The document presents an overview of gene therapy, discussing its definition, approaches, types, and potential applications in treating various genetic disorders, including cystic fibrosis, Parkinson's disease, and cancer. It covers the history, methods of gene delivery, vectors used, and the advantages and disadvantages of gene therapy. The presentation highlights the complexities and challenges associated with gene therapy, such as the immune response and limitations when dealing with disorders caused by multiple genetic defects.

1. PRESENTATION ON
GENE THERAPY
Presented to:
Himanshi Ma'am
Presented By:
Vikas Dagar
Roll No: 190121220010
M. Pharma (P’Ceutics)
2ND Semester
DEPARTMENT OF PHARMACEUTICAL SCIENCES,
GURU JAMBHESHWAR UNIVERSITY OF SCIENCES & TECHNOLOGY, HISAR

2. CONTENTS
• INTRODUCTION
• GENE THERAPY
• What are Genetic Disorders?
• Several approaches to gene therapy
• Why gene therapy?
• HISTORY OF GENE THERAPY
• TYPES OF GENE THERAPY
• IN VIVO GENE THERAPY
• EX VIVO GENE THERAPY
• VECTORS IN GENE THERAPY
• METHOD OF GENE DELIVERY IN PHYSICAL METHOD
• CHEMICAL METHOD
• POTENTIAL TARGET DISEASES
• ADVANTAGES
• DISADVANTAGES

3. INTRODUCTION
• Gene therapy is the introduction of genes
into existing cells to prevent or cure a wide
range of diseases.
• It is a technique for correcting defective
genes responsible for disease
development .
• The first approved gene therapy
experiment occurred on september
14,1990 in US, when Ashanti DeSilva was
treated for ADA-SCID.

4. GENE THERAPY
• Gene therapy is experimental technique that
uses genes to treat or prevent disease. In the
future, this technique may allow doctors to
treat a disorder by inserting a gene into a
patient’s cells inserted of using drugs or
surgery .
• Genes are carried on chromosomes and the
basic physical and functional units of
heredity.
• Genes are specific sequences of nucleotides
that encode instruction on how to make
proteins.

5. What are Genetic Disorders?
 Genetic Disorder is a disease caused by a
“variation” or “mutation” of a gene.
Genetic Disorder can be passed on to family
members who inherit the genetic abnormality.
A small number of rare disorder are caused
by a mistake in a single gene.
Most disorder involving genetic factors, such
as heart disease and most cancers, arises
from a interplay of multiple genetic changes
and environmental factors.

6. Several approaches to gene therapy:
1) Inserting a normal gene to replace
abnormal gene.
2) Inactivating, or “knocking out ,” a mutated
gene that is functioning improperly.
3) Introducing a new gene into the body to
help fight a disease.

7. Why gene therapy?
 Gene therapy can be used for a number
of diseases, such as severe combined
immune- deficiencies, hemophillia ,
Parkinson’s disease, cancer and even HIV
through a number of different approaches.
This technique may allow doctors to treat
a disorder by inserting a gene into a
patient’s cells inserted of using drugs or
surgery.

8. Vector and its ideal properties
• TARGET the right cells.
• INTIGRATE the gene in the cells.
• ACTIVATE the gene.
• AVOID harmful effects.
• No universal vector exists.

9. HISTORY OF GENE THERAPY
• 1953: scientists Francis Crick & James
Watson determined double helical structure
of DNA.
• 1973: American doctor Stanfeild Rogers tried
to treat sisters with Hyperargininemia using
human pappiloma virus.
• 1980: Dr. Martin Cline – first attempted at
human gene therapy in university of
California , L.A.
• 1984: the human gene therapy working group
(HGTG) created.

10. • 1999: Death of Jesse Gelsinger , the first
casuality in gene therapy.

