Plenary Session Spotlight

Thursday, May 3

Catherine Bollard, MBChB, MD, Children’s National Medical Center and The George Washington University, United States

Stanley Riddell, MD, Fred Hutchinson Cancer Research Center, United States
Crystal Mackall, MD, Stanford University School of Medicine, United States
Robert Negrin, MD, Stanford University, United States

With Cellular ImmunoTherapy in the spotlight with recent FDA approvals, the Presidential Plenary will focus on Immunotherapy where we are fortunate to have attracted leaders in the field to speak including Stanley Riddell, Crystall Mackall and one of ISCT’s Past Presidents, Robert Negrin. These speakers will focus on the development of novel cell therapeutics for cancer. Dr Riddell will speak on “Beyond CD19-CARs for Hematologic Malignancies”, Dr Mackall will speak on “Cell Therapies for Solid Tumors” and Dr Negrin will speak on “Targeting the Microenvironment”.

Strategies for Improving Efficacy and Safety of CAR T Cells for Cancer Therapy
Stanley Riddell

Advances in synthetic biology and adoptive T cell transfer are making inroads in cancer therapy. Genetically modified T cells that express synthetic chimeric antigen receptors (CARs)ave had success in patients with advanced B cell malignancies. Our lab conducted the first clinical trials in which the CD4 and CD8 T cell composition of the CD19 CAR-T cell product is uniform in all patients. This approach identified CAR-T cell dose/response and dose/toxicity relationships not apparent in prior studies, and improved the therapeutic index in B cell tumors. The outgrowth of tumor cells with low or absent antigen expression remains an obstacle for durable remissions in some patients. A larger challenge is to extend CAR-T cell therapy to common solid tumors, where identifying molecules expressed on cancer cells that can be targeted safely remains an obstacle. We developed CARs that target the receptor tyrosine kinase like orphan receptor ROR1 expressed in many cancers including non small cell lung cancer (NSCLC) and triple negative breast cancer (TNBC). ROR1 is expressed on some normal tissues and we have used a mouse model to examine how various logic gated CARs might redirect selective recognition of tumor cells and not normal tissues.

Allogeneic Cells with Limited GVHD Potential for Cancer Immunotherapy
Robert Negrin

In this presentation I will discuss the role of different effector cell populations that have limited capacity for induction of graft vs host disease and can therefore be used in the allogeneic setting. These cell populations will include natural killer, invariant natural killer T cells and cytokine induced killer cells. The biology of these different cell populations will be illustrated with clinical studies.

Thursday, May 3

Jacques Galipeau, MD, University of Wisconsin-Madison, United States

Martin Hoogduijn, PhD, Erasmus MC, Netherlands
Bruno Péault, PhD, University of Edinburgh, Scotland, and University of California at Los Angeles, United States
Duncan Stewart, MD, The Ottawa Hospital Research Institute, Canada

Mesenchymal stromal cells and related culture adapted cell pharmaceuticals have met important regulatory milestones in the past year with both EMA and likely FDA approvals for clinical use in the making. As the filed strives to develop MSC v2.0, understanding of ontogeny, biology and mechanism of action will inform on how best to improve potency and use of these cell products in expanded clinical applications. The remarkable observation that killed MSCs effect a response from treated mice suggests important unrecognized cell-function autonomous effects of MSC therapy. A point to be addressed by Dr Hoogduijn. On a more basic biology perspective, the deciphering of pericyte form and function in vivo provides important insights on recruitment of endogenous progenitors to affect an injury response. Dr Péault will highlight the biology of vascular fraction-derived MSC-like pericytes and how this knowledge can inform development of this tissue source for translational use. In closing, the translational use of gene engineered endothelial cells foreshadows the development of MSC engineered counterparts, and the clinical lessons learnt and shared by Dr Stewart will provide guidance to the field as well.

