AN INVESTIGATORY PROJECT REPORT

ON
“GENE THERAPY”

NAME : S. HARINI
CLASS :12TH
SECTION : N2

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 “SAMRAKSHANA"
The Total Protection with Affection

ADITYA VIDYASHRAM RESIDENTIAL SCHOOL

Poraiyor, Villianur, Puducherry-605110
CBSE AISSCE XII 2023-2024
ALL INDIA SENIOR CERTIFICATE EXAMINATION
BIOLOGY PROJECT
CERTIFICATE

Name: __________________ Std & Sec: ______________

Registration Number:

Title of the project:

Certified that this is a bonafide project work done by the above-
mentioned student in our school during Senior Secondary course in
the year 2023-2024

SUBJECT TEACHERPRINCIPAL
Project Submitted for the CBSE-AISSCE Practical examination held
on__/__/ 2024.

INTERNAL EXAMINER EXTERNALEXAMINER

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 Acknowledgement

I would like to extend my sincere and heartfelt obligation towards all

those who have helped me in making this project. Without their active
guidance, help, cooperation and encouragement, I would not have been
able to present the project on time.

I am extremely thankful and pay my sincere gratitude to my teacher

____________________________for valuable guidance and support for

completion of this project.

I extend my sincere gratitude to my principal____________________

for the moral support extended during tenure of this project.

I also acknowledge with a deep sense of reverence, my gratitude towards

my parents, other faculty members of the school and friends for their
valuable suggestions given to me in completing the project.

Date:

Place: Signature of the Student

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 CERTIFICATE

This is to certify that the project work on

_______________________________________ based on the
curriculum of CBSE has been completed by
________________of Class-XII of Aditya Vidyashram
Residential school, Puducherry.

The above-mentioned project work has been completed

under my guidance during the academic year 2023-24.

Signature of the guide teacher

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 INDEX

1. Certificate
2. Acknowledgement
3. Introduction
4. Brief history of gene therapy
5. Types of gene therapy
6. Vectors of gene therapy
7. Methods of gene therapy
8. Success cases of gene therapy
9. Problems with gene therapy
10. Ethical issues
11. Conclusion
12. Bibliography

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INTRODUCTION:
What are genes?
 Genes are functional units of heredity as they are made of DNA.
 The chromosome is made of DNA containing many genes

Who defined gene?

 Danish botanist Wilhelm Johannsen coined the word gene
 1st discovery of gene by Gregor Mendel

Wilhelm Johannsen Gregor Mendel

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What gene therapy?

 It is a technique used to
correct the defective gene
that is responsible for
disease development.
There are four approaches
1. A normal gene inserted to
compensate the
dysfunctional gene.
2. An abnormal gene is traded for normal gene.
3. An abnormal gene is repaired through selective reverse
mutation.
4. Change the regulation of gene pair

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BRIEF HISTORY OF GENE THERAPY

1967- Nobelist Marshall Nirenberg wrote of programming cells.

1974- National Institute of Health took lead in recombinant DNA
(rDNA) research regulation.
1980-Dr. Martin Cline performs first DNA transfer into bone
marrow cells.
1984- US Office of Technology Assessment stressed difference
between somatic and germ-line therapy.
1990- NIH performed first approved gene therapy procedure.
1999- Jesse Gelsinger becomes first fatality in gene therapy.
2003- FDA placed a temporary halt on all gene therapy trials
using retroviral Evectors in blood stem cells.
2005-637 GT clinical trials (3464 patients)

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TYPES OF GENE THERAPY
Gene therapy is classified into two types:
Somatic gene therapy:
 Somatic cells of a patient are targeted for foreign gene
transfer.

 In this case the effects caused by the foreign gene is

restricted to the individual patient only, and not inherited by
the patient's offspring or later generation.
Germ line gene therapy:
 Here the functional genes, which are to be integrated into the
genomes, are inserted in the germ cells, i.e; sperm or eggs

 Trgeting of germ cells makes the thearpy heritable

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Types of Somatic Gene Therapy:

Ex Vivo
 Cells are modified the body
and then transplanted back
in again
 Called ex vivo because the
cells are treated outside the
body

In Vivo
 Genes are changed in cells
when the cells are still in
the body.
 Called in-vivo because the
gene is transferred to cells
inside the patient's body.

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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.

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Example of Ex-Vivo gene therapy:
 In ex vivo gene therapy, specific cells are removed from the
body and sent to a lab. In the lab, these cells are altered.

 Often, new DNA is added to the cells using a viral vector.

 Once the newly altered cells are prepared, they’re delivered

back to you, usually by an intravenous (IV) infusion.

 Ex vivo gene therapy is most often used in blood-related

disorders.

 This includes certain cancers — such as specific types of

leukemia and lymphoma — and genetic disorders.

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  The world's first gene therapy
trial, conducted by Dr French
Anderson and Michael Blaese. The
patient was a four-year-old girl,
named Ashanti DeSilva. She suffered
from ADA (adenosine deaminase
deficiency) and was given genetically
altered white blood cells to correct
potentially fatal abnormalities in her
immune system.

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In Vivo Gene Therapy:
 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 patientseffectively.

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Examples of In vivo gene therapy:
 In patients 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 leads to the accumulation of sticky mucous in

respiratory tract and lungs.

 Treated by in vivo replacement of defective gene by

adenovirus vector.

