This document discusses gene therapy, including its history, mechanisms, approaches, challenges, and future directions. Some key points:
- As of March 2019, there were over 3,800 gene therapy clinical trials registered on ClinicalTrials.gov.
- Gene therapy introduces genetic material into a person's cells to treat or prevent disease, often using modified viruses as vectors to deliver the gene.
- Approaches include gene replacement/addition therapy, gene editing, and epigenetics. Vectors commonly used are retroviruses, lentiviruses, and adeno-associated viruses (AAVs).
- Challenges include pre-existing immunity to viral vectors, potential immune responses against transduced cells, and safety issues like
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Gene Therapy: An Emerging Approach
1. 1. ClinicalTrials.gov. Search of gene therapy. https://clinicaltrials.gov/ct2/results?cond=&term=gene+therapy&cntry=&state=&city=&dist= (Accessed April 2019). 2. Beitelshees M, et al. Discov Med 2017;24(134):313–22.
•As of March 2019, there were 3831
“Gene Therapy” trials registered on
ClinicalTrials.gov1
1
2. 2
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■ What Is Gene Therapy and How Does It
Work?
■ Gene therapy is an experimental treatment that involves
introducing genetic material into a person’s cells to fight
or prevent disease.
■ A gene can be delivered to a cell using a carrier known
as a “vector.” The most common types of vectors used in
gene therapy are viruses. The viruses used in gene
therapy are altered to make them safe, although some
risks still exist with gene therapy.
■ The technology is still in its infancy, but it has been used
with some success.
Mark Lundie, Ph.D; Medical Director, Rare Diseases, Pfizer Canada
■ 1972 : Friedmann and Roblin publish "Gene therapy for human genetic
disease?" in Science
■ 1990 :First patient to be treated with gene therapy – 4 yr old girl with adenosine
deaminase (ADA) deficiency
■ 1999 : Following Jesse Gelsinger’s death the FDA suspended several clinical trials
pending the reevaluation of ethical and procedural practices.
■ 2012 : Glybera became the first treatment to be approved for clinical use in Europe
■ 2017: Kymriah approved in US for certain pediatric and young adult patients with a form of
acute lymphoblastic leukemia.
■ 2017 : Luxterna (Spark therapeutics) approved for retinal dystrophy due to a mutation of
the RPE65 gene
3. 3
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Genetic medicine
Genetic Medicine Represents a Third Wave of Innovation
Second
wave
~1980s
First Wave
~1900s
Third
wave
Now
Small molecule
Biologics
Symptomatic/
Risk
Reduction
Disease
Modifying
Potentially
Transformational
etanercept
Dyslipidaemia medication
rituximab
aspirin
nonacog alfa
adalimumab
Anti-hypertensive medication
4. Add a functioning gene
Permanently remove,
modify,
or add a gene
Modify gene expression
*Figure from National Institutes of Health: https://commonfund.nih.gov/epigenomics/figure
1. Kumar SR, et al. Mol Ther Methods Clin Dev 2016;3:16034. 2. Cox DBT, et al. Nat Med 2015;21(2):121–31. 3. Lomberk GA, et al. Epigenomics 2016;8:831–42. 4. Epigenetic Therapy as Cancer Treatment.
https://www.slideshare.net/janelle_leggere/efficacy-of-epigenetic-therapy-as-cancer-treatment?from_action=save (Accessed April 2019).
Genetic Medicine: Approaches
Genomic DNA
PAM
crDNA 5´
5´
3´
3´
tracrRNA
Cas9 Nuclease
DNA inaccessible, gene inactive
Gene Replacement / Addition
Therapy
Gene Editing2
Epigenetics3,4*
4
5. Adapted from: Kumar SR, et al. 2016
Kumar SR, et al. Mol Ther Methods Clin Dev 2016;3:16034.
