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PROGRAM CHAIRS:
Joseph A. Fraietta, PhD - University of Pennsylvania
Martin A. Giedlin, PhD - Novartis Pharmaceuticals Corporation

Jeffery N. Odum
Global Technology Partner, Biotech and API, and
Managing Partner, Strategic Manufacturing Concept Group, NNE
The Journey to Design Efficient Cell Therapy Manufacturing Assets

Abstract

The challenge of the Project Design Manager is to optimize the design process to be fast, efficient, cost-effective, and produce a quality, risk-focused end product. For cell therapy-based manufacturing assets, this is a journey less traveled by many today. This presentation will focus on navigating this road-less-traveled, identifying key tools, risk mitigation strategies, and knowledge elements necessary to succeed in integrating the product-process-infrastructure elements of a cell therapy-based manufacturing enterprise.

Biography

Jeff Odum is the Managing Partner, Strategic Manufacturing Concept Group, and a Global Technology Partner at NNE Pharmaplan in the US Office located in Research Triangle Park, North Carolina. He has over twenty-five years of management experience in the design, construction, and commissioning of facilities in the process, biotechnology, pharmaceutical, and chemical industries. A recognized expert in biopharmaceutical manufacturing, Mr. Odum has authored over seventy articles and three industry reference books on subjects related to GMP compliance, process improvement, and the design and construction of biopharmaceutical manufacturing facilities. He is a welcomed speaker at numerous international industry forums and conferences, presenting on topics relating to next generation facility design, bioprocess manufacturing, project management, and GMP compliance. Mr. Odum, a Certified Pharmaceutical Industry Professional (CPIP), served as the North American Education Advisor to the International Society of Pharmaceutical Engineering (ISPE), is a member of the ISPE Biotechnology Community of Practice Steering Committee, and a contributing author to numerous industry baseline and reference guides focused on biotechnology manufacturing, process development, project management, and commissioning and qualification. He is also a member of both the Parenteral Drug Association (PDA) and ISPE technical training faculties, is a teaching fellow in North Carolina State University’s Biomanufacturing Training and Education Center (BTEC) graduate program in biomanufacturing, and a guest instructor for the North Carolina Community College System BioNetwork Program. He has led training efforts in fifteen countries, including training for global regulators from the US FDA, Health Canada, and the CFDA (formerly Chinese SFDA).

Brian Bischoff
Principal Investigator, Cellular Therapy Solutions, LLC
Development of a Microfluidic Device for the Removal
of Glycerol from Cultured Red Blood Cells

Abstract

Recent advancements in the ability to produce blood products from expandable stem cells might be used to create a strategic reserve to reduce the strain on volunteer donors. A strategic stem-cell derived frozen supply may reduce seasonal shortages, enhance the ability to respond to emergencies, and address periodically inadequate supplies of rare blood types. Unlike the donor-based system, where small numbers of units are collected in geographically distributed locations, stem-cell derived red blood cell products (RBCs) will be produced in large volumes at a central location and will be frozen to enable distribution and adequate shelf life. In addition to improving the stem cell-derived manufacturing process, new economical ways of preparing the frozen RBCs for transfusion are needed. The principal obstacle to the use of frozen blood has been the difficulty in removing cryoprotective agents (glycerol) post-thaw to enable clinical use. Red blood cell deglycerolization is typically time-consuming, requires expensive centrifuge-based equipment, and results in high cell losses. Without an easy-to-use and economical method to wash frozen RBCs, the practicality of a stem cell-derived strategic supply will be greatly impaired.

The focus of this presentation is to share early results from our work to develop a microfluidic device capable of removing glycerol from cryopreserved red blood cells. This National Institute of Health (NIH)-funded Phase I investigation describes initial steps to take a prototype device developed in an academic laboratory with funding from National Institutes of Biomedical Imaging and Bioengineering (NIBIB) and bring that invention toward commercialization. The device developed will be easy to use, requiring no external power and improve on cell loss. Our central advancement is a vertically oriented microfluidic device, which can be used to remove glycerol with minimal losses and minimal operator intervention.

