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PROGRAM CHAIRS:
Otto-Wilhelm Merten, PhD - Généthon
Barbara A. Thorne, PhD - Thorne Bio-consulting LLC

Joseph G. Joyce, PhD
Director, Vaccine Process Development, Merck Research Laboratories
Challenges in the Development and Scale-Up of an
Attenuated Live Virus Vaccine Candidate for Human Cytomegalovirus Infection

Abstract

Prophylactic live attenuated vaccines (LAV) have been successfully developed for multiple viral disease targets including measles, polio, rotavirus, and rabies. From an immunological perspective this modality offers the benefit of simultaneous stimulation of both the humoral and cellular immune responses which is recognized as having an advantage over subunit vaccines, especially for viral diseases for which the correlates of protection are complex and not well understood. However, the development of manufacturing processes for robust and reproducible production of LAVs at commercially viable scales can be quite challenging. Attenuation has commonly been affected by extensive passage of the virus in a permissible cell line. While this method has generally been accepted as safe by regulatory agencies, instances of reversion and even vaccine-induced disease have been reported. The advent of genetic technologies to modify viruses at the molecular level has enabled novel attenuation strategies which carry a much lower risk of revertant events. Many viruses are restricted in their permissible cell tropism and often do not propagate well or at all in suspension cell culture, thus necessitating the need for scale-up of adherent cell lines. Microcarrier culture offers a potential solution but the mechanics are considerably complex. Finally, downstream purification processes for LAVs are limited by a number of factors including low binding capacity for many viruses on traditional chromatography resins and the requirement for closed-system aseptic processes for those viruses which are too large for terminal sterile-filtration. We are developing a LAV for prevention of congenital human cytomegalovirus infection. The presentation will discuss several of the challenges and solutions which have been identified during process development and scale-up to enable Phase II production. These include virus propagation in an adherent mammalian epithelial cell line, purification options for difficult-to-remove host cell protein contaminants, and maintenance of closed-system aseptic processing.

Biography

Joseph G. Joyce received his PhD in Biochemistry from Lehigh University in 1998. In his current role as Director of Vaccine Process Development, he is responsible for downstream CMC process development within the Bioprocess Research and Development organization of Merck Research Laboratories. His major role is to direct teams responsible for designing and implementing robust and scalable processes for preparation of drug substance for vaccine candidates within the clinical development space. Other functions include providing support for early vaccine discovery work and life cycle management for various legacy vaccines. Joe joined Merck in 1989 working as a protein biochemist. For 15 years he led a group responsible for antigen identification, production, and characterization within the vaccine discovery space prior to assuming his current role in clinical development in late 2013. Joe has worked on multiple microbial and viral vaccine programs encompassing a wide variety of vaccine modalities. Important highlights include contributions to the development of licensed vaccines RecombivaxHB® for hepatitis B, and Gardasil® and Gardasil-9® for human papillomavirus infection, where he played a key role in establishing purification and characterization protocols for HPV virus-like particles produced in yeast. Projects which he currently has responsibility for include investigational pneumococcal conjugate vaccines and cytomegalovirus vaccine, both of which are in clinical trials, and contributions to life cycle management for several licensed Merck vaccines including the varicella franchise. Additional vaccine programs which Joe has worked on include HIV-1, Clostridium difficile, Dengue, malaria, and Chlamydia trachomatis.

Nitin Garg
Assistant Director, Upstream Development
Applied Genetic Technologies Corporation (AGTC)
Bioreactor Production of Recombinant Herpes Simplex Virus

Abstract

Recombinant herpes simplex virus type 1 (rHSV)-assisted recombinant adeno-associated virus (rAAV) vector production provides a highly efficient and scalable method for manufacture of clinical grade rAAV vectors. Large quantities of infectious HSV stocks are requisite for these therapeutic applications, requiring a scalable vector manufacturing and processing platform comprised of unit operations which accommodate the fragility of HSV. Here, we present scale up efforts of a replication deficient rHSV-1 vector bearing the rep and cap genes or ITR-gene of interest (GOI) cassette of AAV-2. Adaptation of rHSV production from CS10 vessels to a microcarrier bioreactor permitted a 3-fold increment in cell density and similar increase in infectious vector concentration (pfu/cell). The fed-batch microcarrier bioreactor system afforded a rHSV vector recovery of 2.25E+08 pfu/mL.

