Day 1 :
Keynote Forum
Thea Sesardic
NIBSC, UK
Keynote: Neutralizing antibodies directed against Botulinum A and B toxin heavy and light chains
Time : 9:30-10:05
Biography:
Thea Sesardic has over 35 years’ experience in biomedical research with 25 years’ experience at NIBSC on regulatory research focusing on the 3Rs - replacement, reduction and refinement of animals in the field of biological products derived from bacterial toxins. Over 10 methods have been developed and implemented as part of the batch release function of the group for toxins, antitoxins and toxoid vaccines at NIBSC, reducing the use of animals by 1,000 a year at NIBSC alone. Several of the methods have also gone through international validation and are now regulatory standards. In her capacity as UK delegate on the European Pharmacopoeia group of experts, she has introduced new monographs and method chapters (eg. potency of human tetanus IgG, safety and potency of DT vaccines, replacing toxin challenge methods and replacing LD50 assays for Botulinum toxin for therapy) that continue to impact animal use in Europe and globally. She has contributed to several major revisions and manuals of methods to assess quality, safety and efficacy of DT vaccines as part of a WHO working group.
Abstract:
Purpose of Study: The antiBotABE is a collaborative consortium project with an aim to generate an oligoclonal cocktail of protective recombinant super-humanized IgGs against neurotoxins secreted by Clostridium botulinum A, B, and E strains. A new strategy has been implemented involving germline-humanized antibodies of macaque (Macaca fascicularis) origin, which already proved successful to generate neutralizing antibodies against ricin and anthrax toxins, as well against encephalitis viruses. These drug lead candidates could become available for biodefense primarily, but also for treatment of natural and iatrogenic cases of botulism. Methods: The recombinant form of the corresponding heavy (HC) and light (LC) chains of botulinum neurotoxin (BoNT) A and B were used to immunize macaques, from which an immune antibody phage-display library was built. The best antibody fragments (scFvs) isolated from that library were selected according to their affinities, and the best binders against the LC and HC were tested against BoNT holotoxin for their in vitro inhibition properties in an endopeptidase assays and ex vivo neutralization in mouse phrenic nerve-hemidiaphragm assay respectively. The most effective scFvs were humanized using human germline sequences as template, expressed as full size IgG and tested individually and in combination for neutralization properties in local flaccid paralysis as well as in lethality bioassay in vivo. Summary of Results: The best scFvs or scFv-Fcs against the LC, selected by inhibition of endopeptidase studies in vitro and subsequently by mouse phrenic nerve-hemidiaphragm assay were SEM120-IIIC1 and BLC3 against BoNT/A and B respectively. The best scFvs or scFv-Fcs against the HC were selected by highest neutralization capacity in the mouse phrenic nerve-hemidiaphragm assay and were AHc38 and B2-7 against BoNT/A and B respectively. When these antibodies were expressed as IgGs and tested in the mouse protection assay AHc38 and BLC3 alone protected mice from toxin induced paralysis, whereas combinations of SEM120-IIIC1 and AHc38 as well as combinations of BLC3 and B2-7 fully protected mice in vivo at doses where protection was not observed by each antibody alone. Conclusions: AntiBotABE project has developed promising germline-humanized IgGs confirming the success of a strategy, based on targeting the HC and LC domains and the use of NHP hyper-immune libraries. These antibodies are potential lead candidates for further clinical development.
- Track 5: Antibodies: Emerging, Reemerging & Infectious Diseases Track 6: Monoclonal Antibodies & Organ Cancers Track 8: Antibodies: Biology & Engineering Track 12: Biosimilars & Current Research Track 15: Antibodies: Medical Applications Track 22: Lymphocytes: Research & novel Strategies
Chair
William G Whitford
GE Health Care, USA
Co-Chair
Michael R Dyson
IONTAS Ltd, UK
Session Introduction
Michael R Dyson
IONTAS Ltd, UK
Title: Construction and Usage of Large Antibody Libraries in Mammalian Cells
Time : 10.05-10.30
Biography:
Mike is a co-founder of IONTAS Ltd, a therapeutic antibody discovery company. He constructed the IONTAS antibody phage display library, is a co-inventor of the IONTAS mammalian cell antibody and T cell receptor display platform and acts as the project leader for several therapeutic antibody discovery projects. Mike was previously Head of Protein Engineering at Acambis plc (acquired by Sanofi) and was the Project Leader, responsible for protein expression, within the Atlas Project at the Wellcome Trust Sanger Institute. He has over 20 years experience in cell and molecular biology gained during research at the Massachusetts Institute of Technology, Universities of Edinburgh and Cambridge and the biotechnology industry. He has published 30 papers and book chapters (cited over 1000 times, h-index 17) and is inventor on 5 US patent and patent applications.