11. TYPES OF GENE THERAPY
SOMATIC CELL GENE
THERAPY
• Therapeutic genes
transferred into
somatic cells.
• Eg. Introduction of
genes into bone
marrow cells, blood
cells, skin cells etc.
GERM LINE GENE
THERAPY
• Therapeutic gene
transferred into the
germ cells.
• Eg. Genes
introduced into
eggs & sperms.

12. SOMATIC CELL GENE
THERAPY
• Will not be
inherited later
generation.
• At present all
researches
directed to correct
genetic defects in
somatic cells.
GERM LINE GENE
THERAPY
• It is heritable &
passed on to later
generations.
• For safety, ethical
& technical
reasons, it is not
being attempted at
present.

13. IN VIVO GENE THERAPY
• Direct delivery of therapeutic gene into target
cell into patients body.
• Carried out by viral or non viral vector
systems.
• It can be the only possible option in patients
where individual cells cannot be cultured in
vitro in sufficient number(e.g. brain cells).
• In vivo gene transfer is necessary when
cultured cells cannot be re-implanted in
patients effectively.

14. Example of IN VIVO gene therapy
• In patient with cystic fibrosis, a protein called
cystic fibrosis transmembrane regulator
(CFTR) is absent due to a gene defect.
• In the absence of CFTR chloride ions
concentrate within the cells and it draws
water from surrounding.
• This lead to the accumulation of sticky
mucous in respiratory tract and lungs.
• Treated by in vivo replacement of defective
gene by adenovirus vector.

15. EX VIVO GENE THERAPY
Isolate cells with genetic defect from a
patient
Grow the cells in culture
Introduce the therapeutic genes.
Select genetically corrected cells and grow.
Transplant the modified cells to the patient.

16. Example of EX VIVO gene therapy
• 1st gene therapy – to correct deficiency of
enzyme, Adenosine deaminase (ADA).
• Performed on a 4yr old girl Ashanthi
DeSilva.
• Was suffering from SCID- severe
combined Immunodeficiency.
• Caused due to the defect in gene coding
for ADA.
• Deoxy adenosine accumulate and
destroys T lymphocytes.

18. VECTORS IN GENE THERAPY
• To transfer the desired gene into a target
cell, a carrier is required. Such vehicles of
gene delivery are known as vectors.
• 2 main classes
- Viral vectors
- Non viral vectors

19. VIRAL VECTORS
1) RETROVIRUS VECTORS SYSTEM
The recombinant retroviruses have the
ability to integrate into the host genome in a
stable fashion.
• Can carry a DNA of
size- less than 3.4kb
• Replication defective virus particles
• Target cell dividing

20. 2) ADENO VIRUS VECTOR SYSTEM
• Adeno virus with a
DNA genome
-good vectors.
• Target- non dividing
human cell.
Eg . Common cold
adenovirus

21. 3) ADENO ASSOCIATED VIRUS VECTOR
• It is human virus that can integrate into
chromosome 19.
• It is a single stranded, non pathogenic small
DNA virus.
• AAV enter host cell , becomes double stranded
and gets integrated into chromosomes.

22. 4) HERPES SIMPLEX VIRUS VECTOR
• Viruses which have natural tendency to
infect a particular type of cell.
• They infect and persist in nervous cells.

23. NON VIRAL VECTOR SYSTEM
1) PURE DNA CONSTRUCT
• Direct introduction of pure DNA construct
into target tissue.
• Efficiency of DNA uptake by the cells and
expression rather low.
• Consequently, large quantities of DNA
have to be injected periodically.

24. 2) LIPOPLEXES
• Lipid DNA complexes; DNA construct
surrounding by artificial lipid layer.
• Most of it gets degraded by lysosomes.
3) DNA MOLECULAR CONJUGATES
• Commonly used synthetic conjugate is
poly-L- lysin bound to specific target cell
receptor.