Biography

Nitin is a bioprocess technology professional with nine years of varied and increasing responsibility and experience in areas of biotherapeutic, viral vaccine manufacturing, and process development. Prior to joining AGTC in December 2012, Nitin was working with another viral vaccine company based in Tarrytown, New York. He received his MS in biomedical engineering from the University of Arlington, Texas and a bachelor's in biotechnology from Kurukshetra University, India.

Renukaradhya J. Gourapura, DVM, PhD
Associate Professor, Food Animal Health Research Program, Ohio Agricultural Research and Development Center (OARDC), Ohio State University
Nanotechnology-Based Viral Vaccines to Elicit Cross-Protective Immunity in a Pig Model

Abstract

Unlike systemic delivery, mucosal delivery of potent vaccines is gaining attention to increase the breadth of cross-protective immunity against infections which cause disease primarily at mucosal sites. In this direction, my team works on developing new vaccine platforms against influenza and other viral diseases in a pig model. Nanotechnology is an important endeavor of the 21st century. Nanometer-scale materials have favorable physicochemical properties for mucosal vaccine delivery, as their size, shape, charge, and composition could be designed. Our current study in the laboratory is aimed to develop and evaluate the ability of particulate candidate peptide and inactivated influenza virus vaccines delivered by intranasal route as a mist in pigs. For this task, we made use of biodegradable FDA-approved poly (lactic-co-glycolic acid) (PLGA) polymer-based nanoparticles to induce cross-protective immune response. Our experimental results so far demonstrated that such a vaccine delivery platform reduces the clinical influenza symptoms by inducing a robust cross-protective cell-mediated immune response against a virulent zoonotic heterologous influenza virus challenge in pigs. Thus, our research approach has confirmed the utility of a pig model for an intranasal particulate influenza vaccine delivery platform to control influenza in humans.

Biography

Aradhya Gourapura is a mucosal immunologist with focused research accomplishments in the area of nanotechnology-based viral vaccine delivery system studies in a unique pig model. He has 25 years of diverse experience in areas of host-pathogen interactions, development of diagnostics, cancer immunology, and new generation innovative intranasal vaccine delivery platforms. He has DVM and MS degrees from the Veterinary College, Bangalore, and a PhD from the Indian Institute of Science in Bangalore, India. Before joining The Ohio State University in May 2008, Aradhya did his postdoctoral training at the Indiana University School of Medicine for over five years in the area of viral and cancer immunology.

Eevi M. Lipponen
Head of Process Development, FinVector Vision Therapies OY
Process Development for Large-Scale Lentivirus Production
in an Adherent iCELLis 500 Bioreactor

Abstract

Lentiviral vectors are a promising tool for gene therapy. However, scaling up the production methods of lentiviral vectors in order to produce high-quality material for clinical purposes has proven to be challenging. At FinVector, we have developed production of lentiviral vectors using the iCELLis bioreactor (Pall Life Sciences). The disposable iCELLis bioreactor is designed for cultivation of anchorage-dependent cells with a scalable 3D growth area ranging between 0.53 m2–500 m2, an integrated perfusion system, and a controllable environment for production. In this talk we present a scalable and efficient method to produce lentiviral vectors with transient transfection of adherent human embryonic kidney 293T (293T) cells in a fixed-bed bioreactor. Several process parameters were optimized on a small scale and scaled up to production in iCELLis 500. Commercial-scale upstream process development was followed by development of scalable downstream purification.

Biography

Eevi M. Lipponen has worked more than 10 years in the field of gene therapy manufacturing, and is currently focusing on early step process development of several viral vectors, e.g. lentivirus, AAV. She worked for many years developing scalable and robust commercial-scale manufacturing processes under the regulatory guidelines for adenoviral vectors and has extensive experience in GMP work. Her personal interest is in chromatographic downstream purification techniques. Eevi`s current role is Head of Process Development in the Research and Development Department at FinVector Vision Therapies in Kuopio, Finland.