Biography

Brian has been developing and commercializing new products for over 24 years and holds 15 patents across a diverse range of fields. He has had the privilege of being the principal investigator on three successful National Institute of Health (NIH) and two Defense Advanced Research Projects Agency (DARPA) grants.

Brian especially enjoys the early stages of identifying, evaluating, and developing new technologies into commercial products. As part of this interest, he developed and teaches a graduate level course at the University of Saint Thomas in St. Paul, Minnesota focused on the unique challenges encountered during the early stages of product development.

Through his work as an advisor for the University of Minnesota’s Discovery Fund, he was introduced to Dr. Allison Hubel’s microfluidic technology, which led to the current NIH-funded project to develop a fluidic device for removal of cyroprotective agents from cells post-thaw. Brian believes that fluidic devices for cell separation, modification, expansion, and inspection will become increasingly important in the manufacturing and point-of-care delivery of cellular therapies.

Kelvin G.M. Brockbank, PhD
CEO, Tissue Testing Technologies LLC
202 — Fundamental Issues in Cryopreservation of Cells and Tissues

Abstract

Cryopreservation methods for cells typically use freezing methods. The major cryopreservation variables are cryoprotectants selection, their concentration, cryoprotectant exposure and removal conditions, pressure, storage temperature, minimization of post-thaw cell death, vehicle solution for the cryoprotectants, and cooling and warming rates. Critical issues in cryopreservation protocols include intracellular ice formation, cryoprotectant cytotoxicity, and apoptosis. Most established cell lines can be preserved adequately in suspension with 5–10% DMSO. However, some cell types, particularly primary cells such as hepatocytes and cardiac myocytes, are difficult and variable from donation to donation. Furthermore, the game changes if the cells need to be cryopreserved on a substrate or if they are within or on a tissue matrix. Cells on tissue culture substrates may require specific growth conditions to prepare them and maintain them on their substrate during cryopreservation by freezing. Cells in tissue matrices, either natural or tissue engineered, often require cryopreservation using ice-free vitrification strategies. Ice-free cryopreservation preserves both the cells and the extracellular matrix integrity. Recent advances in cryoprotectant formulation and rapid warming using Fe nanoparticles and radio frequency inductive heating have overcome some of the barriers to deployment of vitrification for therapeutic tissue products. Finally, validation should always be performed to determine cell viability and cell/tissue function to ensure that the material being preserved is within desired product specifications.

Biography

Dr. Brockbank is the Founder, Managing Partner, and Chief Executive Officer of Tissue Testing Technologies LLC (T3LLC). He previously developed the core technology for two public companies, CryoLife, Inc. and Lifeline Scientific, Inc., formerly known as Organ Recovery Systems. Dr. Brockbank has more than twenty years of project management experience in cell therapies, tissue engineering, medical device research and development, and application of low temperature biology to cell/tissue/organ transplantation. In this work he has also had responsibility for quality systems, regulatory affairs, and clinical research during development of Lifeline Scientific’s LifePort® kidney transport system. His work has been supported by competitive grants from the National Institutes of Health (NIH), the National Institute of Standards and Technology Advanced Technology Program (NIST-ATP), cancer foundations, and the US Department of Defense. He has previously held positions as President, Cell & Tissue Systems, Senior Vice President, Organ Recovery Systems, Inc., Senior Vice President, Life Science Holdings, Inc., Vice President, Research and Development for Edward’s Cardiovascular Surgery Division, Baxter HealthCare Corporation, and Director, Research and Development, for CryoLife, Inc. Dr. Brockbank has also consulted on both business and technical issues for many other companies in the fields of cell therapy and tissue engineering. At present he has a Research Professor academic affiliation with the Department of Bioengineering at Clemson University. He has approximately 200 published research papers and patents related to transplantation, cryopreservation, and tissue banking in addition to more than 250 presentations at national and international conferences. His main interests are in improved preservation of blood vessels, heart valves, cartilage, and tendons as well as development of preservation methods for more complex vascularized composite allografts such as hand and face transplants.