25. • Therapeutic DNA is then made to
combine with the conjugate to form a
complex.
• It avoids lysosomal breakdown of DNA.
4) HUMAN ARTIFICIAL CHROMOSOME
• Can carry a large DNA , with one or more
therapeutic genes with regulatory
elements.

26. METHODS OF GENE DELIVERY
PHYSICAL METHOD
GENE GUN
• Employs a high pressure delivery system
to shoot tissue with gold or tungsten
particles that are coated with DNA

27. MICROINJECTION
• Process of using a glass micropipette to
insert microscopic substances into a single
living cell.
• Normally performed under a specialized
optical microscope setup called a
micromanipulator.

28. SONOPORATION:
• We use ultrasonic frequency to deliver
DNA into cells . Which can disrupt the cells
membrane and allow DNA to move into
cells.

29. ELECTROPORATION:
• In this method we use short pulses of high
voltage to carry DNA across the cell membrane
which makes a shock to cause temporary
formation of pores and thus allow DNA
molecules to pass.

30. • Some of the drawbacks of electroporation
can be using of high- voltage plasma
discharge DNA was efficiently delivered
following very short pulses.

31. CHEMICAL METHODS
• USING DETERGENT MIXTURE
- Certain charged chemical compounds
like Calcium phosphates are mixed with
functioning cDNA of desired function.
- The mixture is introduced near the
vicinity of recipient cells.
- The chemicals disturbs the cell
membrane, widens the pore size and allows
cDNA to pass through the cell.

32. • LIPOFECTION
- It is a technique used to inject genetic
materials into a cell by means of liposomes.
- Liposomes are artificial phospholipid
vehicles used to deliver a variety of molecules
including DNA into the cells.
• INORGANIC NANOPARTICALS:
It is one of the non-viral gene delivery system
where we use gold-silica and iron oxide and
calcium phosphate some of the benefits are
storage stability , low manufacturing cost and
often time , low immunogenecity .

33. POTENTIAL TARGET DISEASES
Gene therapy can be used for a number of
diseases, such as Parkinson’s disease,
cancer Alzheimer's disease and cystic
fibrosis through a number of different
approaches.

34. Gene Therapy Used in Cancer
• Cancer is basically a disease of cells characterized by the
loss of-
-Normal cellular growth
-Maturation
-Multiplication
and thus Homeostasis is disturbed.
• Gene therapy for cancer is currently focused in multiple areas,
including :
1. Genetically engineered viruses that directly kill cancer cells,
2. Gene transfer to alter the abnormal functioning of cancer
cells, and
3. Immunotherapy (which includes CAR T-cell therapy), which
helps the immune system better find and kill tumour cells.

35. Genetically Engineered Viruses
• This approach uses specially modified
viruses (called oncolytic viruses) that
target and destroy cancer cells while
leaving normal cells unharmed.
• The viruses, engineered to contain certain
genes, are designed to infect cancer cells
and, once inside, to produce proteins that
cause the cells to die.

36. Gene Transfer
• In gene transfer, researchers introduce a foreign
gene directly into cancer cells or into
surrounding tissue.
• The goal is that the newly inserted gene will
cause the cancer cells to die or prevent cancer
cells and surrounding tissue from funnelling
blood to tumours, depriving them of nutrients
they need for survival.
• While this approach has a great deal of promise,
it presents scientists with several obstacles as
well, including “gene silencing,” in which the
implanted genes fail to switch on.

37. Immunotherapy
• CAR T-cell therapy, which seeks to enhance the natural
cancer-fighting ability of patient’s own T cells, is one type of
immunotherapy.
• A sample of a patient’s T cells is collected and mixed with
viruses carrying several specific genes.The viruses deliver
these genes to the T cell’s nuclei, where they are incorporated
into the cell’s DNA.
• The genes cause the T cells to express a special protein
called a chimeric antigen receptor, or CAR, on their surface.
The CAR directs the T cell to the tumour cell using a specific
“address,” and the CAR T cell is then equipped to rapidly
destroy the cancer cell.
• When the cells, now called CAR T cells, are infused into the
patient, they seek out tumour cells and then proliferate to
generate many more cancer-killing cells.