Michael L. Roberts, PhD
Founder & CSO, Synpromics Ltd
Enhancing Viral Vector Manufacturing and
Therapeutic Construct Design with Synthetic Promoters

Abstract

Synthetic biology is a relatively new discipline, having leapt onto the scene over a decade ago as a means to introduce engineering principles to improve and exploit bio-based processes. The vision is that genetic parts can be standardized and taken “off-the-shelf” to build complex biological systems to improve upon various industrial processes. The key to this vision is being able to identify parts that perform in a predictable fashion in any given biological system, and this remains its greatest challenge. At Synpromics we have developed a systematic way in which to construct synthetic promoters that mediate unprecedented control of gene expression. By adopting complex bioinformatics analysis of functional genomics datasets, we are able to create promoters that are active under pre-defined transcription profiles. The approach is broadly applicable across all eukaryotic organisms and we have employed it across a diverse array of applications, from agricultural biotechnology to gene therapy. Dr. Roberts will present data detailing the creation of a set of inducible and constitutive promoters designed to regulate the expression of viral vector helper genes to improve the efficiency of vector manufacturing. He will also discuss results from in vivo studies illustrating how synthetic promoters can be used to mediate predictable expression in different tissues and discuss how Synpromics technology can be used to enhance the efficacy gene and cell therapies.

Biography

Dr. Roberts read biochemistry at the University of Glasgow and completed his PhD at the University of Cambridge, where he employed viral vectors to study plasticity in the peripheral nervous system. He then proceeded to a post-doc position at Royal Holloway, University of London, where he worked on developing novel gene therapies for neuromuscular disorders. In 2002 he moved to Greece on a Marie Curie fellowship to set up a functional genomics facility at the National Hellenic Research Foundation. He then spent five years running gene therapy R&D activities for a small US biotech firm called Regulon. Dr. Roberts moved back to Edinburgh to establish Synpromics in 2010; with a view to develop technology to leverage functional genomics datasets to develop novel synthetic gene promoters. He currently serves as the company’s Chief Scientific Officer.

Hugo R. Soares
PhD Student, Instituto de Biologia Experimental e Tecnológica (iBET)
Virus-Like Particles as Scaffolds for Vaccine Development:
A Case Study of Hepatitis C Virus

Abstract

Hepatitis C virus (HCV) infects nearly 3% of the world’s population, mainly in low-income countries where direct-acting antivirals are difficult to access. Virus-like particles (VLPs) are a particular subset of subunit vaccines which are currently being explored as safer alternatives to live attenuated or inactivated vaccines.

Retroviruses have been widely explored as vectors for gene therapy and as scaffolds for vaccine candidates. One of retrovirus-like particle's (retroVLPs) most attractive characteristics is the ability to incorporate heterologous envelope proteins — known as pseudotyping — as a means to manipulate viral tropism or to present foreign antigens to the immune system. As pseudotyping is a non-selective process, host cellular proteins are also included in the retroVLP membrane. Even though many studies have addressed the identity of these host-proteins, their contribution to retrovirus immunogenicity remained unclear.

Previously we have shown that (i) tetraspanins are the major immunogens present in retroVLPs, (ii) that CD81 is highly incorporated in retroVLPs produced in HEK293 cells inducing specific B- and T-cell immune response in mice, and (iii) that there is an increase in the diversity of tetraspanins in retroVLPs after CD81 depletion. In this work we further explore the potential of using retroVLPs as scaffolds to develop vaccine candidates against hepatitis C, and evaluate the impact of cell host on VLP immunogenicity in mice.

Three cell hosts were assessed in their ability to produce HCV VLPs — Sf9, HEK293, and HuH-7. The impact of cell host on VLP production yields, stability during production and purification, antigen incorporation, and immunogenicity were studied. Preliminary results indicate that retroVLP production yields were maintained at 109 particles/mL in HEK293 and Sf9 cells, while productivity in HuH-7 was significantly lower — 107 particles/mL. The correct assembly of VLPs was confirmed by transmission electron microscopy for all cell hosts. Both mammalian cell lines tested were shown to incorporate higher levels of HCV antigens per VLP and to present glycosylation patterns similar to those reported for infectious HCV viruses. VLPs produced in insect cells incorporated lower amounts of low molecular weight HCV antigens. Overall our work indicates that HEK293 are good cell hosts for the production of HCV VLPs allowing for high quality incorporated antigens.

Biography

Hugo Soares is a final year PhD student in Biotechnology at iBET – Instituto de Biologia Experimental e Tecnológica and at Universidade NOVA de Lisboa. Hugo has a degree in Biology and more than 10 years’ experience working with virus, vaccines, and monoclonal antibodies. Hugo’s PhD project focuses on the study of retrovirus-like particles widely used in gene therapy and vaccine development. His work contributed to understanding the role played by host proteins and cell substrates in the production, purification, and quality of the final particles.

