1.
Presentation on
Gene therapy
PRESENTED BY
Vishnu Kumar Dhakad
M.Sc. Agricultral Biotechnology

2.
CONTENTS
1.Introduction
2.History
3.Approaches for gene therapy
4.Steps in gene Therapy
5. Gene therapy based on cell line:
6.Gene transfer method: viral and non viral
7.Non viral gene transfer: physical and chemical method
8.Success cases of gene therapy
9.Advantages of gene therapy
10.Disadvantages of gene therapy
11.Ethical issues related to gene therapy

3.
INTRODUCTION
 It is an experimental technique for correcting defective
genes that are responsible for disease development.
PRINCIPLES
 Replacing a mutated gene that causes disease with a
healthy copy of the gene.
 Inactivating a mutated gene that is functioning
improperly.
 Introducing a new gene into the body to help in fight
with the disease.

4.
HISTORY OF GENE THERAPY
 In 1960 the concepts of Gene Therapy was introduced.
 The first approved gene therapy experiment occurred on 14th
September 1990 at the National Institute of Health ,U.K on a four year
old girl named Ashanti DeSliva.
 It was a treatment for a genetic defect that left her with an immune
system deficiency.
 White cell were extracted from her body and after implantation of
genes that produce ADA(Adenosine deaminase), the cells were
transferred back to the girl’s body.
 Considerable improvement in the immune system of the girl was
noticed.

5.
Approaches of gene therapy
1. Gene modification
a) Replacement therapy
b) Corrective Gene therapy
2. Gene transfer
a) Physical
b) Chemical
c) Biological
3. Gene transfer in specific cell line
a) Somatic gene therapy
b)Germ line gene therapy
4. Eugenic approach (gene insertion)

6.
STEPS IN GENE THERAPY
• Identification of defective gene.
• Cloning of normal healthy gene.
• Identification of target cell/tissue/organ.
• Insertion of normal functional gene into the host
DNA.

7.
TYPES OF GENE THERAPY
SOMATIC CELL GENE THERAPY
• Therapeutic genes transferred
into the somatic cells.
• Eg.Introduction of genes into the
bone marrow cells,blood cells,
skin cells etc.
• It will not be inherited later
generations.
• At present all the researches
directed to correct genetic
defects in somatic cells.
GERM LINE GENE THERAPY
• Therapeutic gene transferred into
the germ cells.
• Eg .Genes introducing into the
eggs and sperms.
• It is heritable and passed on to
later generations.
• For safety, ethical and technical
reasons it is not being attempted
at present.

8.
SOMATIC CELL GENE THERAPY: TWO TYPES
In vivo gene therapy: delivery of new genetic material directly to
target cells within the body.
-The challenge lies in ensuring the specificity and reaching the correct
target cells within the body.
Ex vivo gene therapy: target cells are removed from the body and
then genetically modified.
-The cells are then returned to the body after selection and
amplification.
-This is a safe method but dependent on the type of cells being
targeted.

9.
GENE TRANSFER METHODS
The two major classes of methods:
 VIRAL VECTOR-recombinant viruses
 NON VIRAL VECTOR-naked DNA or DNA comlexes
1.VIRAL VECTOR- Viruses have evolved a way of encapsulating and delivering their
genes to human cells in a pathogenic manner. Scientists have tried to harness this
ability by manipulating the viral genome to remove disease causing genes and
insert therapeutic ones.
 It can include both DNA and RNA viral vectors.
1.RNA based vectors:-Retroviruses and HIV(lentiviruses)
2.DNA based vectors:-Adenoviruses and Adeno-associated viruses.

10.
2.NON VIRAL VECTOR
Methods of non-viral gene delivery have also been explored using
physical(carrier free gene delivery) and chemical approaches (synthetic
vector based gene delivery).
Physical approaches ,includes
 Needle injection
 Electroporation
 Gene gun
 Ultrasound (Sonoporation)
 Magnetofection

11.
CHEMICAL METHODS OF GENE THERAPY
1.LIPOPLEXES
 DNA must be protected from damage and its entry into the cell must be facilitated.
 Plasmid DNA can be covered with lipids in an organized structure like a micelle or a
liposome complexed with DNA is called a lipoplex.
 3 types of lipids:
1.Anionic (negatively charged)
2.Neutral
3.Cationic(positively charged)
Common used of lipoplexes
 Transfer of gene into cancer cells, where supplied genes have activated tumor
suppressor control genes in the cell.
 Decrease the activity of oncogenes.
 Useful in transfecting respiratory epithelial cells, so they may be used for treatment of
genetic respiratory diseases such as cystic fibrosis.

