advances in gene therapy: eyal grunebaum (the hospital for sick children)
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Advances in gene therapy
Eyal Grunebaum MD Head, Division of Immunology and Allergy Senior Scien<st, Developmental and Stem Cell Biology Hospital for Sick Children, Toronto , Ontario
Canadian Expert Pa<ents in Health Technology Conference November 2016, Toronto
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Educational objectives • What is gene therapy (GT) • Why we need GT (examples from immune def. pa<ents) • How we do GT (outside and inside the body) • When do we now use GT • What innova<on in GT are expected (CAR-‐T, CRISPER).
• Goal: Empower you to be able to advocate effec<vely for GT, when appropriate.
No financial “conflicts of interest”. 2
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Gene therapy: De:inition GT is the introduc<on of gene<c material into cells, which will then be translated by the cell’s machinery to a protein, to compensate for exis<ng abnormal gene or to make a beneficial change to a gene.
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Genes in the DNA are the codes for making proteins. Proteins determine the various traits in our body.
Gene Protein Trait
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“Bubbles” temporary protect kids with severe immune defects
• Children born without an immune system, 2nd to gene<c defects.
• Prone to life threatening infec<ons. • Without appropriate interven<on, condi<on fatal in 1st few years.
• Previously, total isola<on to prevent infec<ons (“bubble babies”) .
David Ve\er (1971-‐1983)
• Not long term solu<on. • Poor quality of life, significant financial & mental challenges.
4 Seinfeld, 1992, “The Bubble Boy” episode, George a\acked by a teenager living in a plas<c bubble, who “losses his mind”.
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Bone marrow transplantations can correct severe immune defects
Transplan<ng bone marrow, harvested from normal donors, to restore immunity following irradia<on, chemo or immune defects (i.e. “bubble babies”).
Erythrocytes
Platelets
White blood cells (immune cells to fight infec4ons)
Hematopoie<c stem cells produce: December 28th 1968
Bone marrow
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h\p://chemosabe-‐socks.blogspot.ca/2013/07/grae-‐versus-‐host-‐disease.html
Defenseless receiving pa<ent (host) Ac<vated immune system
of normal donor (grae), primed to a\ack
You must be new here. I am skin
A major complication of bone marrow transplants: graft vs host disease
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Relocated to a new environment Damage to: Skin Liver Gastro Lungs Joints Etc
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Graft versus host response has major impact on transplant outcome.
Grunebaum E, Mazzolari E, Porta F, Dallera D, Atkinson A, Reid B, Notarangelo LD, Roifman CM. Bone marrow transplanta<on for severe combined immune deficiency. Journal of American Medical Associa<on. 2006.
In North America d/t small families, <20% have HLA iden<cal sibling donor
Gene therapy with pa<ents own “corrected” cells
0 12 24 36 48 60 72 84 96 108 120 132 144 156 168
Months after bone marrow transplantation
100
50
10
60
70
80 90
Sibling donors with identical HLA (92.3%)
Parents, only half matched HLA (52.7%)
Survival (%
)
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Example from pa<ents with severe immune defects
(12.5% have GvHD)
(61.4% have GvHD)
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1: Gene therapy “outside of the body” How is it done?
Cells taken from pa<ent’s BM
A gene of interest is embedded into the viruses’ DNA
“Altered” viruses are mixed with the pa<ent’s cells
The new gene integrates into the cells’ DNA and is expressed as a protein in the pa<ent’s cells
Cells injected into the pa<ent
Altered cells expand & func<on inside the body
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In the lab, viruses (most common gene delivery tool) altered so cannot reproduce or cause harm
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Advantages of gene therapy vs bone marrow transplants include:
• Use pa<ent’s own cells, readily available. • No “grae versus host” response. • No risk of exposure to new infec<ons or other abnormali<es donors might have (and not know about). • Less harm.
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Gene therapy for inherited immune defects. • Pa<ents with adenosine deaminase deficiency, type of inherited severe immune deficiency, were the 1st to receive gene therapy (1990), followed by pa<ents with X-‐linked severe combined ID.
