4. Conti…
Gene therapy is an experimental technique in
which introduction of normal genes into cells
in place of missing or defective ones in order
to correct genetic disorders.
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.
5. TYPES OF GENE THERAPY
Therapeutic genes
transferred into
the somatic cells.
Eg. Introduction of genes
into bone marrow cells,
blood cells, skin cells etc.
Will not be inherited
later generations.
At present all researches
directed to correct genetic
defects in somatic cells.
Therapeutic genes
transferred into the germ
cells.
Eg. Genes introduced into
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.
• Somatic cell gene therapy • Germ line gene therapy
8. EX VIVO GENE THERAPY
Transplant the modified cells to the patient.
Select genetically corrected cells and grow.
Introduce the therapeutic genes .
Grow the cells in culture
Isolate cells with genetic defect from a patient
9. 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 defect in gene coding for ADA.
Deoxy adenosine accumulate and destroys T lymphocytes.
Disrupts immunity , suffer from infectious diseases and die at young
age.
10. 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 numbers (e.g. brain cells).
In vivo gene transfer is necessary when cultured cells cannot be
re-implanted in patients effectively.
11. EXAMPLE OF IN VIVO GENE THERAPY
In patients with cystic fibrosis, a protein call 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 leads to the accumulation of sticky mucous in
respiratory tract and lungs.
Treated by in vivo replacement of defective gene by
adenovirus vector .
12. 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
Viral vectors
1) retrovirus vectors system: Can carry a DNA of
size – less than 3.4kb
13. Conti……
Retroviruses used in gene therapy so that any genes that are
harmful to man are removed. Corrective genes are then
added to replace the removed genes, and the new, modified
retrovirus is then introduced into the patient.
Adeno virus vector system
Gene therapy using an Adenovirus vector. A new gene is
inserted into an adenovirus vector, which is used introduce
the modified DNA into a human cell.If the treatment is
successful,the new gene will make a functional protein.
Lentivirus : It is a retrovirus it has a single stranded
RNA genome .used in haemophilia A , Rheumatoid
arthritis , Diabetes mellitus.
14. NON VIRAL VECTORS
1) pure DNA construct
Direct introduction of pure DNA construct into target tissue .
Therapeutic DNA is then made to combine with the conjugate to form a
complex.
It avoids lysosomal breakdown of DNA.
2) Lipoplexes
Lipid DNA complexes; DNA construct surrounded by artificial
lipid layer.
Most of it gets degraded by lysosomes
15. Gene Gun
Employs a high-pressure delivery system to
shoot tissue with gold or tungsten particles that
are coated with DNA
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.
METHODS OF GENE DELIVERY
PHYSICAL METHODS
16. Conti…
Sonoporation
Using ultrasonic frequency to deliver DNA into cell.
Allows disrupt cell membrane and DNA move into
cell.
Eletroporation
Method that uses short pulses of high voltage to carry
DNA across cell membrane. Thus cause temperature
formation of pore in cell membrane allows gene to
pass through.
17. CHEMICAL METHODS
USING DETERGENT MIXTURES
Certain charged chemical compounds like Calcium phosphates are
mixed with functional 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.
LIPOFECTION
It is a technique used to inject genetic materials into a cell by means of
liposomes.
Liposomes are artificial phospholipid vesicles used to deliver a
variety of molecules including DNA into the cells.
18. POLYPLEXES
Complexes of Cationic polymers + DNA
Consist of polymer,DNA binding proteins with ligands
attached to them
Polyethylinemine
Poly-L-Lysine Histone
Synthetic polypeptides
Polyplexes Structure:
Spherical
Globular
Rod-like
Toroids
19. OTHER TYPES OF GENE THERAPY
GENE AUGMENTATION THERAPY
Most common form of gene therapy
Foreign gene replaces missing or defective gene.
Eg. Replacement of defective gene p53 by a normal one in
liver cancer.
.
20. GENE INHIBITION THERAPY
Done to block the overproduction of some proteins.
2 types – Antigene and antisense therapy.
Antigene – blocks transcription using antigene oligonucleotide
Antisense – blocks transalation using antisense oligonucleotide
21. IS GENE THERAPY SAFE?
Gene therapy is under study to determine whether it could
be used to treat disease.