Abstract:
The availability of large libraries of binders expressed within mammalian cells will allow direct screening for function through simultaneous expression and functional reporting within the same cell. In addition, libraries of binders in mammalian cells could also be used for display of binders on the cell surface allowing the screening of millions of clones by flow sorting while providing information on both the level of expression and the extent of binding within individual clones. The main limitation in achieving this has been the inability to construct sufficiently large libraries containing a single antibody gene/cell. We have solved this problem by directing the integration of antibody genes into a single genomic locus through the use of site-specific nucleases. Antibody libraries consisting of many millions of clones have been constructed and binders selected. This includes antibodies formatted as IgGs expressed in the cell type used for production.
Jan Voskuil
Everest Biotech, UK
Title: How antibodies can prevent medical progress and how they can be great tools?
Time : 10:30-10:55
Biography:
Jan Voskuil is the Chief Scientific Officer at antibody manufacturer Everest Biotech in Oxfordshire, UK. After specializing in prokaryotic cell biology through his PhD program in Amsterdam, Netherlands, and a Postdoc position at Stanford, California, he switched to the science of neurodegenerative diseases at Oxford, UK through Postdoc positions at Dunn School of Pathology and at MRC and through a leading position at the Alzheimer drug discovery company Synaptica. He subsequently gained experience in a GLP-regulatory environment in CRO companies both in Oxfordshire and Cambridgeshire, validating assays in Flow Cytometry and ELISA platforms and writing SOPs. His extensive experience with generating and characterizing monoclonal and polyclonal antibodies in combination with accrued knowledge on most immunoassays in academic and commercial environments made him the ideal candidate to take charge in putting Everest Biotech on the global map by ever raising the quality and size of its catalogue and by delivery of adequate technical support. As a result, Everest antibodies are currently part of most globally well-known catalogues, and its products are increasingly recognized as useful alternatives to unfit monoclonal antibodies.
Abstract:
Antibodies play a pivotal role in diagnostics, biomarkers research and increasingly in therapeutics. However in all three fields problems related to specificity and to consistency of the antibodies occur. Such problems are being kept to a minimum by the use of monoclonal antibodies, because of their immortality and for their single molecular basis. However, existing monoclonal antibodies have generally not been screened for the specific application it is currently required for. This leads to the use of antibodies with insufficient affinity for that particular application. In addition different formulations at which one antibody is offered to the market will cause inconsistencies in performance. Any polyclonal antibody is dismissed because it represents a mixture of different molecules, and therefore this mixture will change in composition from animal to animal thus changing the characteristics of the reagent from batch to batch. Although for therapeutics the best way to address these problems may be through the route of recombinant reagents (avoiding the animals altogether), one remains to be vigilant for cross-reactivity of reagents when reacting to a non-unique epitope. Here is a good opportunity to make the most of a cost-effective alternative for in vitro and for ex vivo tests; there is evidence that epitope-specific polyclonal antibodies offer an adequate alternative to unavailable fit-for-purpose monoclonal antibody.
Hiroshi Ueda
Tokyo Institute of Technology, Japan
Title: Quenchbodies - Quench-based antibody probes that emit antigen-dependent fluorescence
Time : 11:10-11:35
Biography:
Hiroshi Ueda has completed his PhD in the University of Tokyo, and studied antibody engineering in Laboratory of Molecular Biology, Medical Research Council, Cambridge, UK. He has been working with antibodies with the aim of developing novel methods for biosensing and diagnostics for years, and received many awards including Biosensors 2012 Best Paper Award. He is now the Vice chairman of Biochemical Assay Society of Japan, and a Councilor of The Japanese Biochemical Society.
Abstract:
In the areas of biological analysis and clinical diagnostics, rapid and specific quantification for various molecules is considered highly important. Here we describe a novel reagent less fluorescent biosensor strategy based on the antigen-dependent removal of a quenching effect on fluorophores attached to an antibody. Using position-specific protein labeling systems, we found that an antibody single chain variable region (scFv) fluoro labeled at the N-terminal region showed a significant antigen-dependent fluorescence increase. Investigation of the enhancement mechanism by mutagenesis, denaturation, and fluorescence lifetime measurement of the carboxytetramethylrhodamine (TAMRA)-labeled anti-osteocalcin scFv showed that antigen-dependency was dependent on semi-conserved tryptophan residues. This suggested that the binding of the antigen led to the removal of quenching effect caused by the proximity of tryptophan residues to the linker-tagged fluorophore. Using TAMRA-scFv, many targets including peptides, proteins, and haptens including narcotics could be quantified. Similar or higher sensitivities to those observed in competitive ELISA were obtained, even in human plasma. Furthermore, we found that a number of Fab fragments fluorescence-labeled at their N-termini with various dyes showed higher fluorescence enhancements probably due to enhanced quenching. Due to its versatility, these “quenchbodies” are expected to have a range of applications, from in vitro diagnostics to imaging of various targets in situ. Latest results for application to cellular imaging will be shown.