Eduard Ayuso, DVM, PhD
Senior Scientist, Head of Vectorology and Innovation, Université de Nantes
Comparative Studies of AAV2 Manufacturing: Mammalian Versus Insect Cells

Abstract

Medicinal products based on recombinant adeno-associated viral vectors are predominantly manufactured by transient transfection of mammalian cells or baculovirus infection of insect cells, the latter method being more suitable for large-scale production. The scientific community agrees that both methods seem to generate fully functional rAAV particles, however, very few studies have compared these systems in a systematic and exhaustive manner. An international network of scientists has decided to perform such a study using AAV from serotype 2 as an example. To this end, the same AAV vector genome was cloned in a plasmid or a recombinant baculovirus expressing vector and was transfected in HEK293 or infected in Sf9 cells, respectively. For downstream processing, cells were treated with detergent, clarified, and purified by immunoaffinity columns (+/- CsCl gradient). Production yields, purity and in vitro infectivity were measured in both systems and results showed a high degree of comparability. Nonetheless, complementary quality control tests are being performed in different laboratories in a blind fashion and in vivo potency will be tested in mice. The conclusions that will be drawn from this comparative project will be of high interest for the scientific community, regulatory bodies, and for the clinical use of rAAV vectors.

Biography

Eduard Ayuso obtained both his degree in Veterinary Medicine in 2001 and his PhD in Biochemistry and Molecular Biology in 2006 from Universitat Autònoma de Barcelona, Spain. Dr. Ayuso is an expert in several viral vector platforms for gene therapy, such as adenoviral vectors, helper-dependent adenoviral vectors, and adeno-associated vectors. He has made significant contributions to the field of in vivo gene transfer in small and large animal models, particularly in metabolic diseases. His training in viral vectors included stays at the University of Ulm (Germany), the International Centre for Genetic Engineering and Biotechnology (Trieste, Italy) and the Children’s Hospital of Philadelphia (USA). Also, Dr. Ayuso has participated in several academic research programs in partnership with pharmaceutical and biotechnology companies. In 2013, he moved to the Institut National de la Santé et de la Recherche Médicale (INSERM, UMR1089) in Nantes (France) where he leads the “Innovative Vectorology” team with the primary mission of improving the efficacy and safety of recombinant adeno-associated vectors for clinical gene transfer. He is a member of the Board of the French Gene Therapy Society and is a key contributor to the AAV Reference Standard Material Working Group.

Suzanna Melotti
Scientist II, Vector Characterization, bluebird bio Inc.
Physiochemical Characterization of Lentiviral Vectors

Abstract

Lentiviral gene therapy vectors have great potential to enable the treatment of severe genetic diseases using genetically modified CD34+ cells and chimeric antigen receptor T cells (CAR T). This is supported by recent positive data in initial clinical trials showing promising therapeutic benefits and safety. The progress from early to late-stage development entails the need for enhanced characterization of the lentiviral vector product. Lentiviral vector preparations produced by transiently transfected HEK293 cells are complex in nature and contain a mixture of infectious and noninfectious nanoparticles. In this talk, we will outline some of the techniques we have developed to assess the size and composition of lentiviral vector preparations. The pros and cons of each method will be evaluated with example data from each method presented.

Biography

Suzanna leads the lentiviral vector characterization group at bluebird bio.

Amitabha Deb, PhD
Senior Investigator I, Cell Therapy Lab 1
Novartis Institutes for Biomedical Research
Stabilizing Lentiviral Vector Formulations for CAR T-Cell Applications

Abstract

There is growing interest in the use of lentiviral vectors, particularly for cancer immunotherapy and treatment of monogenic diseases. Manufacturing of these vectors is challenging primarily due to the cytotoxic effects of vector components resulting in low cell culture titers and vector instability leading to low purification yields. To improve process scalability, lentiviral vectors were produced and purified from serum-free suspension culture. However, vector aggregation is difficult to control leading to a poor recovery in clarification and sterile filtration through 0.22 micron filters. A high-throughput screen resulted in identifying a number of stabilization buffers, pH, and salt combinations that yielded greater stability of vectors. The formulations have been tested in accelerated stress studies for robustness. Thermolability of lentiviral vectors leads to a requirement for storage at less than -65 °C, even for short-term, in-process intermediates. An inexpensive mixture of buffer and sugar shows stability of vectors for weeks at 4 °C and under multiple freeze-thaw conditions. The selected formulations were tested for transduction in primary T-cells for use in CAR T-cell therapy.