12.
2.POLPLEXES
 Complexes of polymers with DNA are called Polyplexes.
 Consist of cationic polymers and their production is regulated by ionic
interactions.
 Large differences compared to lipoplexes is that polyplexes cannot release their
DNA load into their cytoplasm.
3.OLIGONUCLEOTIDE
 Synthetic oligonucleotides are use in gene therapy to inactivate genes
involved in the disease process.
 Strategy involve use of antisense as well as small molecules of RNA called
siRNA.

13.
SUCCESS CASES OF GENE THERAPY
BLINDNESS
PARKINSON’S
DISEASE

14.
GENE THERAPY CURES BLINDNESS
 Cure blindness of inherited condition.
 Inherited disease caused by an abnormality in a gene called RPE65.
 The condition appears at birth or in the first few months of life and causes
progressive loss of vision.
HOW IT WORKS?
 Used harmless viruses.
 Enable access to the cells beneath the retinas of patients.
 By using a very fine needle.
-safe in an extremely fragile tissue and can improve vision in a condition
previously considered wholly untreatable.

15.
GENE THERAPY REDUCES PARKINSON’S DISEASE SYMPTOMS
 It significantly improved the weakness of the symptoms such as tremors, motor skill
problems, and rigidity.
 Main overactive brain region :the subthalamic nucleus should be introduced with gene
that would produce GABA an inhibitory chemical then they could potentially quiet that
brain region and alleviate tremors.
HOW IT WORKS?
 Done with local anesthesia, used a harmless inactive virus[AAV-2GAD].
 Deliver the GAD gene into patient’s subthalamic nucleus.
 The gene instructs cells to begin making GABA neurotransmitters to re-
establish the normal chemical balance that becomes dysfunctional as the
disease progresses.

16.
Researchers are studying gene therapy for many other diseases,
such as
 Severe combined immuno-deficiencies(SCID).
 Hemophilia A &B
 Cancer
 HIV
 Cystic fibrosis
 Muscular dystrophy
 Diabetes

17.
ADVANTAGES OF GENE THERAPY
 In case of ‘silence’ a gene as in the case of HIV, which had not yet
developed into AIDS, scientists could save them the pain and
suffering of the disease by using gene therapy to ‘silence’ the
disease before its onset.
 Gene therapy has the potential to eliminate and prevent
hereditary diseases such as cystic fibrosis and is a possible cure
for heart disease, AIDS and cancer.
 Give a chance of a normal life for those who born with a genetic
disease.
 For certain disease that do not have any cure except gene therapy
it could save many lives.

18.
DISADVANTAGES OF GENE THERAPY
Short-lived nature of gene therapy.
 Immune response - Genes injected with a virus may
trigger an immune response against the virus.
Problems with viral vectors
(once inside the patient, the viral vector could recover its
ability to cause disease).
Multigene disorders - The genetic material might not get
into the right cell, or the right place in the cell’s DNA.

19.
ETHICAL QUESTION SURROUNDING THE
GENE THERAPY
 How can “good” and bad uses of gene therapy be distinguished?
 Who decides which traits are normal and which constitute a
disability or disorder?
 Will the therapy only benefit the wealthy due to its high cost?
 Should people be allowed to use gene therapy to enhance basic
human traits such as height, intelligence, or athetic ability?

20.
Gene therapy: A review article
1Faculty of Pharmacy and Alternative Medicine, the Islamia University of
Bahawalpur, Pakistan.
2Faculty of Eastern Medicine, Hamdard University Karachi, Pakistan.
3Faculty of Pharmacy, Bahauddin Zakariya University, Multan, Pakistan.
Accepted 14 March, 2011
The introduction of nucleic acids into cells has as a purpose of medical condition or
disease. Currently, gene therapy studies a broad range of potential therapeutic
interventions, including the body's immune reaction to tumors, new blood vessels
in the heart to alleviate heart attacks and to stop HIV-replication in patients with
AIDS (Coleman et al., 2003). There is also renewed emphasis on the gene therapy
of genetic diseases, such as hemophilia A and B, and cystic fibrosis. Human gene
therapy experimentation raises many issues. In this review article, background of
gene therapy, introduction, genetic diseases, gene function, germ line gene
therapy, hurdles in gene therapy, methods for gene therapy, ex vivo, in vitro and in
vivo-gene therapy, risks associated with gene therapy

21.
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