Day 1 :
Keynote Forum
Simone Pacini
University of Pisa, Italy
Keynote: Mesangiogenic progenitor cells (MPCs) in orthopedics, a new tool for cell-based medicinal products?
Time : 10:00-10:40
Biography:
Simone Pacini has completed his PhD in 1997. Until 2003 he was active in oncohematology and responsible of flow cytometry and cell sorting facility. Later, his studies focused on clinical use of MSCs and participated actively on isolation of a novel progenitor of MSC in adult human bone marrow. He is currently working in collaboration reputed Universities and Medical Colleges. Actually, he is a Lab Director of the “Michele Cavaliere Laboratories for Cell Therapy” at University of Pisa. He has published more than 40 papers in reputed journals and he is Editorial Board Member of the Frontiers in Cell Developmental Biology journal.
Abstract:
Mesangiogenic progenitor cells (MPCs) have been firstly described in 2008 in human bone marrow (hBM) mononuclear cell cultures, intended to isolate mesenchymal stromal cells (MSCs) in animal-free conditions. Later, we developed a clinical-grade and selective culture method to isolate MPCs with high grade of purity with yields around 1% of plated cells. MPC are characterized by lack of MSC markers, specific integrin profile and phenotype that include CD31 and surprisingly CD45. From the first report on MPCs, these cells showed both mesengenic and angiogenic potential in vitro. Interestingly, pluripotency-associated gene, as OCT-4, NANOG and nestin expression were also detected. Mesengenic differentiation protocol has been set up in chemical defined conditions and more recently, the angiogenic potential was clearly demonstrated also in vivo, applying MPC constructs on chicken chorioallantoic membrane. Surprisingly, the ex vivo precursor of MPCs in hBM has been identified in CD45dimCD64brightCD31brightCD14neg population with a morphology resembling the monoblast. For their peculiar differentiation properties and clinical-grade isolating methods, MPCs could represent a new tool for the implementation of cell-based medicinal products (CBMPs) applicable for skeletal tissue regeneration, as these cells could also support the neo-vascularization. In fact, future studies on tissue reconstruction should take in consideration that the newly formed tissue growth and integration should be supported by concomitant neo-vessels formation. The co-existence of mesengenic and angiogenic potential in MPCs could significantly improve the regeneration potential of new therapeutic approaches that involve these interesting cells.
Keynote Forum
Jehad Zweiri
University of Liverpool School of Medicine, UK
Keynote: The impact of IR radiation on the induction of bystander killing by genetically engineered ovarian tumor cells: implications for clinical use of cancer vaccines
Time : 10:40-11:20
Biography:
Jehad Zweiri, Lecturer in cancer studies at the University of Liverpool Medical School, received his Bachelor’s degree from the University of Jordan in 1990. He obtained his Master’s degree from London School of Hygiene and Tropical Medicine/University of London, and then obtained his PhD degree in 2000 from Kings College Medical School/University of London, in the field of Immune Gene Therapy of Cancer under the supervision of Professor Farzin Farzaneh. He then started his work as Postdoctoral Associate at the Department of Immunology and Medicine at the University of Liverpool in 2002. In 2010 he was appointed as a Lecturer in the University of Liverpool Medical School and he is currently Fellow of the British Higher Education Academy since 2012.
Abstract:
Cellular based therapeutic approaches for cancer rely on careful consideration of finding the optimal cell to execute the cellular goal of cancer treatment. Cell lines and primary cell cultures have been used in some studies to compare the in vitro and in vivo efficacy of autologous vs. allogeneic tumour cell vaccines. This study examines the effect of g-irradiation on a range of tumor cell lines in conjunction with suicide gene therapy of cancer. To determine the efficacy of this modality, a series of in vitro and in vivo experiments were conducted using genetically modified and unmodified tumor cell lines. Following co-culture of herpes simplex virus thymidine kinase (HSV-TK) modified tumor cells and unmodified tumor cells both in vitro and in vivo we observed that the PA-STK ovarian tumor cells were sensitive to g-irradiation, completely abolishing their ability to induce bystander killing of unmodified tumor cells. In contrast, TK-modified human and mouse mesothelioma cells were found to retain their in vitro and in vivo bystander killing effect after g-irradiation. Characterisation of tumor cell death showed that PA-STK cells underwent pyknosis (necrosis) after g-irradiation. These results suggest that PA-STK cells are not suitable for clinical application of suicide gene therapy of cancer, as lethal g-irradiation (100Gy) interferes with their bystander killing activity. However, the human mesothelioma cell line CRL-5830-TK retained its bystander killing potential after exposure to similarly lethal g-irradiation (100Gy). CRL-5830 may therefore be a suitable vehicle for HSV-TK suicide gene therapy. This study highlights the diversity among tumor cell lines and the careful considerations needed to find the optimal tumor cell line for this type of suicide gene therapy of cancer.