• Done only aeer extensive work in labs (cells, animals, etc). • Used only for pa<ents with no other treatment op<ons. Decade of disappointments: • Difficul<es in introducing the new genes into the cells. • Difficul<es in geqng genes to func<on & produce proteins. • Difficul<es ensuring only 2 gene copies entered (normally there are only 2 gene copies in a cell).
• Difficul<es in controlling the expression of the new genes. • Viruses integrated randomly in the cells’ DNA, ac<va<ng “cancer genes”, leading to leukemia.
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Improvements over time in gene therapy : • Learned that “gene corrected” cells need “head-‐start” to overtake pa<ent’s exis<ng cells low dose chemotherapy used in most GT protocols. • Developed be\er delivery tools with improved safety and efficacy. • Be\er mechanisms to control gene expression, using endogenous promoters (“drivers”) that determine expression. • Enhanced understanding of specific disease biology, thereby choosing condi<ons more likely to benefit from GT. • Earlier iden<fica<on of pa<ents through newborn screening, enabling therapy of kids before becoming sick. 11
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In 2006, Parker was the 1st Canadian to receive “outside” GT (for adenosine deaminase de:iciency) through the “Milan” GT trial, 2016, clinically well, normal immunity.
Aug 2006
Aug 2016
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Long-‐term follow-‐up of gene therapy for ADA de:iciency demonstrates its success • All 18 ADA-‐deficient pa<ents who received GT in the Milan trial are alive. None developed any malignancy. • 90% of them have normal immune func<on. • (Cicalese MP, et al. Update on the safety and efficacy of retroviral gene therapy for immunodeficiency due to ADA deficiency. Blood. 2016) • May 2016: “The European Marke<ng Authoriza<on Commi\ee”, the FDA equivalent, approved commercial use of GT for adenosine deaminase deficiency. [1st out-‐of-‐body GT licensed in Western countries!]
• Clinical trials of GT for ADA deficiency are currently being done in Los Angeles and London. 13
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Current status of gene therapy for immune defects (outside of the body) Clinical trials • Adenosine deaminase def. • IL2Rg deficiency • Chronic granulomatous disease • Wisko\ Aldrich syndrome
Pre-‐clinical research stages • CD40 ligand deficiency • ZAP70 deficiency • RAG1 deficiency • RAG2 deficiency • Artemis deficiency • Leukocyte adhesion defect • Etc
Example: We have been working on GT for PNP deficiency for a decade, and have at least 5 years <ll clinical trials. (Liao P, Toro A, Min W, Lee S, Roifman CM, Grunebaum E. Len<virus gene therapy for purine nucleoside phosphorylase deficiency. J Gene Med. 2008) 14
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Gene therapy for immune defects-‐ remaining challenges. 1. Life-‐long benefits and risks are not known. 2. GT needs to be developed separately for each disease
(>300 genes muta<ons are already known to cause immune defects).
3. Each of these condi<ons requires inves<ng significant resources and many years of research.
4. Limited access in USA, not (yet?) in Canada. 5. Pa<ents and families need to travel to US/Europe. 6. Very expensive (US$250,000/pa<ent). Support by MOH
appreciated, however non-‐sustainable, par<cularly if we plan to increase the # of pa<ents receiving GT. 15
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“Out side of the body” GT for many other non-‐immune conditions
• Gene therapy where bone marrow derived cells are treated with virus outside of the body, and injected back.
• Sickle cell anemia • Fanconi Anemia • Thalassemia
• Metachroma<c Leukodystrophy • Adrenoleukodystrophy
For addi<onal condi<ons: Clinical.Trails.gov
Storage disorders
Hematological diseases
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• DNA of interest delivered directly into the blood or <ssue/organ using viruses (or other vehicles). • Virus inserts itself, and the DNA of interest, into the cells where protein is expressed by the cell’s machinery.
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2. Gene therapy in the body
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Gene therapy directly in the body • Advantages: • No need to remove cells from the pa<ent. • When disease is limited to specific <ssue/organ, the gene directly delivered to <ssue/organ (liver, muscle, brain, tumor, etc).
• More delivery methods are available (viruses, electricity, lipids). • These “delivery methods” can deliver larger genes. • Easy to perform.
• Disadvantages: • The targeted cells usually do not replicate (nor the virus), hence effect is rela<vely short, oeen necessita<ng repeated injec<ons.