Gene Replacement / Addition Therapy
Vector
Therapeutic
gene
In vivo Ex vivo
Capsid
5
6. 6
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AAV Vector Background
• Single-stranded genome containing gene of
interest (GOI), transcriptional regulatory
elements (enhancer and promoter) and
AAV-ITR (AAV-inverted terminal repeats)
• Size limit about 4.8-5.0 kB
• Extrachromosomal
GOIEnh/Pro
• Genome is encapsulated in a shell
consisting of a viral capsid (cap) protein
• Different capsid serotypes can direct virus
to specific tissues (e.g. AAV9 for muscle)
Intron PolyA
AAV Vector Consists of Two Components
7. Thomas CE, et al. Nat Rev Genet 2003;4:346–58.
Delivery: Vectors
The success of gene therapy critically depends on effective vehicles for gene transfer, which are
based on viral platforms (but are not viruses)
RETROVIRUS
LENTIVIRUSAdeno-associated
virus (AAV)
7
Retrovirus Lentivirus AAV
Genetic material ssRNA ssRNA ssDNA
Tropism Dividing cells only
Dividing and
nondividing cells
Dividing and
nondividing cells
Vector genome
forms
Integrated Integrated
Nonintegrated
(Primarily episomal)
Carrying capacity 8 kb 8 kb <5 kb
8. Figure adapted from Mingozzi F, High KA. (2013).5
1. Ohmori T, et al. J Thromb Haemost 2015;13(Suppl 1):S133–S142. 2. Rapti K, et al. Mol Ther 2012;20:73–83. 3. Nathwani AC, et al. N Engl J Med 2014;137(21):1994–2004. 4. George LA. Blood Adv 2017;1(26):2591–
2599. 5. Mingozzi F, High KA. Blood 2013;122:23–36.
Immunologic Challenges in Gene Therapy
• Pre-formed neutralizing antibodies to
vector capsid1
• Humoral immunity may prevent retreatment2,3
Humoral Immunity Cellular Immunity
• Pieces of capsid displayed on transduced cells4
• T cells respond to vector capsid1,4
• T cells attack / kill transduced cells4
• Loss of cells expressing the donated gene and
producing clotting factor3,4
Vector
T cell
9. 1. George LA, et al. N Eng J Med 2017;377:2215–2227. 2. Nathwani AC, et al. N Engl J Med 2014;371(21):1994–2004. 3. Kattenhorn LM, et al. Hum Gene Ther 2016;27(12):947-961. 4. Baruteau J, et al. J Inherit Metab
Dis 2017;40(4):497–517. 5. Grieger JC, et al. Mol Ther 2016;24(2):287–297.
Future considerations
• Durable efficacy and long-term safety
• Minimizing Immune response
• Optimizing viral vectors
• Multigene disorders.
• Germ line vs somatic cell line gene therapy
• Insertional mutagenesis.
9
10. 1. George LA, et al. N Eng J Med 2017;377:2215–2227. 2. Nathwani AC, et al. N Engl J Med 2014;371(21):1994–2004. 3. Kattenhorn LM, et al. Hum Gene Ther 2016;27(12):947-961. 4. Baruteau J, et al. J Inherit Metab
Dis 2017;40(4):497–517. 5. Grieger JC, et al. Mol Ther 2016;24(2):287–297.
Other considerations
• Capsid and DNA engineering
• Viral tropism for selection of target organ
• DNA optimization (highly expressing gene variants, optimized codons, strong
promotors)
• Reduce immunogenicity; allow for broader treatable populations, possibly allow repeat
infusions
• Manufacturing
• Scalability from experimental studies to clinical supply
10
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Dosing and Immunological response
One ‘Quadrillion’
(1000 Trillion)
100 Trillion
One Trillion
One Hundred Billion
12. For Proactive Use Pfizer Inc / MRC approved 06/23/2017
Importance of Transforming AAV Production Capabilities Enable Broader Gene
Therapy Application
12
AAV=adeno-associated virus; vg=vector genome.
~1 x 1012 vg/roller bottle
40,000x Roller Bottles ~4x iCELLis 500
Attached Cell Reactor
~1x 250 L Cell
Suspension Bioreactor
From: Lab-Scale Culturing To: Contemporary, Scalable Processes
~1 x 1016 vg/iCELLis 500
cell reactor
~4 x 1016 vg/250 L cell
suspension bioreactor
13. 13
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Potential to Transform Therapy for Monogenic Rare Diseases
Liver
Hemophilia B,
Hemophilia A
Muscle
Duchenne Muscular Dystrophy
CNS and LSDs
GAN, ALS, MPS’s
Disorder associated with
lifelong need for 1x to 3x
weekly infusions
Incurable disorder with
shortened life expectancy
Progressive, degenerative
disorders leading to incapacitation
and, potentially, early death
Gene therapy offers the potential to provide
one-time, transformational treatment