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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
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
“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”.
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
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)
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
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
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
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
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
“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
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
• 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
• 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!
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
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.
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!!
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