Allison Hubel, PhD
Professor, Mechanical Engineering;
Director, Biopreservation Core Resource (BioCoR), University of Minnesota
Transforming Preservation of Cells Used Therapeutically:
Improved Post-Thaw Function of Mesenchymal Stem Cells

Abstract

Effective methods of preservation are essential for clinical use of cell therapies. It is common for cells to be collected in one location, processed in a second location at a later time, and administered in a third location. The critical biological properties must be retained. We have developed a novel approach to preservation of cellular therapies. A computational algorithm accelerates optimization of composition and cooling rate for a preservation protocol thereby enabling rapid development of a fit-for-purpose protocol. The algorithm uses metrics of function (versus membrane integrity) thereby optimizing post-thaw function of the cells. Cryopreservation solutions developed contain combinations of osmolytes that act in concert to preserve post-thaw function of cells. This approach was applied to the preservation of mesenchymal stromal cells with favorable outcome and this approach can be applied as well to other cell therapies.

Biography

Dr. Allison Hubel is Director of the Biopreservation Core Resource (BioCoR, www.biocor.umn.edu), a national resource in biopreservation. Dr. Hubel is currently a professor in Mechanical Engineering at the University of Minnesota. For more than 20 years, Dr. Hubel has studied both basic science and translational issues behind preservation. Her research focuses on improved methods of preserving cells, development of microfluidic technology to improve preservation, and understanding molecular mechanisms of damage. Dr. Hubel has created and offered professional short courses in the field of preservation and is co-principal investigator (PI) of a training grant in the area. She is a former deputy editor of Biopreservation and Biobanking.

David F. Stroncek, MD
Chief, Cell Processing Section, NIH Clinical Center
Chimeric Antigen Receptor T-Cells: Manufacturing Challenges

Abstract

Clinical trials of chimeric antigen receptor (CAR) T-cells for the treatment of hematologic malignancies have been promising. Currently, CAR T-cells are manufactured using autologous peripheral blood mononuclear cells collected by apheresis. Our center has manufactured clinical CAR T-cell therapies using a variety of different methods. While all methods worked well when tested with cells collected from healthy subjects, we have encountered some problems related to ex vivo cell expansion when manufacturing CAR T-cells for autologous therapy in children and young adults with acute lymphocytic leukemia and osteosarcoma. We found that the presence of large quantities of monocytes or granulocytes in autologous peripheral blood mononuclear cells used to start manufacturing is associated with poor T-cell expansion. The initiation of more rigorous procedures to select lymphocytes from the peripheral blood mononuclear cells have improved CAR T-cell yields, particularly for patients with osteosacrcoma. These results suggest that circulating myeloid-derived suppressor cells are present in patients with cancer and acute lymphocytic leukemia. These suppressor cells are collected by apheresis with mononuclear cells and, if not removed, can inhibit ex vivo expansion.

Biography

David Stroncek, MD is Chief of the Cell Processing Section of the National Institutes of Health (NIH) Clinical Center. His laboratory manufactures and develops a wide variety of Investigational New Drug (IND) cellular and gene therapies to support NIH intramural program Phase I/II clinical trials including chimeric antigen receptor (CAR) T-cells, dendritic cells, natural killer cells, and bone marrow stromal and culture-expanded T-cells. His group also processes hematopoietic progenitors and donor lymphocytes to support hematopoietic stem cell transplantation protocols.

