GENE THERAPY

Learning Outcomes

1. Describe gene therapy and its various

forms;
2. Discuss the prevalence of gene
therapy in daily life; and
3. Explore the opportunities that may be
opened by gene therapy in the future.
GENE THERAPY
“Doctors of the future
will no longer treat
with medicines…”
- Thomas Edison

“What an amazing time

to be a doctor!”
- Anonymous
Why Gene Therapy…

…when we already have excellent

treatments?
Patient with a hereditary or genetic disease?

• These patients after diagnosis live their lives on a

daily spoonful of medication.
 Gene Therapy Conventional Therapy
• A cure • Symptomatic/Palliative Rx

• A permanent effect • No effect once drug is stopped

• New gene is inherited • Disease is inherited

• Sense of well being: • Constant reminder:

“I AM NORMAL!” “I HAVE A DISEASE…”

• Hope in living a normal life • Maybe improves quality of life

The Human Genome
• Genetic code found in the DNA of 23 chromosome pairs of
Homo sapiens plus the DNA found in human mitochondria

Haploid Genome: Egg and sperm cells

23 chromosomes
3 billion DNA base pairs

Diploid Genome: Somatic cells

23 chromosome pairs
6 billion DNA base pairs
Comparison of Genetic Information with a Library
BOOK COMPONENT BIOLOGICAL EQUIVALENT
Letters Nucleotides
Words Codons
Sentences Genes
Paragraph DNA
Chapter Chromosomes
Books Cells
Library Human Genome
The Human Genome Project
• Goal  To make a map of the human genome

• The base pair order of 24 chromosomes was to be

determined

• Planning started in 1984

• Launched in 1990 and completed in 2003

IMPORTANCE OF HGP

• Can be used to help replace genes that are defective or missing in genetic diseases

• To predict how a patient will respond to a treatment based on a genetic profile

GENE THERAPY

Definition:

A novel approach to treat, cure or prevent a disease

by changing the expression of a person’s genes
Approaches:
1. Gene Replacement/Correction
–Replacing a mutated gene with a healthy copy
Ex.) Severe Combined Immunodeficiency

2. Gene Silencing/Gene Interference

–Inactivating or “knocking out” a mutated gene
Ex.) Sickle Cell Disease

3. Gene Augmentation/Gene Addition

–Introducing a new gene to help fight disease
Ex.) Parkinson’s Disease

4. “Suicide Gene”
– Can cause a cell to kill itself through apoptosis
– Makes cancer cells more vulnerable and sensitive to anticancer drugs
Ex.) Solid Tumors
Types of Gene Therapy

Germ Line
Therapy
TYPES Ex Vivo
Somatic Cell
Therapy
In Vivo
 Germ Line Therapy
• Altering a gene of an egg or a
sperm cell
or
• Altering the genetic composition of
a blastomere during an early stage
of its division

• Any zygote produced as a result of

this germ cell will have a correct
version of the defective gene and
will continue passing it on to their
offspring

• Considered unethical
Somatic Cell Therapy
• Involves altering the genetic code of a person’s
somatic cells

• Inserting therapeutic genes into somatic cells like:

• Fibroblasts
• Myoblasts
• Epithelial cells
• Nerve cells
• Glial cells

• It is mostly performed in fully grown organisms

• Less controversial
Gene Delivery in Somatic Cell Therapy

Ex Vivo

• Somatic Cell
Therapy
• In Vivo
Ex Vivo Approach
• Commonest method of the two
• Called as “Ex Vivo” because the
cells treated outside body
In Vivo Approach
• Called “In Vivo” because events
take place within patient’s body
• Uses vectors
Vectors for Gene Therapy

The Ideal Vector

• Safe
• Transfection efficient
• Selective
• Persistent gene transfer
• Unrecognized by immune system

Ideal vector is yet to exist!

