Gene Therapy- types, past, present and future, advantages and disadvantages

1. GENE THERAPY
By: Dr. Shruthi Rammohan
2nd Year PG- Dept. of Pharmacology
RRMCH

2. “Doctors of the future
will no longer treat
with medicines…”
- Thomas Edison
“What an amazing time
to be a doctor!”
- Anonymous

3. Why Gene Therapy…
…when we already have excellent
treatments?

5. Patient with a hereditary or genetic disease?
•These patients after diagnosis live their lives on a
daily spoonful of medication.

6. Gene Therapy
• A cure
• A permanent effect
• New gene is inherited
• Sense of well being:
“I AM NORMAL!”
• Hope in living a normal life
Conventional Therapy
• Symptomatic/Palliative Rx
• No effect once drug is stopped
• Disease is inherited
• Constant reminder:
“I HAVE A DISEASE…”
• Maybe improves quality of life

7. 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

8. BOOK COMPONENT BIOLOGICAL EQUIVALENT
Letters Nucleotides
Words Codons
Sentences Genes
Paragraph DNA
Chapter Chromosomes
Books Cells
Library Human Genome
Comparison of Genetic Information with a Library

9. 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

10. GENE THERAPY
A novel approach to treat, cure or prevent a disease
by changing the expression of a person’s genes
Definition:

11. 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
Approaches:

12. Types of Gene Therapy
TYPES
Germ Line
Therapy
Somatic Cell
Therapy
Ex Vivo
In Vivo

13. 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

14. 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

15. Gene Delivery in Somatic Cell Therapy
Somatic Cell
Therapy
Ex Vivo
In Vivo

16. Ex Vivo Approach
• Commonest method of the two
• Called as “Ex Vivo” because the
cells treated outside body

17. In Vivo Approach
• Called “In Vivo” because events
take place within patient’s body
• Uses vectors

18. Vectors for Gene Therapy
The Ideal Vector
• Safe
• Transfection efficient
• Selective
• Persistent gene transfer
• Unrecognized by immune system
Ideal vector is yet to exist!

19. Vectors for Gene Therapy
• Different carrier systems are used for gene delivery
Physical Chemical Biological
Non- viral Vectors Viral Vectors

20. Physical
• Microinjections
• Electroporation
• Sonoporation
• Gene Gun
Advantages: - Effective in transfecting primary, progenitor
and stem cells
Disadvantages: - Cell may sustain heavy trauma and initiate
apoptotic mechanisms
- Requires high precision and accuracy for
success

21. Chemical
• Oligonucleotides
• Liposomes
• Dendrimers
• Calcium phosphate
Advantages: - Target ligand ensures delivery to correct
cells
- Very effective at targeting cancer cells
Disadvantages: - Effective doses can be toxic
- Difficulty transfecting primary, progenitor
and stem cells

22. Biological
• Retrovirus
• Adenovirus
• Adeno associated
viruses
• Herpes Simplex Virus
Advantages: - Integrates into cell efficiently
- High expression of therapeutic gene
Disadvantages: - Immune reactions
- May reactivate in the body
- Transfecting wrong cells

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

24. Gene Therapy Based Drugs
• First country to introduce a gene based drug was China
• Gendicine
• An adenovirus-p53 based gene
• 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
• Alipogene tiparvovec – adeno-associated virus
• Treatment of familial lipoprotein lipase deficiency
• Most expensive medicine in the world ($1.6 million/treatment)
2004
2012

25. 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:
(>5%)
Conjunctival hyperemia, Cataract, increased IOP,
Retinal tear, Eye inflammation, irritation and pain
Cost: $425,000 / eye

26. 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:
(>25%)
Fatigue, Chills, Pyrexia, Nausea, Influenza-like
illness, and Injection site pain
Cost: ~ $65,000 / patient  depends on dosing

27. 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:
(>20%)
Hypogammaglobinemia, Infections, Pyrexia,
Decreased appetite, Headache, Encephalopathy,
Bleeding, Hypotension, Tachycardia, Nausea,
Diarrhea, Vomiting, Fatigue, Acute kidney injury
Cost: $475,000 for entire treatment

28. Successfully Treated Diseases
with Gene Therapy
• Cystic fibrosis
• Haemophilia
• Thalassemia
• Sickle Cell Anaemia
• Familial Hypercholesterolemia
• Severe Combined Immunodeficiency
• Gene therapy is still considered an experimental
discipline

29. Recent Developments
• The majority of gene therapy trials are being conducted in the United States and Europe

30. 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

31. Gene Therapy in India
• Between 2005 – 2013  increased surge in the number of gene therapy research labs
• There is generous financial support from the Indian government
Accomplishments:
Principle Investigator Institution Area of Work Year
Dr. Debi Prasad Sarkar University of Delhi, New Delhi Development of synthetic and
viral delivery systems
2002
Dr. Subrata Banerjee Saha Institute of nuclear physics,
Kolkata
Gene therapy for haematological
disorders
2004
Dr. Kumaravel Somasundaram Indian Institute of Science, Bangalore Gene therapy for cervical cancer 2005
Dr. Jayandharan Rao Center for Stem Cell Research, Vellore Gene therapy for haemophilia
and leukaemia
2010
Dr. Rupesh Dash Institute of Life Sciences,
Bhubaneswar
Gene therapy for prostate cancer 2012
Dr. Arkasubhra Ghosh Narayana Nethralaya, Bangalore Gene therapy using AAV vectors 2013

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

33. 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?

34. Ashanti DeSilva
• 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
First approved gene therapy experiment done in the United States on September 14th, 1990

35. Ashanti DeSilva

36. Jesse Gelsinger
• 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
A victim of a gene therapy treatment that went dreadfully wrong

37. 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

38. 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

39. “The day shall come when every disease has gene therapy”