Hazel Gorham
PRA HealthSciences, UK
Title: Challenges in demonstrating biosimilarity and interchangeability of biosimilar products
Time : 11.35:12.00
Biography:
Hazel Gorham, PhD, Director, Biosimilars Development, Scientific Affairs, has 26 years of experience in both academic and pharmaceutical research/industry across a wide range of roles including developing Clinical Development Plans for biosimilars. Dr. Gorham has experience in all aspects of biosimilar development including study design and regulatory agency discussions. She has experience in all aspects of clinical trial project management and has managed the conduct of multiple Phase I through IV clinical trials and post-marketing studies across geographical regions and differing therapeutic areas. Dr Gorham completed her post-doctoral graduate studies in microbiology at the University of Warwick, UK.
Abstract:
The market of biologics is growing at nearly twice the rate of pharma as a whole. The expiration of patents and other intellectual property rights for originator biologicals over the next decade opens up opportunities for biosimilars to enter the market and increase industry competition. In order to be cost effective a biosimilar product needs access to global markets based on a single development programme that meets the requirement of regulators internationally. Despite increasing alignment in the regulatory requirements for biosimilars between EMA, FDA, WHO and other jurisdiction, there are still many scientific and practical challenges for demonstrating biosimilarity and interchangeability including scientific factors, drug interchangeability and statistical considerations.
Trairak Pisitkun
Chulalongkorn University Systems Biology (CUSB), Thailand
Title: Integrated design of antibodies for systems biology using AbDesigner
Time : 12:00-12:25
Biography:
Pisitkun Trairak is currently working as Principal Investigator, Chulalongkorn University Systems Biology (CUSB) Center, Faculty of Medicine, Chulalongkorn University, Bangkok, Thailand. He has successfully completed his Administrative responsibilities as Senior Scientist, InterPrET Research Center, Department of Biomedicine, Aarhus University, Denmark. His research has included Systems Biology, Computer Software and Web Application Development. Based on this research and fellowship training, he has received several awards and honors, such as Outstanding Mentorship Award, National Heart, Lung, and Blood Institute, National Institutes of Health.
Abstract:
In the current era of large-scale biology, systems biology has evolved as a powerful approach to identify complex interactions within biological systems. In addition to high throughput identification and quantification techniques, methods based on high-quality mono-specific antibodies remain an essential element of the approach. To assist the large-scale design and production of peptide-directed antibodies for systems biology studies, we developed a fully integrated online application, AbDesigner to help researchers select optimal peptide immunogens for antibody generation against relatively disordered regions of target proteins. Here we describe AbDesigner in terms of its features, comparing it to other software tools, and the applications of it to design various antibodies.
Saroj Velamakanni
Fahy Gurteen Laboratories, UK
Title: Multiparameter analysis using cell cycle biomarkers for breast cancer: Prognostic and predictive implications
Time : 13:10-13:35
Biography:
Saroj Velamakanni earned his PhD degree in Molecular Microbiology at University of Cambridge, UK. In 2002, he was awarded the prestigious Cambridge Nehru Scholarship to pursue his Doctoral studies. After Post-doctoral work at the Universities of Cambridge, he received a research fellowship from Wolfson College, Cambridge to work on Calcium Signalling in mammalian systems. During this time his work was published in prestigious journal Nature. In 2012, he became associated with the start-up Biotech/in vitro diagnostics company Fahy Gurteen Labs based in Cambridge Research Park first as a Senior R&D Scientist and currently as a Research Director. He has 22 peer reviewed high impact publications and six patents to his credit.
Abstract:
Cell cycle related molecules play a pivotal role in maintaining genomic stability and can be used as biomarkers for cell cycle phase distribution. In this study, we performed immunohistochemical multiparameter analysis using five cell cycle related biomarkers for a cohort of 182 patients and linked it with clinicopathological information. Various stages involved in the design, development and validation of proprietary antibodies against these biomarkers will be discussed. Three unique cell cycle phenotypes were identified 1) out-of-cycle state 2) G1 delayed/arrested state and 3) accelerated cell cycle progression state. Our algorithm sheds new light into the cell cycle state of dynamic tumour populations in vivo. The information obtained is of major prognostic significance and may impact on individualised therapeutic decisions.