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Vectors in Gene Therapy:
Vectors are essentially vehicles designed to deliver
therapeutic genetic material, such as a working gene, directly
into a cell.
There are two types of vectors,

1. viral
2. non-viral

 Viral vectors are currently

a delivery vehicle used in FDA-
approved gene therapies.

 Non-viral techniques are

currently being studied as a safe
and effective way to deliver
genetic material to cells for
therapeutic effect.

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Ideal vectors:
 TARGET the right cells
 INTEGRATE the gene in the cells.
 ACTIVATE the gene.
 AVOID harmful side effects.
 No universal vector exists.

Viral vector:
 Viruses introduce their genetic material into the host cell as part
of their replication cycle.

 Remove the viral DNA and using the virus as a vehicle to deliver
the therapeutic DNA.

 The viruses used are altered to make them safe, although some
risks still exist with gene therapy.

Types of viral vector:

A number of viruses have been used for human gene therapy,
including:

1. Retrovirus
2. Adenovirus
3. Adeno-associated virus
4. Herpes simplex virus

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Retrovirus:
 A type of virus that has RNA instead of DNA as its genetic
material.
 It uses an enzyme called reverse transcriptase to become
part of the host cells' DNA.
 This allows many copies of the virus to be made in the host
cells.
 The recombinant retroviruses have the ability to integrate
into the host genome.
 Can carry a DNA of size - less than 3.4kb
 Target cell - dividing

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ACTIONS OF RETROVIRUS:

Lentivirus:
 Lentivirus is a genus of retroviruses that cause chronic and
deadly diseases characterized by long incubation periods, in
humans and other mammalian species.
 Subclass of Retroviruses.
 The viral genome in the form of RNA is reverse-transcribed
 When the virus enters the cell to produce DNA, which is then
inserted into the genome at a random position via viral
integrase enzyme.
 Target cells- dividing, non-dividing.

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Adenovirus:
 Adenoviral DNA does not integrate into the genome and is not
replicated during cell division.
 Humans commonly come in contact with adenovirus, majority of
patients have already developed neutralizing antibodies which can
inactivate the virus.
 Target- non dividing, dividing cells

Adeno-associated
Virus:
 It is a human virus.
 It is single stranded.
 AAV enters host cell, becomes
 double stranded and gets
integrated into chromosome.
 AAV is not currently known to
cause disease and consequently
the virus causes a very mild
immune response.
 Target- non dividing, dividing cells.
Herpes Simplex virus:
 Viruses which have natural tendency to
infect a particular type of cell.
 The Herpes simplex virus is a human
neurotropic virus.
 This is mostly examined for gene
transfer in the nervous system.

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Advantages and Disadvantages of using
viral vectors:
ADVANTAGES:
 Target specific types of cells.
 They're very good at targeting and
entering cells.
 They can be modified so that they can't
replicate and destroy cells.
DISADVANTAG:
 They can cause immune responses in patients
 They can carry a limited amount of genetic
material.
 Therefore, some genes may be too big to fit
into some viruses.
Non-Viral Vectors:
1. Pure DNA extract
 Direct introduction of pure DNA construct into target
tissue.
 Efficiency of DNA uptake by cells and expression rather
low.
 Consequently, large quantities of DNA have to be
injected periodically.

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 2. DNA molecular conjugates
 Commonly used synthetic conjugate is poly-L-lysine
bound to specific target cell receptor.
 Therapeutic DNA is then made to combine with the
conjugate to form a complex.
 It avoids lysosomal breakdown of DNA

3. Lipoplexes

 Lipid DNA complexes;

 DNA construct surrounded by artificial lipidlayer.
 Most of it gets degraded by lysosomes.

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4. Human artificial chromosome

 Can carry a large DNA i.e., with one or more therapeutic

genes.

Methods of Gene delivery

1. Physical methods
a. Gene Gun:
 Employs a high-pressure delivery system to shoot tissue with
gold or tungsten particles that are coated with DNA

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b. 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

2. Chemical method
A.Using detergent mixture
 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 disturb the cell membrane, widens the pore
size and allows cDNA to pass through the cell.

B. 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.
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 SUCCESS CASE OF GENE THERAPY

 After 50 years of research and 20 years of clinical

development, a clear benefit of gene therapy has been
demonstrated for 12 genetic diseases.

 In the future, genetic therapies may be used to prevent, treat,

or cure certain inherited disorders, such as cystic fibrosis,
alpha-1 antitrypsin deficiency, hemophilia, beta thalassemia,
and sickle cell disease. They also may be used to treat
cancers or infections, including HIV.

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Problems with gene therapy:

Short lived nature of gene therapy

 Patients will have to undergo multiple rounds of Gene
therapy
Immune response
 Stimulates the immune system that reduces gene therapy
effectiveness
Problems with viral vectors Potential
 Problems to the patient like toxicity, Immune and
inflammatory responses and gene control targeting are
created
Multi Gene disorder
 Disorders like heart diseases, high blood pressure, arthritis
and diabetes are caused by combined effects of many genes

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 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

 cells and tissues to treat chronic disease and genetic

disorders in which an impaired mutant gene is replaced with
a functional gene.

 There are many ways gene therapy works such as in viral

vectors e.g., Adeno-associated viruses.

 In experiments it has been proven to be a successful

treatment for diseases (like haemophilia in large animals).

 Like many treatments, it has its risks. Such as the potential

for infection or an immune system reaction as previously
stated.

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Bibliography

 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_treat
ment.htm

 http://en.wikipedia.o.g/wiki/Gene_therapy

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