Martin A. Giedlin, PhD
Executive Director, Process Development
Novartis Pharmaceuticals Corporation
Bench to Commercial-Scale Process Development for a CAR T-Cell Therapy

Abstract

Autologous gene-modified adoptive cell therapy is a rapidly emerging option for treatment of refractory/relapsing leukemias and lymphomas. Numerous clinical trials have demonstrated that the use of chimeric antigen receptors (CARs) or modified T-cell receptors (TcRs) targeting these malignancies have a remarkable ability to traffic, expand, and kill target tumor cells post-infusion. However, there are a number of challenges to overcome in order to ensure the safety and efficacy of these cellular-based immunotherapies in a commercial setting. Characterization and control of patient leukapheresis starting material, vector for delivering the target gene, optimal activation/expansion conditions, and in some cases, cryopreservation process and stability, all need to be defined and controlled. This presentation will review an approach utilized to progress these therapeutics from academia to commercial manufacturing and highlight the roles of process development, characterization, and validation, specifically, within the context of a CD19 targeted CAR T immunotherapy.

Biography

Dr. Giedlin is currently the lead for the cellular process development group within Cell and Gene Therapy at Novartis. Prior to this opportunity, he was VP Pharmacology and Manufacturing at Sangamo, supporting the gene-modified adoptive cell therapy programs and the systemic AAV gene therapy projects. He also supported the cancer vaccine programs at Cerus, oncolytic adenoviruses at Onyx, and recombinant proteins, antibodies, and vaccine adjuvants at Chiron. He has his BS in Biology from the University of Notre Dame, his PhD from Virginia Commonwealth University (VCU) Medical Center (MCV), and fellowships at Scripps Research Institute in La Jolla and Dnax in Palo Alto, California.

John M. Baust, PhD
President and Lead Scientist, CPSI Biotech
New Technologies for Enhanced Recovery of Cryopreserved Cells and Tissues

Abstract

Cryopreservation (CP) is an enabling process providing on-demand access to biological material (cells and tissues) whose ultimate use may include starting, intermediate, or final product material. While serving a critical role, CP protocols, approaches, and technologies have evolved little over the last several decades. This often results in a bottleneck for progress impacting cell therapy, tissue engineering, and bioproduction applications. Studies continue to illustrate the impact and benefit of controlling cryopreservation-induced delayed-onset cell death (CIDOCD). In order to overcome these issues, new approaches to CP, including new media and cryoprotective agents (CPAs), molecular control and buffering of cell stress response, and new devices for improved sample freezing and thawing are providing improved sample processing and quality. The majority of these efforts have focused on new “front-end” technologies, such as specialized media or new protective agents. While beneficial to newly developed products or samples not currently processed under established/validated SOPs, they do not provide a path for improving the post-thaw quality of the millions of samples and products currently frozen or preserved under established and validated protocols. Given the known impact of CIDOCD over the initial 24–48 hour (cell type dependent) post-thaw recovery phase, we are developing a series of new molecular based post-thaw conditioning strategies and technologies (RevitalICE) to reduce the impact of CIDOCD in an effort to improve recovery of previously cryopreserved samples. To this end, we investigated the impact of cell stress response modulation during the initial 24 hours post-thaw culture period, examining paths to improve overall survival. Investigations focused on oxidative stress and apoptotic pathway modulation during the post-thaw culture utilizing human endothelial (HUVEC) and smooth muscle (CASMC) cell models. Samples in suspension and in monolayers were cryopreserved utilizing standard controlled rate cooling or a new rapid freezing protocol in various CPAs, and thawed using the novel dry thawing system, SmartThaw. Our studies revealed a significant improvement in cell recovery using the SmartThaw system as a result of rapid and reproducible thawing of samples, regardless of the sample volume or container configuration in comparison with traditional manual water bath thawing. When post-thaw conditioning was incorporated, a further increase in cell recovery was attained. Post-thaw conditioning of endothelial cell samples using n-acetylcysteine or caspase-8 inhibitor resulted in a ~20% and ~35% increase in 24 hour survival, respectively. This improvement in initial survival translated to increased repopulation in culture over a 3 day interval. This recovery media-based conditioning strategy was found to yield improved survival across a number of cryopreservation media and CPA concentrations. The combination of pre-freeze and post-thaw conditioning for freshly cryopreserved samples was also investigated and revealed additional benefit of this combined approach in improving outcome. Overall our findings suggest that there is a viable path for improving recovery of currently cryopreserved samples through the modulation of cell stress response during the initial post-thaw recovery interval (i.e. in culture). This presentation will discuss development efforts focusing on the impact of targeted modulation of the cellular-molecular response to CP as a path to improve cell recovery and function. Data presented will include thermal profile, cell viability, and molecular stress results from several cell systems including CHO, PC3, HUVEC, CASMC, and hMSCs. The objective of these studies is to provide a technology platform enabling improved overall sample viability and function of cryopreserved samples while reducing processing time and end-user variability. Our results suggest that the incorporation of more effective sample thawing devices and protocols can further increase sample quality, process efficiency, and standardization.