Biography

Amitabha Deb has worked in the pharma industry for the last 16+ years serving different roles in CMC development of biologics and gene therapy products. After completing postdoctoral work with Bryan Williams in Cleveland Clinic on the JAK-STAT pathway and role of NF-kappaB (NF-kB) in inflammation, he joined the biotech industry. In his early years, he served as group leader in downstream purification at MassBiologics. For the last eight years, he has been associated with Novartis Pharma, where he served as Lab-Head for Process Science and Associate Director for Viral Vector Development. Currently he is a Senior Investigator in Immuno-Oncology working on next-generation processes for CAR T-cells.

Marco Schmeer
Project Manager, PlasmidFactory GmbH & Co. KG
Use of Minicircles for AAV Production

Abstract

Especially in gene therapy applications, certain sequence motifs contained in plasmid DNA have to be avoided wherever possible. Such sequences are, e.g. the bacterial ori or selection markers, only used for controlling the bacterial replication of the plasmid or to select for the plasmid during cloning or during production. Such sequence motifs are redundant in the intended therapeutic application and are completely removed in minicircles, i.e. circular and ccc-supercoiled expression cassettes.

Since adeno-associated viral (AAV) vectors are produced by co-transfection of HEK293 producer cells, such bacterial sequence motifs may be an issue of an AAV vector-mediated gene transfer as well. Here, as a result of so-called reverse packaging events, an AAV vector-mediated transfer of not only the therapeutic gene but also of the antibiotic resistance gene into the target cells has been reported. Hence, this appears to be a potential risk of plasmid derived AAV vectors which can be overcome by using minicircle DNA for AAV production.

We could show that both constructs, the helper and packaging plasmid, as well as the transfer plasmid can be produced as minicircles although certain sequence motifs such as the inverted terminal repeat sequences (ITRs) were identified as an issue in minicircle production which has been overcome. These minicircles have been used for efficient AAV vector productions. However, only by replacing both the helper and packaging plasmid as well as the transfer plasmid, encapsidation of the antibiotic resistance gene can be avoided.

Biography

Dr. Marco Schmeer studied chemistry and holds a PhD from the University of Bielefeld, Germany. After further post-doctoral training in the fields of electroporative gene and drug transfer, he joint PlasmidFactory as Product and Project Manager in 2005 with responsibility for client-related plasmid and minicircle production projects.

Axel Schambach, MD, PhD
Acting Director, Institute of Experimental Hematology and Professor
Hannover Medical School
Alpharetroviral Self-Inactivating (SIN) Vectors: Biology, Applications, and Production

Abstract

Gene therapy enabled by integrating retroviral vectors has proven its general effectiveness for applications in basic science and clinical treatment. However, vector integration-associated adverse events related to insertional mutagenesis were documented in clinical trials, fueling the efforts to develop safer vector systems. Of note, alpharetroviruses, originally identified as a cancer-causing agent, have a more random and potentially safer integration pattern, when compared to gammaretro- and lentiviruses. This special integration pattern has likely evolved, as alpharetroviruses transfer their incorporated proto-oncogenes and are thus not dependent on insertional mutagenesis for transformation. Moreover, the special biology of alpharetroviruses is reflected by their inability to replicate in mammalian cells, as they are only replication-competent in bird cells. Here, we outline how alpharetroviruses can be converted into state-of-the-art vectors with improved safety features, capable of efficiently transducing human cells and thus harboring a perspective for clinical applications. Based on a split-packaging technology with high titer production in human cells, we effectively transduced human and murine hematopoietic stem cells as well as human T and NK cells, i.e. target populations for anti-cancer therapies. In this presentation, we will explain and discuss clinically relevant examples, in which alpharetroviral self-inactivating (SIN) vectors could be used for translation into clinical application. In summary, together with a potential production perspective from stable producer clones and the associated upscaling potential, alpharetroviral SIN vectors represent promising tools for clinical application in gene therapy of inherited diseases and anti-cancer immunotherapy.