Vectors for Gene Therapy
• Different carrier systems are used for gene delivery

Physical Chemical Biological

Non- viral Vectors Viral Vectors

 Physical
Advantages: - Effective in transfecting primary, progenitor
• Microinjections and stem cells

• Electroporation Disadvantages: - Cell may sustain heavy trauma and initiate

apoptotic mechanisms
• Sonoporation - Requires high precision and accuracy for
success
• Gene Gun
 Chemical Advantages: - Target ligand ensures delivery to correct
• Oligonucleotides cells
- Very effective at targeting cancer cells
• Liposomes
Disadvantages: - Effective doses can be toxic
• Dendrimers - Difficulty transfecting primary, progenitor
and stem cells
• Calcium phosphate
 Biological
Advantages: - Integrates into cell efficiently
• Retrovirus - High expression of therapeutic gene

• Adenovirus Disadvantages: - Immune reactions

- May reactivate in the body
• Adeno associated - Transfecting wrong cells
viruses

• Herpes Simplex Virus

 Timeline of Gene Therapy
• Scientists were able to deliver target genes into mammalian cells
1977
• Sickle cell disease successfully treated in mice with gene therapy
• 1990 • First human trial experiment with gene therapy
s • First journal published (Human Gene Therapy)
• First death reported due to gene therapy
• Number of diseases with successful gene therapy increases
• 2000
s • First gene therapy drug introduced by China

• 2010 • Focus shifts from single gene defects to multigene defects

+

•2011
• Medical community accepts gene therapy to successfully treat HIV

•2017 • Gene therapy based drugs approved

Gene Therapy Based Drugs
• First country to introduce a gene based drug was China
• Gendicine
• An adenovirus-p53 based gene
2004 • Treatment of patients with head and neck squamous cell carcinoma
• No overt adverse side effects have been reported
• Therapeutic efficacy is still controversial

• Europe came out with first commercially available gene therapeutic

product in the Western world
• Glybera
2012 • Alipogene tiparvovec – adeno-associated virus
• Treatment of familial lipoprotein lipase deficiency
• Most expensive medicine in the world ($1.6 million/treatment)
 2017 Gene Therapy Based Approved Drugs

Trade Name: LUXTURNA

Generic Name: voretigene neparvovec-rzyl
Indication: Confirmed bi-allelic RPE65 mutation-associated
retinal dystrophy
• An adeno-associated virus vector-based gene therapy
• Patients must have viable retinal cells as determined by the
treating physician
MOA: Transduction of some RPE cells with a cDNA
encoding normal human RPE65 protein, thus
providing the potential to restore the visual cycle
Side Effects: Conjunctival hyperemia, Cataract, increased IOP,
(>5%) Retinal tear, Eye inflammation, irritation and pain
Cost: $425,000 / eye
 2017 Gene Therapy Based Approved Drugs

Trade Name: IMLYGIC

Generic Name: talimogene laherparepvec
Indication: Local treatment of melanoma with unresectable
cutaneous, subcutaneous, and nodal lesions
recurrent after initial surgery
• Genetically modified oncolytic viral therapy
MOA: Replicates within tumors to produce immune
stimulatory protein (GM-CSF) causing lysis of
tumors and promoting an antitumor response
Side Effects: Fatigue, Chills, Pyrexia, Nausea, Influenza-like
(>25%) illness, and Injection site pain
Cost: ~ $65,000 / patient  depends on dosing
 2017 Gene Therapy Based Approved Drugs
Trade Name: KYMRIAH
Generic Name: tisagenlecleucel
Indication: Patients up to 25 years of age with B-cell
precursor acute lymphoblastic leukemia (ALL)
that is refractory or in second or later relapse.
• CD19-directed genetically modified T cell immunotherapy
• Vector used is HIV or Lentivirus
MOA: Reprograms patient’s T cells with a transgene
encoding for a chimeric antigen receptor to
identify and eliminate CD19-expressing malignant
and normal cells
Side Effects: Hypogammaglobinemia, Infections, Pyrexia,
(>20%) Decreased appetite, Headache, Encephalopathy,
Bleeding, Hypotension, Tachycardia, Nausea,
Diarrhea, Vomiting, Fatigue, Acute kidney injury
Cost: $475,000 for entire treatment
Successfully Treated Diseases
with Gene Therapy
• Gene therapy is still considered an experimental
discipline

• Cystic fibrosis
• Haemophilia
• Thalassemia
• Sickle Cell Anaemia
• Familial Hypercholesterolemia
• Severe Combined Immunodeficiency
 Recent Developments
• The majority of gene therapy trials are being conducted in the United States and Europe
Recent Developments & Ongoing Trials
Recent gene therapy projects are targeted at conditions such as:

• Cancers - Phase I and II clinical trials for brain, skin, liver, colon, breast and kidney
cancer