Mesenchymal Stem Cells: Native Identity, Diversity, and Culture Induced Alterations
Bruno Péault

Mesenchymal stem cells were indirectly selected by long-term culture for decades before their innate perivascular identity was revealed. Stringently purified peri-endothelial pericytes and stromal cells located in the tunica adventitia of arteries and veins become spontaneously re-programmed into MSCs in vitro. Single cell transcriptome analysis has shown that perivascular presumptive MSCs are ordered as a hierarchy dominated by developmentally more primitive adventitial cells. A proteomic approach has also revealed the expression by perivascular cells of novel markers which typify subsets of functionally distinct presumptive MSCs, in terms of proliferation potential and progenitor ability. In order to understand the molecular transition between native perivascular cells and their MSC descent in culture, total RNA was sequenced from both sorted human perivascular cells and the MSC progeny thereof after 4 passages in vitro. Systematic comparison showed that culture results in dramatic changes in gene expression, up to 33% of all genes expressed becoming modified. This was confirmed by comparing the growth factors and cytokines secreted by either intact perivascular cells or their MSC progeny, which also considerably differ. The general conclusion of these studies is that MSC phenotype and function are largely imposed by in vitro culture.

What do we really know about MSC-mediated immune regulation?
Martin Hoogduijn

In my presentation I will discuss the known immunoregulatory properties of MSC and link them to observations made after infusing MSC for therapy concerning survival and biological activity of the cells. It turns out there are some considerable discrepancies. I will discuss the effects of infusion of inactivated MSC and compare them to the effects of fully active MSC. I will end by proposing a model system to be used for finding out the real immunomodulatory mechanisms of action of MSC.

Gene-enhanced angiogenic cell therapy for cardiovascular diseases: an update on the ENACT-AMI and SAPPHIRE clinical trials
Duncan Stewart

Adult progenitor and stem cells have been used to promote cardiac and blood vessel regeneration but randomized clinical trials have yielded variable results for clinical efficacy. Barriers to the efficacy of cell therapies include limited cell retention or engraftment, heterogeneity and marginal potency of many cell products, and the deleterious effect of host factors of the activity of autologous cell therapies. Our group has optimized the selection and manufacture of a well characterized mononuclear cell product called early outgrowth endothelial progenitor cells (EPC), derived from non-mobilized apheresis collections. To enhance proangiogenic potency, autologous EPCs (aka circulating angiogenic cells) are transiently transfected to overexpress endothelial NO-synthase (eNOS). The ENhanced Angiogenic Cell Therapy for Acute Myocardial Infarction (ENACT-AMI) trial is a multicenter double-blind study with 3-way randomization to saline (control), transfected and non-transfected EPCs delivered up to 30 days post AMI. Change in global EF by MRI is the primary endpoint, and the trial is about half-way to the enrolment target of 100 patients. The Study of Angiogenic cell therapy for Progressive Pulmonary Hypertension: Intervention with Repeat dosing of eNOS-enhanced EPCs (SAPPHIRE) is Canada-wide muticentre trial which has recently initiated enrolment. Up to 8 doses of the eNOS-enhanced EPCs (160M cells in total) will be delivered IV over one year in an innovative 3-arm cross over.

Friday, May 4

Daniel J. Weiss, MD, PhD, University of Vermont School of Medicine, United States

Eva Rohde, MD, Paracelsus Medical University Salzburg, Austria
Linda Marban, PhD, Capricor, United States
Sai-Kiang Lim, PhD, A*STAR Institute of Medical Biology, Singapore

Extracellular Vesicles (EVs) and other particles released by cells are increasingly recognized as a means of cell to cell communication and a powerful means by which physiologic and disease processes are mediated.  However, the field is still in relative infancy and there remain many basic questions about particle identity, isolation and purification techniques, overall homeostasis, and potential use in therapeutics.  These topics will be explored by three leading international experts in EV biology.

Manufacturing and Characterization of MSC-EVs for Clinical Testing
Eva Rohde

Extracellular vesicles (EVs) derived from stem and progenitor cells may have therapeutic effects comparable to their parental cells and are considered promising agents for the treatment of a variety of diseases. Strategies are needed to successfully translate EV research and to develop safe and efficacious therapies.
Based on the experience in manufacturing biological therapeutics for transfusions or for clinical studies, the presentation will focus on the development of mesenchymal stromal cell (MSC)-derived EV-based therapeutics. MSC-EVs are currently under investigation for their immunomodulatory, neuroprotective and tissue regenerative potential.
The requirements for manufacturing, safety, and efficacy testing of MSC-EVs during the translational development are comparable to those for various biologicals. The anticipated heterogeneity and complex molecular structure of EV-therapeutics may require thoroughly defined procedures and standardization of production will be a major issue. The mode of action (MoA) of putative EV-therapeutics will depend on their parental cells and will vary with the intended use for specific target diseases.
While the scientific community, pharmaceutical companies and clinicians are at the point of entering into clinical trials to test various EV-based therapeutics, the identification of the MoA underlying a suggested potency in each therapeutic approach remains a major challenge to the translational path.