Biography

Prof. Axel Schambach is Acting Director of the Institute of Experimental Hematology at Hannover Medical School and Professor for gene modification of somatic cells. Axel Schambach studied medicine in Hamburg, San Diego (UCSD), San Francisco (UCSF), Dallas, and Zürich. After his MD he moved to Hannover Medical School to join the Institute of Experimental Hematology and received his PhD in Molecular Medicine in 2005 from the Hannover Biomedical Research School. Since 2007 he has been a group leader for hematopoietic gene and cell therapy at the Excellence cluster REBIRTH, a cluster with a strong focus in regenerative medicine. Since 2012 he has been Associate Faculty and Lecturer at Boston Children’s Hospital (Division of Hematology/Oncology, Harvard Medical School) to develop new gene therapies, including e.g. immunodeficiencies. Prof. Schambach’s main interest is to understand the molecular pathophysiology of inborn and acquired diseases of the blood and immune system and to develop tailor-made gene therapy strategies for selected metabolic diseases and immunodeficiencies. He has authored and co-authored more than 200 publications in this field.

Marian Bendik
Head of Gene Therapy Technologies, Shire
AAV Vector Quantification Strategy for Clinical Trials

Abstract

How can we assure appropriate dose escalation in Phase 1/2 clinical trials? This talk will present a current approach for titering a manufacturing-scale AAV vector product to assign the potency for clinical trials and also will bring an overview of additional characterization methods that are necessary to support the current dosing approach. Further it will also discuss potential causes of undesired variability in gene therapy clinical trials caused by chemistry, manufacturing, and controls (CMC) aspects.

Els Verhoeyen
Research Director, International Center for Infectiology Research (CIRI)/
Enveloped Viruses, Vectors and Innate Responses (EVIR), Ens de Lyon
New Lentiviral Vector Pseudotypes for Gene Therapy of Human Hematopoietic Cells

Abstract

Transductional targeting relies on the modification of the vector surface either by incorporation of foreign envelope glycoproteins that have a natural restricted tropism. The vector specificity is then determined at the level of cell entry and therefore only cells carrying the specific receptor will be transduced.

Lentiviral vectors present a very powerful and flexible tool to mimic the surface of heterologous viruses because they can display heterologous viral envelope glycoproteins at their surface. The original HIV envelope gp120 recognizes the CD4 receptor, which is present on T-cells and macrophages. Fortunately, the HIV envelope can be replaced by the corresponding protein of another virus. This process, which alters the tropism of the vector, is called pseudotyping. Very often, the G protein of vesicular stomatitis virus (VSV-G) is used to pseudotype lentiviral vectors (LVs) because it is highly stable and it recognizes a yet unknown phospholipid receptor which is ubiquitously expressed in mammalian cells. Unfortunately, important hematopoietic target cells such as resting human immune cells (T, B, and DC) or blood stem cells do not express the VSV receptor and are poorly transduced by VSVG-LVs.

A major breakthrough was the engineering of novel lentiviral vectors by our lab that allowed efficient gene transfer into resting immune cells. Indeed, a major limitation of current lentiviral vectors (LVs) such as the generally used VSVG-pseudotyped LVs is their inability to govern efficient gene transfer into quiescent cells such as primary T-cells and B-cells, which hampers their application for gene therapy and immunotherapy. We invented a novel LV incorporating measles virus (MV) H and F surface glycoproteins (MV-LVs). Importantly, a single-cell exposure to these MV-LVs allowed efficient and stable gene transfer of quiescent T-cells and B-cells, which are not susceptible to classical VSVG-LVs. Rather unexpectedly, MV-LVs did not induce cell-cycle entry upon transduction in either quiescent cell type and conserved the naïve or memory phenotypes of transduced resting T-cells and B-cells. Of utmost importance, MV-LVs allowed efficient gene transfer in both healthy and patient plasma B-cells. Additionally, high-level transduction of immature dendritic cells (DCs) was also obtained without induction of their maturation. These MV-LVs also have the extraordinary capacity to be redirected to specific cell types such as CD4, CD8 T-cells, CD19 T-cells, and hematopoietic stem cells. Thus, these MV-LVs are revolutionary for basic research as well as for gene therapy applications.

Another major breakthrough is the development of new lentiviral vectors that carry a baboon endogenous retrovirus (BaEV-LVs) at the surface that efficiently transduces hematopoietic stem cells without any cytokine stimulation, thereby conserving their long-term reconstituting capacity which makes them valuable tools for gene therapy. Moreover, vectors that were able to efficiently transduce were tested for FIX expression and ectopic anti-hepatitis C antibody expression from human B-cells in a humanized mice model. Another important characteristic is that the BaEV-LVs represent a highly efficient new tool to genetically manipulate T-cell acute lymphoblastic leukemia-initiating cells.