• Repeated injec<ons might cause an immune response against the virus, thereby jeopardizing the efficacy of gene therapy.
• Might “infect” and therefore affect neighboring cells. 18
Because of rela<ve ease, became very popular
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• Acute Intermi\ent Porphyria • Spinal Muscular Atrophy 1 • Duchenne Muscular Dystrophy • Limb girdle muscular dystrophy • Amyotrophic lateral sclerosis-‐ (HGF) • Painful diabe<c neuropathy-‐ (HGF) • Leber's Hereditary Op<c Neuropathy • Choroideremia-‐ done in Edmonton • Rare: Neuronal Ceroid Lipofuscinosis • Common: Parkinson’s disease • Very common: Myocardial infarct-‐ into coronary arteries
Direct gene delivery-‐ commonly used
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Into the blood
Into the muscles
Into the brain
Into the eye
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• Skin melanoma (delivers a tumor suppressor molecule). • Recurrent Prostate Cancer (increases chemo uptake). • Advanced stage head and neck malignancies • Breast cancer (delivers IL12) • Advanced Pancrea<c Cancer
• For addi<onal condi<ons: Clinical.Trails.gov
Direct gene therapy very promising in treating
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Cancer!
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Chimeric antigen receptor (CAR)-‐ T cells
Treatment of B‑cell malignancies using anF-‐CD19 CAR T cells. Nat. Rev. Clin. Oncol 2014
T cell ac<va<on T cell
expansion
Refractory lymphoma
Viral delivery of an<-‐CD19 CAR
“sensor”
CAR-‐T infusion
chemo-‐therapy
T cell Isola<on
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“Arm” pa<ents’ immune cells, outside of the body, with an engineered “sensor” that searches for malignant cells
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Chimeric antigen receptor (CAR)-‐ T cells
• Clinical trials of CAR-‐T cells to leukemia, lymphoma, mul<ple myeloma, cervical cancer, and many more.
• Caveats: • Some pa<ents do not have enough T cells. • Difficult to isolate T cells and insert genes into them. • T cells have a short biological half life. • Might a\ack “innocent bystanders” (similar to GvHD) • Long-‐term benefits not known yet. • Accessibility, as very expensive (>$350,000/treatment).
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Next generation gene therapy (1) • Cells source: usage of “induced pleuri-‐potent stem cells” such as pa<ent’s skin cells that are “re-‐programed” into bone marrow cells or T cells, and then are corrected by gene therapy outside of the body.
• Safer delivery tools, including “destruc<on switch” that can be turned on if cells are causing uncontrollable damage, or an “insulator” to prevent effects on neighboring genes.
• More efficient viruses.
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Next generation gene therapy (2) • CRISPER/Cas9 is revolu<onary targeted gene edi<ng technology. • Instead of “adding” an exogenous gene, correct the defect in the exis<ng gene (outside of the body).
• Advantage: use the cell’s own regulatory mechanisms.
• No need to worry about the number of copies inserted.
• However, each defect in each gene needs to be corrected independently (hundreds of muta<ons in each of the hundreds of affected genes.
Very promising technology!!
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Conclusions: Gene therapy has moved from vision to clinical reality • Early, GT was impeded by adverse effects and low efficacy. • Understanding mechanisms led to sophis<cated tools with improved safety and efficacy.
• In recent years, there has been promising progress, sugges<ng that GT is an appropriate treatment approach.
• Further improvements are expected in the near future, par<cularly in controlling gene expression and protein func<on, making gene therapy even more a\rac<ve therapeu<c op<on.
• Remaining biological limita<ons & financial accessibility will need to be addressed by scien<sts and the community, respec<vely.
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Acknowledgments • Suppor<ve medical community (Hospital for Sick Children, The Blood & Marrow Transplant unit, Dr. Roifman & SK colleagues).
• Na<onal and Interna<onal colleagues (Aiu<-‐ Milan, Kohn-‐ L.A.) • Funding agencies (SK Founda<on, D & A Campbell, CIHR, etc). • Ontario Ministry of Health (“Out of Country” sec<on). • !! Trus<ng pa<ents and families !!
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3 of our recent children who received gene therapy