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Gene Therapy-a novel

approach to treat diseases

By: Dhiraj kumar Bhuyan

What is gene therapy?
• Gene therapy is a technique for correcting
defective genes responsible for disease
• The basic definition of gene therapy is altering a
person’s genes in order to achieve a more desirable
• In most gene therapy studies, a "normal" gene is
inserted into the genome to replace an
"abnormal,“or disease-causing gene.
Aim Of Gene Therapy
• Is to introduce Therapeutic Material
into Target cells.
• Gene therapy typically aims to
supplement a defective mutant allele
with a functional one.
• At Present Gene Therapy is still at the
clinical research stage.
Successful gene therapy requires

1. Genetic nature of disease is understood.

2. The affected tissues are known and
3. Correct gene expression is achieved.
4. Harmful side effects if any are
Gene Therapy Principle



Retrovirus/Lentivirus Protein

Naked DNA
Types Of Gene Therapy
• Germ line gene therapy: Gene therapy can be
targeted to germ (egg and sperm) cells.

• Somatic cell gene therapy: Gene therapy can

be targeted to somatic (body cells).
- most common
three types
1. in vivo
2. ex vivo
3. Gene targeting
In Vivo Gene Therapy
• The genetic material is transferred directly
into the body of patient.
• Only available option for tissues that
cannot be grown in vitro; or if grown cells
cannot be transferred back.
• It is more or less a random process,
require less manipulations.
• Requires an efficient and selective delivery
Ex Vivo Gene Therapy
• The genetic material
is first transferred
into the cells grown
in vitro
• Controlled process,
transfected cells are
selected and
• more manipulations.
• Cells are usually
autologous, they are
then returned back
to the patient
in vivo and ex vivo schemes

Methods of gene delivery
(therapeutic constructs)

It Includes two methods:

• virus-mediated gene-delivery
• nonviral gene-delivery systems
Non viral gene-delivery

• Naked DNA injection

• Gene gun

• Electroporation

• Lipofection
Naked DNA gene therapy

• This is the simplest method i.e. the

direct introduction of therapeutic
DNA into target cells.
• Very cheap.
Limitations of naked DNA gene
 This approach is limited as it can

be used only with certain tissues

and requires large amount of
 Results in a prolonged low level
Microprojectile gene transfer (gene gun)

• Gene guns which shoots

DNA coated gold particles
into the cell using high
pressure gas.
• Fire particles in cells at
high speed so particles
enter cells.
• Invented for DNA transfer to
plant cells.
• Yet not available in human
Gene transfer through
•Positively charged lipid droplets can
interact with negatively charged DNA
to wrap it up and deliver to cells.

•Inside liposomes DNA is resistant to

degradation and is capable of passing
the DNA through the target cell's

•Used in cystic fibrosis, cancer,

Therapeutic drugs
parkinson’s disease.
Non viral gene-delivery
• DNA can be of any size
• Non-infectious and
• Can’t replicate; no inflammatory

• low efficiency
• non-specific
virus-mediated gene-delivery systems

• Include Biological vehicles (vectors) such as

viruses and bacteria.

• Viruses are currently the most efficient means of

gene transfer.

• Viruses attack their hosts and introduce their

genetic material into the host cell as part of their
replication cycle.
Some viral vectors used in gene
therapy are:

• Retroviruses
• Adenoviruses
• Adeno-associated viruses
• Herpes simplex virus
• Lentiviruses
Ideal Viral Characteristics
• Insert size: should be large
• Targeted: limited to a cell type.
• Immune response: none.
• Stable: not mutated.
• Production: easy to produce high
• Regulatable: produce enough protein to make
a difference.
Applications include a variety of

• Monogenic inherited disease,

• Cancer,
• Cardiovascular disease,
• Infectious disease,
• Autoimmune disease.
In humans

Cancer 69%
Genetic Defects that are
Candidates for Gene Therapy
Successful Gene Therapy for
Severe Combine Immunodeficiency

• SCID caused by a number of defects:

Example: ADA (adenosine deaminase) gene
X – linked SCID.

• Lack of functional lymphocytes.

• No T-cell dependent antibody response.
SCID: Ex vivo gene therapy
The First Case
• The first gene therapy was performed on
September 14th, 1990
– Ashanti DeSilva was treated for SCID
– Doctors removed her white blood cells,
inserted the missing gene into the WBC, and
then put them back into her blood stream.
– This strengthened her immune system
Gene Therapy Successes
Ashanti de Silva
Photo courtesy of Van de Silva

successfully treated for

ADA deficiency - 1990

Ryes Evans successfully

treated for SCID - 2001
Gene Therapy Problems

• Two boys treated for SCID developed

leukemia due to disruption of a gene that
regulates cell division.
•Extremely labour intensive.
•Repeated many times to achieve success.
Cancer gene therapy
Three ways

1. Construcing antisense of oncogene

2. Introducing correct functional version of tumor
suppresser gene
3. Introducing a gene encoding a toxic protein
Adenovirus p53
First commercially approved
gene therapy product
• For elevated milk production.

• Provide nutrition to milk

• Mice with a genetically engineered deficiency

which mimics some human disease.

• Methods for Tissue Specific Targeting .

Advantages of gene
• Treatment of a genetic disease at the root of the
problem, at the DNA level.
• Potential to treat a disease for which no treatment
is currently available.
• Potential for life-long treatment from a single
• Once a treatment for one genetic disease has been
developed, similar diseases should be equally
treatable, using a different disease-specific gene.
Hurdles In Gene Therapy
• Short lived nature of gene therapy
• Immune response
• Problems with viral vectors
• Multigene disorders
• Specific scientific knowledge and regulatory
expertise has to be available.
• Insertional mutagenesis.
Major Problems that Scientists
Must Overcome

• Develope more efficient ways to deliver the genes to the

patients’ genetic material
• Develop vectors that can specifically focus on the targeted cells
• Ensure that vectors will successfully insert the desired genes
into each of these target cells
• Deliver genes to a precise location in the patient’s DNA
• Ensure that transplanted genes are precisely controlled by the
body’s normal physiologic signals