Biography

John M. Baust, PhD is the founder, President, and Lead Scientist of CPSI Biotech. Dr. Baust has over 15 years’ experience in research & medical device development and is a co-inventor on over 30 patents. Dr. Baust is a recognized innovator and entrepreneur in cryomedicine and is a pioneer in the area of molecular mechanisms of cell death and low temperature stress. Dr. Baust has published over 100 papers, reviews, book chapters, abstracts, and patents in the area of low temperature biology and has been instrumental in the advancement of the field of cryobiology into the molecular biology era, focusing on signal transduction and apoptosis. In this regard, Dr. Baust is credited with the discovery of cryopreservation-induced delayed-onset cell death. Dr. Baust has led the development of numerous medical devices, including the Supercritical Nitrogen (SCN) and Pressurized Nitrogen (PSN) cryoablation devices for the treatment of cancer and cardiac arrhythmias. This is in addition to spearheading the development of the SmartThaw and SmartFreeze devices for improved cell and tissue cryopreservation. Coupled with these technical engineering developments, he leads life science research programs focused on the cell-molecular actions of cryoablation. These efforts have resulted in the identification of a significant molecular stress response component to freezing injury which is responsible for the differential sensitivity of various cancers to thermal ablation. In addition to these activities, Dr. Baust serves on the editorial boards of Biopreservation and Biobanking, as well as Technology in Cancer Research and Treatment. He is also a reviewer for several other scientific journals. Dr. Baust co-edited the book Advances in Biopreservation, is a past board member of the Society for Cryobiology, and currently serves on the Board and as Treasurer of the American College of Cryosurgery. Dr. Baust completed his studies at Cornell University, State University of New York at Binghamton, and Harvard Medical School.

Kathleen Meyer, PhD
Vice President, Nonclinical Development, Sangamo BioSciences, Inc.
Preclinical Evaluation to Support the First-in-Human Study for Ex Vivo Zinc Finger Nuclease Genome Editing of the CCR5 Locus in Hematopoietic Stem and Progenitor Cells

Abstract

Sangamo Therapeutics is a clinical stage biopharmaceutical company focused on the discovery, development, and commercialization of novel therapeutics for various monogenic and infectious diseases with unmet medical needs. Sangamo’s therapeutic products are based on our engineered zinc finger DNA-binding protein (ZFP) genome editing and adeno-associated virus (AAV) gene therapy platforms. We are leaders in the fields of gene therapy and genome editing. Our proprietary ZFP technology platform enables highly specific genome modification and gene regulation. ZFPs are a naturally occurring class of transcription factors, which we can engineer to bind to any DNA sequence with singular specificity and drive desired therapeutic outcomes.