Exosomes- CAP-2003
Linda Marban

Capricor Therapeutics is developing CAP-2003, an exosome product, for use in diseases of inflammation and fibrosis. Exosomes are nanometer sized vesicles that serve as cellular messengers. They were discovered over thirty years ago and were originally thought to be involved in cellular waste removal. Since then, exosomes have slowly risen to the fore of scientific curiosity as mediators of everything from metastasis to tissue regeneration. The exosomes in development by Capricor are proprietary to their cell source, the CDC, and have been shown to be markedly immunomodulatory as well as anti-fibrotic and pro-regenerative. Exosomes are a novel way to achieve cellular repair without the living material of the cell. The first indication will be to treat HLHS, otherwise known as hypoplastic left heart syndrome. HLHS is a congenital anomaly of the heart that requires immediate surgical repair at birth followed by two other surgeries during childhood. Exosomes delivered at the time of surgery may delay time to transplant as most of these children develop heart failure and require transplant relatively early in life. Process development of an exosome product will also be discussed.

MSC exosomes and small EVs: a comparative review
Sai-Kiang Lim

The rationale for the therapeutic use of MSCs has been increasingly attributed to their secretion, in particular exosomes or small extracellular vesicles (EVs). The term “small MSC EVs” generally refers to 50-200 nm EVs and is often used interchangeably with the term “exosomes” which refers to similarly sized EVs that are derived from endosomes. In practice, most MSC exosomes are purified by size and not by properties unique to exosomes. Hence most MSC exosome preparations are at best exosome-enriched small EV preparations, and by extrapolation, most small MSC EV preparations despite having a narrow size distribution are heterogenous. This presentation will review the degree of heterogeneity in small MSC EV preparations and the challenges that it poses to understanding MSC exosome biology and applications.

Friday, May 3

Miguel Forte, MD, PhD, Zelluna Immunotherapy, Norway

Carl June, MD, University of Pennsylvania, United States
Pascal Touchon, DVM, MBA Novartis, United States
Diane Parks, MBA, Kite Pharma Inc, United States

The session will cover the recent developments and success with CAR-T therapies.
The vision of translation from academia to industrial development and eventual market launch will be discussed focussed on the recent market launches. This will represent an opportunity to give the audience, both the academics as well as the industry, to hear and discuss the different angles and models that lead to the CAR-T success stories. These examples will be looked with an aim to the future as lessons learned and the impact on the field of cell and gene therapy and the commitment of different stakeholders to this emerging value adding immunotherapy approach to unmet medical needs.


CAR T: Current Status and Perspectives from the Academic Lens
Carl June

Reimagining Cancer Care and Delivering Kymriah to Patients
Pascal Touchon

Saturday, May 5

Donald Kohn, MD, University of California, Los Angeles, United States
Helen Heslop, MD, Baylor College of Medicine, United States

Matthew Porteus, MD, PhD, Stanford University School of Medicine, United States
John Zaia, MD, Beckman Research Institute of City of Hope, United States
Michael Holmes, PhD, Sangamo Therapeutics, United States

In this joint session hosted by ISCT and ASGCT on Genome Editing for Benign & Malignant Diseases, recent advances in gene editing will be reviewed.
Presentations will include: Ex Vivo Gene Modification of HSC by Matthew Porteus (Stanford University); ZFN Targeting of CCR5 in HSC for HIV by John Zaia (City of Hope Medical Center); and In Vivo Targeting of Liver by Michael Homes (Sangamo Biotherapeutics). The goal of the session is to provide attendees with the latest information on the approaches and status of genome editing.