• Heart Disease - A phase I clinical trial just completed showing SDF-1 gene therapy
improved HF symptoms

• Parkinson’s Disease - Phase I completed- Modified virus delivers 3 genes to striatum to boost
production of dopamine

• Diabetes Mellitus - Studying approaches of transferring the insulin gene into other cells

• Arthritis - Delivery to synovium achieved with a retrovirus

• Alzheimer’s Disease - Switching off Alzheimer’s gene and using exosomes to deliver drugs to
brain

Major disadvantage is that these diseases are multigenic

Advantages & Disadvantages of Gene Therapy
ADVANTAGES
• Can be used to “silence” a disease before
its onset
Ex.) HIV in AIDS
• Potential to eliminate and prevent
hereditary diseases
• Viral vectors could recover the ability to
• The “Last Chance” or “Last Hope” therapy
• Achieving pharmacological effects
Ex) Making cancer cells susceptible to
anticancer drugs
DISADVANTAGES
• Short- lived nature  Repeated therapy
• Severe Adverse Effects
Ex) Viral vectors may trigger an extreme
immune response
cause disease once inside the patient
• Insertional mutagenesis
(Virus targets wrong cell  new disease)
• Difficulty to effectively treat multigenic
disorders
Ex) Diabetes, Heart Disease
Ethical & Social Considerations

• High cost makes this “promising” therapy available only to the

wealthy

• Can the widespread use of gene therapy make society less accepting
of people who are different?

• Does it interfere with God’s plan?

• Should people be allowed to use gene therapy to enhance basic

human traits such as height, intelligence, or athletic ability?
 Ashanti DeSilva
First approved gene therapy experiment done in the United States on September 14th, 1990
• Born with ADA (adenosine deaminase) deficiency

T- cells taken and placed in a tissue culture

Stimulated to proliferate with IL-2

Infected with a retroviral vector (MoMLV-ADA)

Injected back in a series of treatments

• First patient to be cured by continuous gene therapy

• With gene therapy, she lives a healthy and productive life
Ashanti DeSilva
 Jesse Gelsinger
A victim of a gene therapy treatment that went dreadfully wrong

• Suffered from OTC (ornithine transcarbamylase deficiency)

• Was on a low protein diet taking 32 drugs/day

Using an adenovirus as a vector, his malfunctioning genes were

replaced with healthy ones

As a result, he started to suffer from symptoms not seen in

other clinical trials performed at Penn Hospital

Doctors are still unsure what happened, but suspect extreme

inflammatory reactions due to vector

• First documented death due to gene therapy

Conclusion

• Gene therapy is still new and experimental

• Most EXCITING application of DNA science

• “Genes are the new medicines”

• Many technological, toxicological and ethical issues should be solved

• Holds GREAT PROMISE for curing a number of genetic diseases

References
1. KD Tripathi. 2006. Essentials of Medical Pharmacology, 6th edition. New Delhi
2. King R.C. Stansfield W.D. & Mulligan. P.K. 2006. A dictionary of genetics, 7th ed. Oxford.
3. http://gene-therapy.yolasite.com/ethics.php
4. http://web.ornl.gov/sci/techresources/Human_Genome/redirect.shtml
5. Helmenstine AM. Introduction to the Human Genome Project. Thought Co 2017
6. Kaufmann KB, Büning H, Galy A, Schambach A, Grez M. Gene therapy on the move. EMBO Molecular
Medicine 2013; 5(11):1642-61
7. Misra S. Human gene therapy: a brief overview of the genetic revolution. J Assoc Physicians India 2013;
61:127–33
8. Patil PM, Chaudhari PD, Sahu M, Duragkar NJ. Review article on gene therapy. International Journal of
Genetics 2012; 1(4):74-9
9. Wirth T, Parker N, Herttuala SY. History of Gene Therapy. Gene 525 2013; 162-9
10. Kachroo S, Gowdar SJT. Gene Therapy: An Overview. Gene Technol 2016; 1(5)
11. Haynes MT, Huang L. Lipid Coated Calcium Phosphate Nanoparticles for Nonviral Gene Therapy. Adv Genet 2014;
(88):205-29
12. Gene Therapy Clinical Trials Worldwide. Journal of Gene Medicine 2017
“The day shall come when every disease has gene therapy”