Our ZFPs can be linked to functional domains that normally activate or repress gene expression to create ZFP transcription factors (ZFP TFs) capable of turning genes on or off. We can also link ZFPs to endonuclease domains to create zinc finger nucleases (ZFNs) which enable precise genome editing in cells. Sangamo's engineered ZFNs can modify a cell's DNA at a precise location, thereby facilitating the correction or disruption of a specific gene or the targeted addition of a new DNA sequence, without the unwanted consequences of off-target DNA cleavage activity. Our ZFP technology can be broadly applied to the development of novel human therapeutics. We have proprietary programs in monogenic diseases that we are developing in the first-ever in vivo human clinical trials in 2017. We also have a preclinical pipeline that is focused on monogenic and rare diseases, as well as diseases of the central nervous system (CNS). This presentation will describe the preclinical safety evaluation programs supporting Sangamo’s first clinical studies using genome editing technology, ex vivo therapy targeting the CCR5 locus in hematopoietic stem and progenitor cells to treat subjects with HIV. CCR5 is a co-receptor for HIV entry into T-cells, and our therapeutic approach aims to use our ZFN-mediated genome editing technology to replicate a naturally occurring human CCR5 knockout mutation which renders individuals largely resistant to infection with the most common serotype of HIV. T-cells that lack CCR5 on their cell surface can only be infected by HIV at low efficiencies. We are using our ZFN-mediated genome editing technology to disrupt the CCR5 gene in cells of a patient’s immune system to make these cells permanently resistant to infection by the most common form of HIV. The aim is to provide a population of HIV-resistant immune cells that are able to fight HIV and its associated opportunistic infections, thereby mimicking the characteristics of individuals that carry the natural CCR5 delta-32 mutation.

Biography

Dr. Kathleen Meyer has served as Vice President, Nonclinical Development at Sangamo Therapeutics since January 2017, and leads the nonclinical development of Sangamo’s zinc finger protein-based genome editing and AAV-based gene therapy candidates. Prior to this, she served as Senior Director, Pharmacology, Toxicology and Bioanalytical Sciences at Sangamo since 2014. Dr. Meyer has over 18 years of industry experience in toxicology, bioanalytical development, pharmacokinetics, and nonclinical safety evaluation strategy and implementation of various biopharmaceuticals, including small molecule, monoclonal antibody, enzyme replacement, and gene therapies. Prior to joining Sangamo, Dr. Meyer served as Principal Scientist, Pharmacology and Toxicology at BioMarin Pharmaceutical Inc. where she guided small molecule and biologic drug candidates through the nonclinical development process supporting clinical trials and marketing authorization. From 2009 to 2012, she served as Senior Director, Nonclinical Safety Evaluation at XOMA LLC and, prior to that, held positions as a Scientist and Principal Scientist at Elan Pharmaceuticals from 1997–2003. Before joining industry, she worked as a postdoctoral fellow evaluating nonviral methods of gene delivery at the University of California, San Francisco. Dr. Meyer received an AB in Physiology, a Master’s degree in Public Health specializing in Toxicology and Epidemiology, and her PhD in Environmental Health Science/Toxicology from the University of California, Berkeley. Dr. Meyer is a Diplomat of the American Board of Toxicology.

Ilya Shestopalov, PhD
Senior Scientist, Cellular Characterization, bluebird bio Inc.
Advanced Cellular Analytics for CD34 Drug Products

Abstract

Cell-based drug products manufactured with lentiviral vectors (LVVs) require advances in analytical methods to characterize their safety and efficacy. This seminar will cover implementation of novel technologies to address unique challenges posed by gene therapy drug products. Cellular drug products are composed of a heterogeneous mixture of phenotypically and functionally distinct cells, and complete phenotypic composition analysis can be challenging. To that aim, we employed high-dimensional mass cytometry (CyTOF) to reliably quantify the phenotypic composition of autologous CD34 drug products. LVV transduction poses a further challenge, as historical assessments of LVV transduction efficiency have relied on calculating an average vector copy number (VCN), yet this measurement does not directly inform on the percentage of cells containing at least one transgene. Quantification of LVV-positive cells (%LVV+) can be complicated by the lack of expression of the transgenic protein in the assayed cells, absence of fluorescent reporters in clinical vectors, and/or lack of suitable methods for detection of transgene expression. To address this challenge, we developed a single-cell PCR assay to detect individual cells with one or more integrations of LVV sequences; enabling the quantification of the %LVV+ cells in a population. Finally, to evaluate drug product potency we developed an assay that utilizes our understanding of diserythropoiesis in beta thalassemia and rescue of erythropoiesis upon transduction with hemoglobin beta (HBB) LVV. Altogether, we demonstrate that unique analytical challenges posed by gene therapy drug products can be addressed with state-of-the-art analytical methods involving CyTOF, single cell PCR, and in vitro disease modeling.