Keynote Forum
Hongjun Wang
Medical University of South Carolina, USA
Keynote: Mesenchymal stem cells from chronic pancreatitis patients prolongs mice and human islet survival
Time : 11:35-12:15
Biography:
Hongjun Wang has completed her PhD from China Agricultural University and Postdoctoral studies at the University of Rome, “La Sapienza” and St. Jude Children’s Hospital. She was an Instructor and Assistant Professor at the Harvard Medical School/Beth Israel Deaconess Medical Center from 2001-2011. She is now a full Professor and the Scientific Director of the Center for Cellular Therapy, at the Medical University of South Carolina, USA. She has published more than 50 papers in reputed journals and has been serving as an editorial board member of repute.
Abstract:
Mesenchymal stem/stromal cells (MSCs) have tissue repair abilities and immunoregulatory effect. We investigated whether MSCs derived from chronic pancreatitis (CP) patients are suitable for use in cell therapy. We first compared MSCs from CP patients with those from healthy donors. We found that cell surface markers, ability of colony formation and multi lineage differentiation abilities were similar between healthy MSCs (H-MSCs) and CP-MSCs. Gene profile study indicated 4 out of 84 human MSC-related genes were differentially expressed in CP-MSCs in comparison with healthy MSCs, among which growth differentiation factor 6 (GDF6), hepatocyte growth factor (HGF) were down-regulated, whereas transforming growth factor (TGF) β3 and matrix metalloproteinase-2 (MMP2) were upregulated. CP-MSCs displayed great potential of inhibition of T cells proliferation to the same extent as healthy MSCs, with even higher indoleamine 2, 3-dioxygenase (IDO) expression upon IFN-γ stimulation. The protective effects of MSCs on hypoxia-induced β cell death are also comparable between CP-MSCs and H-MSCs. We further tested the protective effects of MSC in a marginal mass mouse islet transplantation model, and found that co-transplantation of islets with CP-MSCs improved islet survival and function after transplantation. The effects are in part mediated by paracrine secretion of insulin-like growth factor-1 (IGF-1), suppression of inflammation, and promotion of angiogenesis. In addition, in a pilot clinical trial, co-transplantation of patient islets with autologous MSCs showed safety and primary efficacy. Therefore our rodent and clinical trial data demonstrated that CP-MSCs have the potential to be used as a synergistic therapy to enhance the efficacy of islet transplantation.
Keynote Forum
Farzin Farzaneh
King’s College London, UK
Keynote: Development of therapeutic vaccines against cancer associated antigens
Time : 14:45-15:25
Biography:
Farzin Farzaneh holds the Chair of Molecular Medicine at King’s College London. He has published over 250 research articles with an average citation of over 30, and an impact factor of 47. He has run a licensed GMP facility at King’s College London, since 2001, for the production of cell and gene therapy based investigational medicinal products. He has extensive industrial and academic collaborations, including research council, charitable and pharmaceutical sponsorships of £25 M. He has initiated a number of clinical trials in novel applications of gene therapy and holds MHRA licences (IMPs and “Specials"). He is a Qualified Person (QP) for release of cell and gene therapy products in UK and EU and an Individual Designate under a Human Tissue Authority licence that allows procurement, testing, processing, distribution and/or import/export of tissues and/or cells intended for human applications. He is also appointed by the Commission on Human Medicines, as a member of the Clinical Trials, Biologicals & Vaccines Expert Advisory Group since 2016.
Abstract:
Autologous chimeric antigen receptor (CAR) T cells, one of the most effective of the new immune therapy strategies, are extremely expensive, as they are made for each patient individually. Deletion of the endogenous T cell receptor allows the use of allogeneic CAR T cells, which if proven to be safe and effective, will be substantially cheaper and suitable for large populations of patients. We have helped with the development of allogeneic CAR T cells, now in a number of phase-I clinical trials, providing preliminary evidence of safety and efficacy. In an entirely different form of immune gene therapy, we have shown that myeloid leukaemia cells expressing immune co-stimulatory molecules and appropriate cytokines can induce the rejection of previously established cancers in several mouse models. In two phase-I clinical trials, in poor prognosis acute myeloid leukaemia (AML) patients, we are now assessing the safety and potential efficacy of autologous CD80/IL-2 expressing AML cells. Preliminary data shows evidence of vaccination induced antigen specific cellular immunity and clinical efficacy. In order to develop therapeutic cancer vaccinations, we have identified combinations of adjuvants for synergistic activation of cytotoxicity (CASAC) that can induce antigen-specific cellular immunity, even in immune senescent aged mice. Using a simpler version of CASAC in combination with a selected library of hTERT peptides, we are completing a phase-I trial in patients with therapy resistant, progressive, metastatic disease. Early results indicate safety and stimulation of immunological responses, as well as objective clinical responses and disease stasis in up to 40% of patients.