Reem Hamdy A Mohammed
Cairo University Hospitals, Egypt
Title: B cell targeted strategies
Time : 13:35-14:00
Biography:
Reem Hamdy A Mohammed is a Assistant Professor of Rheumatology, graduated from the School of Medicine, Cairo University, MB, BSCh, 1995 excellent grades with honors. She finished her Master degree of Rheumatology and Rehabilitation in May 2000, and MD Rheumatology and Rehabilitation, PhD rheumatology, at Cairo University in 2004. She is recognized as international fellow by the American college of Rheumatology in 2011, nominated as Fellow by the Royal College of Physicians UK in May, 2014 and member of the American Association of Immunologists. As a rheumatologist, she is committed to disseminating evidence based knowledge in the field of rheumatology and autoimmune diseases to all her students, advancing research and academic standards in my organization. In the field of research, she shared in multiple researches in the field of autoimmune diseases many of which has been published in addition to sharing as an author in two book publications about vasculitis. She is also serving as an editor and reviewer in a number of Journals in the field of Rheumatology and Clinical Immunology.
Abstract:
With the evolution of biologic drugs in the last one and half decades, the treatment paradigm of autoimmune diseases experienced a dramatic shift that ultimately improved outcome in the setting of refractory illness. Targeting the antibody producing B lymphocyte at its different developmental stages represents one important approach nowadays in the treatment strategy of a multiplicity of autoimmune diseases especially rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). The production and differentiation of the B cells passes through various developmental stages with the earliest identifiable precursor being the stem cells also known as the progenitor B-Cell, then pro-B, pre-B, immature B, virgin B-Cell and plasma cells. Therapeutic antibodies to cell surface receptors on B lymphocytes as well as their survival factors have been extensively tested with the anti-CD-20 therapy Rituximab being on top of the list used for refractory rheumatoid and in refractory systemic lupus particularly after failure of conventional immunosuppressive drugs, in addition to anti B lymphocyte survival factor anti- BLys like Atacicept and Belimumab, the recently approved biologic therapy for systemic lupus erythematosus. Other uprising B cell targeted strategies include Blisimod (A623) soluble and membrane bound BLyS, and Tabalumab (LY2127399) soluble and membrane BLyS, such novel drugs are being currently evaluated in SLE and RA
Lijuan Zhang
National Institute for Communicable Disease Control and Prevention, China
Title: Agrarian residents in china are at increased risk of tick-borne anaplasmosis and ehrlichiosis and spotted fever exposure
Time : 14:00-14:25
Biography:
Dr Zhang-lijuan (MD, phD ) is the Director of the Dept. of Rickettisology, National Institute for Communicable Disease Control and Prevention, China CDC. She is and PhD Candidate Supervisor and one of the members of the Committee of Natural Focus Disease, Ministry of Hygiene of People’s Republic of China, and a member of the National Zoonoses Committee. Her research interests are national surveillance of Rickettsioses, Anaplasmosis and Ehrlichiosis and molecular epidemiological and pathogenesis researches of rickettsiae, anaplasma and ehrlichia .She has published more than 60 papers on the surveillance of rickettsiae and basic research on rickettsiae agents.
Abstract:
More and more epidemiological evidences indicated that the annual numbers of the emerging anapalsema and ehrlichia and the reemerging rickettsiae have steadily increased in China and the outbreaks of these zoonoses have been frequently reported in recent years. Of the traditional rickettsioses, scrub typhus, caused by Orientia tsutsugamushi (Ot), is one of the serious acute febrile illness in China. In some severe endemic areas of southern China, hundreds of farmers suffered scrub typhus in small villages each year. Similarly, murine typhus, caused by R. typhi is highly prevalent in some rural areas although the louse borne epidemiological typhus has been controlled in China. Noticeably, a big outbreak of murine typhus involved 76 patients occurred in drug detoxification program from Ruili city, Yunnan province in 2010. Besides R. sibirica and R. heilongjiangensis, emerging spotted fever group rickettsiae (SFGR) including R. raoulti, R. felis, R. massiliae and SFGR Hainan-1 were successively detected in China in recent years. Specifically, a highly seroprevalence of the SFGR Hainan-1 were identified in 46.1% of the adult population and 37.5% of the preschool children in the local areas. An active national investigation indicated that the emerging tick born E. chaffeensis and A. phagocytophilum were highly prevalent among agrarian residents, domestic animals and some wild animals. In particular, an unusual transmission of nosocomial cases of human granulocytic anaplasmosis occurred in Anhui Province in 2006. In China, the rickettsial diseases are the greatest challenge to public health. Notably, rural residents are not only at increased risk of these zoonoses but also these diseases are mostly under-recognized.