Biography

Ilya Shestopalov, PhD currently leads the cell analytics group within cellular process development at bluebird bio. His research focuses on development of cell-based assays for hematopoietic stem/progenitor and CAR T-cell products. Prior to bluebird bio, Dr. Shestopalov was a postdoctoral fellow in stem cell biology at Boston Children’s Hospital and Harvard University working with zebrafish hematopoietic stem cells. Dr. Shestopalov received his PhD in 2010 from the Chemical Biology program at Stanford University and a BS in Biological Chemistry from the University of Chicago.

Leyla Diaz, PhD
Scientist, BioReliance Corporation
Testing Approaches for Cell-Based Therapeutics:
Rapid Testing and Regulatory Expectations

Abstract

Ensuring the safety and quality of cell-based therapeutic products is achieved through a multi-tiered approach that examines several factors to establish product safety and manufacturing consistency.

Over the past years there has been a dramatic increase in the number of clinical trials involving cellular-based therapies. The nature of these novel therapeutic strategies presents testing challenges when compared to traditional therapies that may include limited shelf life and limited testing sample size as well as a need for reduced turnaround times. As a consequence, existing testing approaches are often not suitable for these products.

We provide an analysis of the current state of the testing regimes undertaken to ensure product quality. Testing strategies for raw materials, production intermediates, and final product are outlined including cell characterization, identity and stability, microbial testing, and detection of adventitious viruses. The implementation and validation of rapid assay methods and suitability of new testing platforms that allow for streamlining testing and reporting are also discussed.

Rachel Legmann, PhD
Manager, Cell Culture PDS Lab & Application Specialist, Pall Life Sciences
Clinical-Scale Manufacturing of Autologous Insulin Producing Liver-Derived Cells
for the Treatment of Diabetes: Culture Dish to a 200 Plate Bioreactor

Abstract

Diabetes is a major global health problem with over 370 million diabetics and an estimated 550 million by 2030. Current therapies rely on recombinant insulin injection to the patients several times a day to control glucose level but do not address the fundamental problem; the loss of insulin producing cells of the pancreas. Orgenesis has developed an autologous cell therapy approach that allows the diabetic patient to be a donor of their own therapeutic tissue. Starting with a small biopsy from a patient’s liver, the biopsied cells are expanded in flatware, trans-differentiated into autologous insulin producing (AIP) cells using three purified adenovirus vectors and then infused back into the patient for long-term amelioration of insulin dependency. To commercialize this cell therapy approach, Orgenesis and Pall have combined their respective expertise to develop a manufacturing process for cell amplification without compromise on cell quality. Xpansion® 200 was used for growing the primary human liver cells under controlled culture conditions to generate cell mass required for curing a diabetic patient. In this study, we have successfully scaled up the cell amplification process to the Xpansion® 200 platform technology while designing a fully closed system. Results showed that 1–2 gr (10–15 million cells) of patient’s liver biopsy was expanded to around 1.8 billion cells in an Xpansion® 200 bioreactor representing more than the targeted dose requirement of 1 billion cells per patient. Results showed that the liver-derived cells had a comparable population doubling time when grown in an Xpansion® bioreactor versus a multilayer system while maintaining cell morphology. The trans-differentiation efficiency demonstrated by similar levels of expression of the pancreatic genes and glucose induced C-peptide secretion. The next step of the study is to focus on incorporating the viral trans-differentiation step into the developed cGMP cell expansion process. As a result, Orgenesis is moving forward with this process for pre-clinical and clinical testing of AIP cells for the treatment of diabetes.