Gene Editing of Hematopoietic Stem and Progenitor Cells
Matthew Porteus

Genome editing provides a method to precisely change the DNA sequence of a cell, including somatic stem cells. We have developed a system that combines the use of CRISPR/Cas9 delivered as ribonucleoprotein complex with AAV6 transduction to generate high frequencies of geneome editing in a wide variety of human cell types including hematopoietic stem cells, T-cells, mesenchymal stromal cells, airway basal stem cells, neural stem cells and pluripotent cells. In this presentation I will discuss our application of this system to engineering human cells to treat human disease.

Genome editing with ZFN using blood progenitor cells in HIV/AIDS
John Zaia

Gene therapy and genome editing are modalities with application to HIV/AIDS, and this presentation will review the current state of the field for this area of clinical research. The speaker will then focus on the feasibility issues regarding scale-up for ZFN-based genome editing of hematopoietic stem/progenitor (HSPC) cell transplantation. The first clinical trial of CCR5-targeted ZFN-modified HSPC is being performed in HIV/AIDS patients, and the laboratory and regulatory hurdles required for initiation of this first-in-human trial will be reviewed. This active outpatient study has now treated 7 subjects, and selected preliminary results will be reviewed. All patients have engrafted with CCR5-disrupted cells and the conditioning regimen, utilizing busulfan, was well tolerated in these HIV-infected subjects.

Highly efficient and specific multiplexed gene editing in T cells using enhanced zinc-finger nucleases (ZFNs) enables strategic engineering of allogeneic T cell immunotherapies
Michael Holmes

Engineered allogeneic CAR-T cells generated from healthy donors would provide a well-characterized, consistent off-the-shelf product that could be administered to a broad population of patients. To pursue this approach, a highly efficient and precise gene editing capability is imperative to generate the desired T cell product profile: TCR/CD3-negative achieved by knocking out T-cell receptor alpha constant region (TRAC), HLA class I-negative achieved by disrupting HLA complex formation by knocking out the β2-microglobulin (β2M) gene, and positive expression of an antigen-specific CAR or TCR achieved by site-directed gene insertion into the disrupted TRAC locus.
Designed ZFNs provide an attractive platform for therapeutic genome editing, as they can be designed to target virtually any site in the genome with a high degree of precision, efficiency, and specificity. Using ZFNs, we have developed multiplexed gene editing capabilities that can simultaneously disrupt the TRAC and β2M loci at a double knock-out (KO) efficiency of >90%, and achieve targeted insertion of a transgene into TRAC at >90%. Furthermore, a rigorous and deep molecular interrogation of the ZFN-treated T cells revealed undetectable off-target nuclease activity, highlighting the specificity of our enhanced ZFN technology. These industry leading capabilities enable the development of defined and well-characterized allogeneic CAR-T cell products for application in hematological malignancies.

Saturday, May 5

Andras Nagy, PhD, Lunenfeld-Tanenbaum Research Institute, Sinai Health System, Canada

Maria Mirotsou, PhD, Astellas Institute of Regenerative Medicine, United States
Mahendra Rao, MD, PhD, New York Stem Cell Foundation, United States
Andras Nagy, PhD, Lunenfeld-Tanenbaum Research Institute, Sinai Health System, Canada

A solution for cell therapy safety and engineered allotolerance
Andras Nagy

Pluripotent stem cells have accelerated the development of new avenues for targeting degenerative diseases. Numerous cell therapies are currently on their way to treat devastating conditions. However, concerns about the cell-safety hold back the full utilisation of these promising new treatments. Here I introduce a concept and show the associated genome engineering strategy that addresses this issue and provides a solution for “fail-safe” cell therapies.
Using published and our experimental measures, we defined mathematically the level of safety of therapeutic batches of cells. Our general approach to assess and quantify the safety will be critical to make informed decisions by the regulators, doctors, and patients to advance this modern medicine-transforming therapies.
Building on the fail-safe technology, we addressed the next hurdle faced by cell therapies; a solution for induced allograft tolerance. I show that the expression of eight local-acting, immunomodulatory transgenes introduced into embryonic stem cells is sufficient to protect cell derivatives against rejection in allogenic, immune-competent recipients. Allografts survive long-term, in different MHC-mismatched recipients, and without immunosuppressive drugs.
The combination of the fail-safe and immune tolerance genome editing will make the “One for All” cell line and therapeutic cell development a reality.