Santa Comba

Case Study Icon
Antonio Ribeiro
08:35 Nanoparticle Tracking Analysis for Studying Aggregation Profile of Particles

Antonio Ribeiro, Ph.D., Professor, Pharmaceutical Technology, University of Coimbra Azinhaga de Santa Comba

Subvisible particles do not constitute a mass fraction to be quantified by using size exclusion chromatography (SEC). Moreover, SEC requires high dilution of the sample, which itself can change the aggregation profile. Nanoparticle tracking analysis (NTA) can count and measure size individual species in undiluted particles and may be more appropriate than SEC for studying particles aggregation. Moreover, using fluorescent labeled particles, NTA may allow distinguishing individual from aggregated particles.

Case Study Icon09:05 Hydrodynamic Diameter (By DLS) and Molecular Mass Measurement (By SLS After FFF) Can Characterise Aggregation Level of A HMW Protein Not Characterisable by SE-HPLC

Peter Matthiessen, Ph.D., Senior Manager, Formulation, Fill/Finish, Baxter Innovations GmbH

The aggregation level of a HMW Protein cannot be quantified by SE-HPLC. Field flow fractionation can partially separate potential aggregates and SLS was used for quantitation of molecular mass increase. Also DLS was used to monitor hydrodynamic diameter increase by potential aggregates. Various temperature stress conditions in liquid and lyophilized form and mechanical stress by shaking or stirring, both known to induce aggregation, were investigated by DLS and SLS. The correlation of aggregation and subvisible particle levels was investigated.

Case Study and Unpublished Data Icon
Arvinte Tudor
09:35 New, Orthogonal Methods to Detect Protein Aggregation at High and Low Concentration

Tudor Arvinte, Ph.D., Chairman & CEO, Therapeomic, Inc.; Titular Professor of Biopharmaceutics, School of Pharmacy, University of Geneva

Based on case studies, different orthogonal methods will be presented that permit detection and characterisation of protein aggregates and particulate matter in liquid formulations of biopharmaceuticals at high and low a protein concentrations. A method alone cannot provide absolute information on the aggregates present in a sample. By using different methods to analyse a sample, we can obtain strong conclusions and a broad picture on the protein aggregation states and particulates present in the solutions.

10:05 Protein-Protein Interactions, in Multi Protein-Albumin or Peptide-Albumin Co-Formulations Using Recombinant Human Serum Albumin (rHSA) to Prevent Aggregation

Jens Thostrup Bukrinski, Ph.D., Senior Scientist, Biopharma R&D, Novozymes A/S

It is well-known that rHSA has the ability to prevent protein and peptide aggregation. At drug concentrations <1mg/mL coating of hydrophobic and hydrophilic surfaces of the primary packaging material and process equipment is expected to prevent depletion and surface induced aggregation. When aggregation is independent of the surfaces (>1mg/mL) the mechanism of aggregation prevention is less well understood and is likely to depend on the aggregation pathway of the drug. Some case studies suggest that hydrophobic patches on rHSA interact with hydrophobic patches on the drugs forming an rHSA-drug complex where such surface areas are shielded and hereby preventing self-association. Other case studies suggest an excluded volume effect with no rHSA-drug complex formation.

10:35 Coffee Break in the Exhibit Hall with Poster Viewing

11:15 Detection and Characterisation of Visible, Sub Visible Particles and Other Aggregates: Achievements and Challenges

Anacelia Rios QuirozAnacelia Rios Quiroz, MSc, Late Stage Pharmaceutical and Process Development, Pharmaceutical Development & Supplies, PTD Biologics Europe, (PTDE-PF), F. Hoffmann-La Roche, Ltd.

The talk will give an overview of required and commercially available counting methodologies for detection of protein aggregates and visible and sub visible particles (SbVP); species ubiquitously present in protein formulations. Focus will be SbVP as they are gaining attention regarding immunogenicity and quality attributes. Lack of a well-defined methodology for SbVP makes it important to increase our knowledge of emerging instruments’ performance. Applicability towards the assessment of a meaningful array of particle counting techniques will be discussed.

11:45 Chemical Kinetics and Microfluidic Sizing for the Analysis of the Aggregation of Therapeutic Proteins

Paolo ArosioPaolo Arosio, Ph.D., Marie Curie Postdoc Fellow, Department of Chemistry, University of Cambridge

In this presentation, we show how chemical kinetic analysis can identify the protein aggregation path-ways at the molecular level. We demonstrate the potential of this approach by analyzing the aggregation mechanisms of different IgGs and of human insulin under conditions that are relevant for downstream and formulation. Finally, we show a novel microfluidic technique that enables the sizing of polydisperse protein samples under native conditions on a second timescale.

12:15 Luncheon Presentation (Sponsorship Opportunity Available) or Lunch on Your Own

13:00 Dessert Break in the Exhibit Hall with Poster Viewing

13:30 End of Protein Aggregates & Particles

Unpublished Data Icon: Unpublished Data | Case Study Icon: Case Study

Day 1 | Day 2 | Speaker Biographies | Download Brochure