Joseph A. Fraietta, PhD
Associate Director, Product Development & Correlative Studies
University of Pennsylvania
Predictive Correlates of Response to CD19-Directed T-Cell Therapy

Biography

After obtaining his degree in Bioscience and Biotechnology, Dr. Fraietta moved to Philadelphia, Pennsylvania to complete his graduate studies at Drexel University College of Medicine. He conducted research to interrogate the signaling and molecular requirements for the generation of successful effector and memory T cells and to determine why these cells fail to control certain chronic infections. In 2012, Dr. Fraietta graduated with a PhD in Microbiology and Immunology. He then held a position as a research fellow in the Institute for Molecular Medicine and Infectious Disease at Drexel where he discovered and developed novel inhibitors of human immunodeficiency virus (HIV).

Dr. Fraietta continued his training as a postdoctoral scientist in Carl June’s laboratory at the University of Pennsylvania where for the next four years he conducted research on the mechanisms underlying the persistence of chimeric antigen receptor (CAR) T cells in patients with chronic lymphocytic leukemia. In 2016, Dr. Fraietta assumed the directorship of the Process/Product Development Laboratory (PDL) in the first-of-its-kind Center for Advanced Cellular Therapeutics in the Perelman Center for Advanced Medicine. He has published multiple primary research and review articles in the field of T cell immunity to cancer and viruses. Dr. Fraietta also directs graduate as well as undergraduate courses in immunology and virology.

Boro Dropulić, PhD
Chief Science Officer & General Manager
Lentigen Technology Inc., a Miltenyi Biotec Company
Integration of Work Flows for the Manufacture of CAR T-Cell Products

Biography

Dr. Boro Dropulić, PhD, MBA is the Chief Science Officer and General Manager of Lentigen Technology Inc (LTI), a wholly owned subsidiary of Miltenyi Biotec GmbH. Prior to LTI, Dr. Dropulic founded Lentigen Corporation in December 2004 and served as its Chief Scientific Officer and President. Previously, he was the Founder and Chief Scientific Officer at VIRxSYS Corporation, where he successfully led a multidisciplinary team to initiate and complete the first lentiviral vector clinical trial in humans. Prior to that, Dr. Dropulic was an Instructor and Adjunct Assistant Professor at The Johns Hopkins University School of Medicine, where he was the first to develop an HIV-based vector targeted to inhibit the replication of the HIV/AIDS virus. He was previously a Fogarty Fellow at the National Institutes of Health, where he worked on developing transgenic animals using embryonic stem cell technology, understanding molecular aspects of HIV replication and gene therapy for HIV/AIDS. Dr. Dropulic obtained his PhD from the University of Western Australia, focusing upon how viruses gain entry into the brain to cause encephalitis. He also holds an MBA from The Johns Hopkins University and a BSc, with honors, from the University of Western Australia. Dr. Dropulic has served on a number of committees, including grant review committees at the NIH, EU, and CIRM, and as a member and Chair of several committees for the American Society for Gene and Cell Therapy.

Michael C. Holmes, PhD
Vice President, Research, Sangamo BioSciences, Inc.
Molecular Assays for the Detection of Off-Target and On-Target Cutting
Using ZFN in CD34 Autologous Stem Cell Transplants
Cenk Sumen, PhD
Senior Manager, Business Development, PCT, a Caladrius company
Successful Manufacturing of CAR T-Cell Therapies: Challenges and Opportunities
Behnam Ahmadian Baghbaderani, PhD
Head of Cell Therapy Development, Emerging Technologies
Lonza Walkersville, Inc.
Important Design Considerations for the Development of a
cGMP Compliant Manufacturing Process for Cell Therapy Applications
Vijay Chiruvolu, PhD
Senior Director, Product Sciences, Kite Pharma, Inc.
Overcoming Manufacturing Challenges for Production
of Autologous T-Cell Therapy Products
Nirali N. Shah, MD
Staff Clinician, Pediatric Oncology Branch, Center for Cancer Research
National Cancer Institute
Lessons Learned With CAR T-Cell Therapies for Acute Lymphoblastic Leukemia


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