Ramjay Vatsan, PhD
Team Lead, Gene Therapy Branch, Division of Cellular & Gene Therapies (DCGT), Office of Tissues and Advanced Therapies (OTAT), FDA CBER
FDA Update on Regulation of AAV, CAR T-Cell, and Gene Editing Products

Biography

Dr. Ramjay Vatsan, PhD is a Team Lead in the Gene Therapy Branch of the Office of Tissue and Advanced Therapies (OTAT) in CBER/FDA. He joined CBER in 2006, and prior to that had worked in basic and translational research at Walter Reed Army Institute of Research (WRAIR)/Naval Medical Research Center (NMRC), the National Cancer Institute (NCI)/National Institutes of Health (NIH), Washington University in St. Louis, and at the National Institute of Immunology, India. Dr. Vatsan has extensive hands-on expertise in the development and use of viral and bacterial vectors used in gene and immunotherapy applications. Dr. Vatsan is an expert in the CMC aspects of gene therapy and immunotherapy and has co-authored a number of CMC guidance documents at OTAT/FDA. Dr. Vatsan is an ASQ Certified Quality Auditor and is a full-time CMC Master Reviewer at FDA.

Haifeng Chen, PhD
CEO, Virovek, Inc.
Manufacturing Adeno-Associated Virus (AAV) in Insect Cells:
Optimizations to Increase Recombinant Baculovirus (rBV) Stability and AAV Yield

Biography

Dr. Haifeng Chen earned his PhD in 1992 from the University of Saarland in Germany. He was then awarded the Marion-Merrell-Dow Post-Doctoral Fellowship to perform studies on adenovirus replication at the University of Kansas Medical Center in Kansas City, Kansas. In 1996, Dr. Chen joined Cell Genesys, Inc. and continued his postdoctoral work focusing on the development of adenovirus and adeno-associated virus (AAV) as gene therapy vectors. In 1997, he took a Research Scientist position at Genovo, Inc. During his time there, he developed two AAV production systems, a baculovirus-based system and an adenovirus-based system. Both resulted in patents and he was promoted to Group Leader in 1998. In 2000, Dr. Chen joined Avigen, Inc. as a Research Scientist focusing on the development of novel AAV vector production technologies. In June 2005, he joined Asklepios BioPharmaceutical, Inc. as Vice President of Production. In June 2006, Dr. Chen started his own company, Virovek, Inc. to provide AAV vector production services to the academic and biopharmaceutical research communities. Besides providing services, Dr. Chen has invented two novel technologies, one for large-scale AAV vector production (patent granted) and the other for production of baculoviral and AAV vectors harboring toxin genes (patent filed), which can be used for cancer gene therapy. Under Dr. Chen’s leadership, Virovek has developed the capacity to provide purified AAV vectors exceeding 3 x 1016 vg per production run. Dr. Chen is currently a member of the editorial board for Molecular Therapy — Methods and Clinical Development, and a member of the Viral Gene Transfer Vector Committee of the American Society of Gene & Cell Therapy (ASGCT). Dr. Chen is also a member of the Society for Neuroscience (SfN) and the American Association for Cancer Research (AACR).

Matthias Hebben, PhD
Head of Bioprocess Development, Généthon
Large-Scale Manufacturing of AAV Vectors: Scale-Up and Transfer of
AAV Production Process for Crigler-Najjar Syndrome Gene Therapy

Biography

After obtaining his PhD in molecular and cell biology in 2001, Matthias Hebben worked both in pharmaceutical industries and biotech companies, occupying several positions related to the virology field: Scientist at VIRBAC, Preclinical Specialist at Intervet/Schering-Plough, and Head of Virology at Vivalis. Since January 2013, he has lead the Bioprocess Development Department at Généthon, a French non-profit organization, where he is in charge of designing and characterizing upstream and downstream processes for manufacturing AAV and lentivirus vectors for gene therapy of rare diseases.

Elizabeth Larimore, PhD
Scientist II, Process Development, Immune Design Corp.
Process Development and Manufacturing of ZVex,
a Lentiviral Vector, for Treatment of Soft Tissue Sarcoma
Gwen Wilmes, PhD
Associate Director, Analytical Development, Dimension Therapeutics
Potency Assay Development for AAV Vectors
Matthias Renner, PhD
Deputy Head, Non-Viral Gene Transfer Medicinal Products
Paul-Ehrlich-Institut (PEI)
Viral Vector Manufacturing and Control: Regulatory Considerations and Challenges


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