- Cell Therapy | Tissue Science & Regenerative Medicine | Cell and Gene Therapy for Rare & Common Diseases | Bioengineering Therapeutics | Viral gene therapy | Advanced Gene Therapeutics | stem cell therapies
Location: Olimpica 3+4
Session Introduction
Xuexin Duan
Tianjin University, China
Title: Gigahertz acoustic streaming induced cell membrane poration towards intracellular delivery
Time : 12:15-12:45
Biography:
Abstract:
Efficient intracellular delivery of exogenous materials remains a critical issue in fundamental biological researches and clinical applications. Here, we developed a novel chemical-free method for intracellular delivery enhancement using a designed gigahertz ultrasonic electromechanical resonator. When excited by a sinusoidal electric signal, the propagation and attenuation of acoustic wave in liquid will generate high-speed acoustic streaming. The liquid above the device working area will be accelerated and strike the substrate surface, thus generates pressure on cells, induces deformation and membrane poration, and finally realizes delivery of exogenous materials. To verify the intracellular delivery ability, DOX was selected as an example, and an enhanced fluorescence of DOX in cells exposed to resonator stimulation can be seen. We also realized the delivery of fluorescent-labeled DNA strains and plasmids. Besides, different power applied to the resonator can induce different fluid velocity, thus generate different force intensity and control the deliver efficiency. Pores on membranes induced by acoustic streaming treatment were observed by SEM. Disrupted cell membranes and porous structures can be seen after treatment, and resealed after 10 min recovery, indicating a strong fluid force exerted on cells and the influence is temporary and reversible.
Tiziano Ingegnere
IRCSS Bambino Gesù Pediatric Hospital, Italy
Title: Human CAR-NK cells: A new non-viral method allowing high efficient transfection and target cell killing
Time : 12.45-13:15
Biography:
Ingegnere T has conducted his studies needed for both the first grade degree (laurea triennale) and second grade degree (laurea specialistica) at Sapienza University of Rome. After his graduation he did his PhD at CNR in Rome. From 2014 to 2016 he spent his first Postdoc in the Immunology Lab of Patrizio Giacomini at IFO-Regina Elena Cancer Institute. Since April 2016 he is in the lab of Prof. Moretta as Postdoc in the Immunology Unit of Pediatric Hospital Bambino Gesù.
Abstract:
Cell-mediated immune responses play a central role in the control of infections and tumor growth. In particular, cytolytic T lymphocytes (CTL) and natural killer (NK) cells are fundamental effectors against virus-infected, tumor and leukemia cells. Both T and NK cells are particularly efficient also in allogeneic settings such as the allogeneic haemopoietic stem cell transplantation (HSCT) to cure hematologic malignancies. Another particularly promising approach of cellular therapy is the use of genetically-engineered autologous T cells with chimeric antigen receptors (CAR) conferring specificity for antigens expressed by tumor cells. Also NK cells can be genetically engineered with CAR. Different from CAR-T, NK cells, equipped with an array of receptors involved in tumor cell recognition and killing, retain their ability to target neoplastic cells through such receptors, possibly making tumor escape mechanisms less effective. In addition, they may be complementary to CAR-T cells. However, NK cell transfection resulted quite challenging. Thus, viral transduction display to have variable levels of transgene expression and may compromise NK cell viability. Moreover, viral transduction requires dedicated facilities, high costs and lengthy preparation. Recently, electroporation of mRNA has been proposed as alternative of viral methods. Although the mRNA electroporation has a very low effect on the vitality and good efficacy, a relevant drawback are represented by the short-time expression of the transgene. Here we show a new procedure for NK cells transfection with plasmid DNA. With an efficiency of up to 50% and viability up to 65% it is the most efficient, non-viral, methodology existing so far to deliver exogenous DNA into NK cells. By applying this method, we transfected exogenous CCR7 chemokine receptor conferring to the NK cells the ability to efficiently migrate in response to the chemokines. Moreover, the introduction of an anti-CD19 CAR confer to transfected NK cells a specific and powerful cytotoxicity against CD19+ leukemic cells. These results illustrate some of potential important applications of this novel transfection approach. Notably, the electroporation of DNA may allow to a non-integrating gene transfer with episomal vectors.