Several studies have already shown that this approach can
have very serious health risks, such as toxicity,
inflammation, and cancer.
some of the risks may be unpredictable; however, medical
researchers, institutions, and regulatory agencies are
working to ensure that gene therapy research is as safe as
possible.
The U.S. Food and Drug Administration (FDA) regulates all
gene therapy products in the United States and oversees
research in this area.
22. ADVANTAGES
Gene therapy has the potential to eliminate and prevent
hereditary diseases such as cystic fibrosis, ADA- SCID etc.
It is a 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 diseases from the future
generations.
23. DISADVATAGES
Long lasting therapy is not achieved by gene therapy; Due
to rapid dividing of cells benefits of gene therapy is short
lived.
Immune response to the transferred gene stimulates a
potential risk to gene therapy.
Viruses used as vectors for gene transfer may cause
toxicity, immune responses, and inflammatory reactions in
the host.
Disorders caused by defects in multiple genes cannot be
treated effectively using gene therapy.
24. APPLICATION OF GENE THERAPY
Gene Therapy Reduces parkinson’s disease symptoms
It significantly improved the weakness of the symptoms such as tremors motor
skill problem and rigidity
Main overactive brain region :the subthalamic nucleus should be introduced
with GAD gene
That would produce GABA Neurotransmitters. It is an inhibitor chemical then
they could potentially quiet . that brain region and alleviate tremors
Cystic fibrosis
Cystic fibrosis affects the epithelial cells of the body, but the life threatening
problems mainly affect the lungs
CF is caused by a mutation in the gene cystic fibrosis transmembrane
coductance regulator.CFTR gene found in chromosome 7. product f this gene
is CFTR protein .
The aim is to get the gene into the cells so that it can make the normal protein,
i.e. CFTR
25. Contd..
In Cancer
Multiple gene therapy have been developed to treat a wide variety of cancers
including suicide gene therapy, anti-angiogenesis and therapeutic gene
vaccines.
Gene therapy against cancer include the introduction of tumor suppressor
genes, gene that induces apoptosis or gene inhibitor tumor angiogenesis.
Hemophilia
For purpose with hemophilia therapies could be designed to deliver genes that
express missing factor viii and ix meaning individual would no longer need to
injected exogenous clotting factors
Fat metabolism disorder
In 2012 glybera became the first viral gene-therapy treatment to be approved in
Europe. Treatment uses in cure lipoprotein lipase deficiency.
26. Recent advances in Gene therapy
In march 2017 French scientists reported on clinical
research of gene therapy to treat sickle-cell disease.
In August 2017 , the FDA approved tisagenlecleucel for
acute lymphoblastic leukemia.
Tisagenlecleucel is an adoptive cell transfer therapy for B-
cell acute lymphoblastic leukemia.
T cells from a person with cancer are removed, genetically
engineered to make a specific T-cell receptor that reacts to
the cancer, and are administered back to the person.
27. Conti …
This is the first form of gene therapy to be approved in
the United States. In October, a similar therapy
called axicabtagene ciloleucel was approved
for lymphoma.
In a new gene therapy method developed by University
of Florida researchers found treatment for a
common form of blindness that strikes both
youngsters and adults.
28. ETHICAL QUESTION SURROUNDING
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?
Could the widespread use of gene therapy make the
society less accepting of people who are different?
Should people be allowed to use gene therapy to enhance
basic human traits such as height, intelligence, or athletic
ability?
29. CONCLUSION
Theoretically, gene therapy is the permanent solution for
genetic diseases.
But it has several complexities. At its current stage, it is not
accessible to most people due to its huge cost.
A breakthrough may come anytime and a day may come
when almost every disease will have a gene therapy
Gene therapy have the potential to revolutionize the
practice of medicine.
30. REFERENCES
Dubey R.C, A textbook of biotechnology, 1st edition(2004), S
Chand and company, New Delhi
Gupta P.K, Elements of Biotechnology, 1st edition(2001), Rastogi
Publications, Meerut.
Satyanarayana U, Biotechnology, 1st edition, Book and allied (P)
Ltd, Kolkata.
http://www.medindia.net/articles/genetherapy_treatment.htm
http://en.wikipedia.org/wiki/Gene_therapy