38. Gene Therapy Used in Cystic Fibrosis
• Cystic fibrosis is a genetic disease that causes mucus to
build up in a patient’s lungs. As a result, patients suffer
from blocked airways and bacterial infections.
• A set of industry collaborations could help bring a gene
therapy developed by the UK Cystic Fibrosis Gene
Therapy Consortium into clinical testing. The treatment
uses a type of virus called a lentivirus to deliver a healthy
copy of a gene called CFTR (Cystic Fibrosis
Transmembrane Regulator), which causes cystic fibrosis
when it carries a mutation. The gene therapy will be
given by inhalation to better target the right cells.

39. • Different mutations have been identified in the CFTR
gene that can cause cystic fibrosis, and gene therapy
may be the most effective way to combat all of them.
• UK Cystic Fibrosis Gene Therapy Consortium was the
first to show that repeated doses of a gene therapy
delivered in fat droplets could be effective in treating
cystic fibrosis in a Phase IIb trial, but the therapy was not
equally effective in all patients.
• Dutch biotech ProQR is developing a drug that binds to
mutated RNA of the CFTR gene to restore the
production of a healthy CFTR protein.

40. • London biotech Verona Pharma recently
obtained positive Phase II results for a
drug that simultaneously inhibits two
enzymes in order to reduce inflammation,
clear mucus membranes and dilate the
lungs.

41. Gene therapy Used in Parkinson's disease
• Parkinson's disease (PD) is a progressive neurological
condition that is the result of the death of the cell that
contains and produces dopamine in substantia nigra.
• People with PD may develop disturbance in their motor
activities.
• Gene therapy in Parkinson's disease consists of the
creation of new cells that produce a specific
neurotransmitter (dopamine), protect the neural system,
or the modification of genes that are related to the
disease. Then these cells are transplanted to a patient
with the disease.

42. Gene Therapy Used in Alzheimer Disease
• Alzheimer is a neurodegenerative disorder
in which there is loss of cholinergic
neurons at the basal forebrain.
• Treatment of this disease involves the
delivery of Nerve Growth Factor (NGF)
which prevents the neuronal loss in
addition to ameliorating deficits in learning
and memory associated cells.

43. • The intracerebral transplantation of genetically modified
fibroblasts has been applied to an Animal Model wherein
a sparingly small amount of the Cholinergic neurons has
been observed to produce NGF.
• Current pharmacological intervention consist on the
administration of L-dopa, a dopamine precursor. The L-
dopa therapy increases dopamine production of the
remaining nigral neurons.
• These treatments try to reduce the symptoms of the
patient focusing on increasing the production of
dopamine but they do not cure the disease.

44. • The new treatments for PD are in clinical trials
and most of them are centred on gene therapy.
• Researchers expect to compensate the loss of
dopamine or to protect the dopamine neurons
from degeneration. The pharmacological and
surgical therapies for PD focus on compensating
the ganglia dysfunction caused by the
degeneration of the dopaminergic neuron from
substantia nigra

45. ADVANTAGES
• Gene therapy has the potential to
eliminate and prevent hereditary disease
such as cystic fibrosis , ADS –SCID etc.
• It is possible cure for heart disease , AIDS
and cancer .
• It gives someone born with a genetic
disease a chance to life.
• It can be used to eradicate disease from
the future generations.

46. DISADVANTAGES
• Long testing therapy is not achieved by gene
therapy due to rapid dividing of cells benefits
of gene therapy is short lived .
• Immune response due the transferred gene
stimulate a potential risk to gene therapy.
• Disorders caused by defect in multiple genes
cannot be treated effectively using gene
therapy.
• Viruses used as vectors for gene transfer
may cause toxicity , immune responses , and
inflammatory reactions in the host.