Sharmila Fagoonee
Institute of Biostructure and Bioimaging, CNR, Italy
Title: Stem cell therapy for Crigler Najjar syndrome type I
Time : 14:15-14:45
Biography:
Sharmila Fagoonee did her PhD thesis on heme and hemoglobin metabolism at the University of Turin. During her Post-doc studies, she started working in the field of Regenerative Medicine after a training at BIDMC, Harvard, Boston. As a CNR Researcher, she is currently studying new therapies and biomarkers for liver diseases. She is Co-Investigator in several research projects (Telethon, PRIN, Regional grants). She has published more than 142 scientific papers. She has served as Reviewer for several scientific journals, and is an Editorial Board Member of Journal of Clinical Medicine and Minerva Biotecnologica.
Abstract:
The Crigler Najjar syndrome type I (CNSI) is a rare recessive disorder caused by mutations in the Ugt1a1 gene. There is no permanent cure except for liver transplantation, and current therapies present several shortcomings. Since stem cell-based therapy offers a promising alternative for the treatment of this disorder, we evaluated the therapeutic potential of a population of stem cells isolated from cryopreserved hepatocytes known as human liver stem cells (HLSC) in immune-compromised NOD SCID Gamma (NSG)/Ugt1-/- mice, which closely mimic the pathological manifestations in CNSI patients. In order to assess whether HLSC expressed UGT1A1, decellularised mouse liver scaffolds were repopulated with these cells. After 15 days' culture in this 3D setting ex vivo, HLSC differentiated into hepatocyte-like cells expressing markers such as albumin and cytochrome 1a1. For the in vivo human cell engraftment and recovery experiments in the Crigler-Najjar mouse model, NSG/Ugt1-/- mice were generated. A single dose of HLSC was injected in the liver parenchyma of 5 days old phototherapy-treated NSG/Ugt1-/- pups and HLSC functionality and phenotype rescue were assessed in vivo at post-natal Day 21. HLSC expressed UGT1A1 in vivo, induced a decrease in serum unconjugated bilirubin, and improved phenotype and survival compared to untreated controls. A significant reduction in eosinophilic neurons was also observed in HLSC-injected mutant mice hippocampus and cerebellum reflecting recovery from brain damage versus controls. Our results show that HLSC express UGT1A1 in vivo and improve the phenotype and survival of NSG/Ugt1-/- mice, and show promises for the treatment of CNSI.
- Workshop
Location: Olimpica 3+4
Session Introduction
Farzin Farzaneh
King’s College London, UK
Title: The translation of fundamental research discoveries into cell and gene based medicinal products
Time : 15:25-16:10
Biography:
Farzin Farzaneh holds the Chair of Molecular Medicine at King’s College London. He has published over 250 research articles with an average citation of over 30, and an Impact Factor of 47. He has run a licensed GMP facility at King’s College London, since 2001, for the production of cell and gene therapy based investigational medicinal products. Farzin has extensive industrial and academic collaborations, including research council, charitable and pharmaceutical sponsorships of £25M. He has initiated a number of clinical trials in novel applications of gene therapy and holds MHRA licences (IMPs and “Specials"). He is a Qualified Person (QP) for release of cell and gene therapy products in UK and EU and an Individual Designate under a Human Tissue Authority licence that allows procurement, testing, processing, distribution and/or import/export of tissues and or cells intended for human applications. He is also appointed by the Commission on Human Medicines, as a member of the Clinical Trials, Biologicals & Vaccines Expert Advisory Group since 2016.
Abstract:
The therapeutic potential of cell and gene therapies has created a great deal of excitement in a range of biomedical fields. The path from fundamental research, through to licensing applications involves several important components. These include, studies assessing the validity of critical concepts, pre-clinical potential safety and efficacy studies, the design of clinical trials, production of products that are suitable for clinical investigations. Similarly important is the process leading to regulatory authorisation of clinical studies, the efficient conduct of meaningful clinical investigations, proof of concept and pivotal trials through to licensing applications. These processes constitute a complex path, with multiple critical steps and potential bottlenecks, involving important design and implementation challenges.
In an open discussion forum, some of the most prominent of these challenges will be identified and strategies to address them will be examined.