Gene therapy is an 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 instead of using drugs or surgery. Gene therapy is used to correct defective genes in order to cure a disease or help your body better fight disease. Researchers are investigating several ways to do this, including: Replacing mutated genes. Some cells become diseased because certain genes work incorrectly or no longer work at all.

1. Amity Institute of Virology and Immunology
BY: -
SHIVANI TIWARI
A12173420002
PGD AIVI DEPARTMENT
TECHNIQUES TO ENGINEER
VIRAL GENOME AS A VECTOR FOR
GENE THERAPY

2. GENE THERAPY
An approach of treating diseasesby either modifying the expressions
of an individual’s genes or correction of abnormal genes
 This can be accomplished by:
• Replacing amutated gene that causes diseasewith ahealthy copy of
the gene.
• Inactivating,or“knockingout,”a mutated gene that is
functioning improperly.
• Introducing anewgeneinto the body to help fight adisease.
Amity Institute of Virology and Immunology

3. TYPES OF GENE THERAPY
Amity Institute of Virology and Immunology
SOMATICCELLGENETHERAPY GERMLINEGENETHERAPY
 Therapeutic genes do transferred into the
somatic cells.
 Therapeutic genes transferred into the germ
cells.
 E.g..Introduction of genes into bone
marrow cells, blood cells, skin cells etc.
 E.g..Genesintroduced into eggs and sperms.
 Will not be inherited later generations.  It is heritable and passed on to later
generations.
 At present all researches directed to correct
genetic defects in somatic cells.
 For safety, ethical and technical reasons, it is
not being attempted at Present.

4. TYPES OF SOMATIC GENE THERAPY
Amity Institute of Virology and Immunology
IN VIVO GENE THERAPY EX VIVO GENE THERAPY

5. EXAMPLE 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 .
Amity Institute of Virology and Immunology

6. EXAMPLE OF EX VIVO GENE THERAPY
 1st gene therapy – to correct deficiency ofenzyme,
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.
Amity Institute of Virology and Immunology

7. TECHNIQUES TO ENGINEER VIRAL GENOME AS A VECTOR FOR GENE
THERAPY
Technique by which we can transfer the desired gene into a target cell, a carrier is required.
Such vehicles of gene delivery are known as vectors.
Ideal Vector includes following characteristics: -
• TARGETthe right cells
• INTEGRA
TEthe gene in the cells.
• ACTIV
ATE the gene.
• AVOID harmful sideeffects.
 2 main classes of Vector are: -
 Viral vectors
 Non viral vectors
Amity Institute of Virology and Immunology

8. VIRAL VECTOR
 Viruses introduce their genetic material into the host cell as
part of their replication cycle.
 It remove the viral DNA and usingthevirus asavehicle to
deliver the therapeutic DNA.
 The viruses used are altered to makethem safe,although some
risks still exist with gene therapy.
Amity Institute of Virology and Immunology

9. RETRO-VIRUS VIRAL VECTOR SYSTEM
 The recombinant retroviruses have the
ability to integrate into the host genome
in a stable manner.
 Can carry a DNAof size – less than
3.4kb.
 Replication defective virus particles.
 Target cell - dividing
Amity Institute of Virology and Immunology

10. ADENO VIRUS VIRAL VECTOR SYSTEM
 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.
Amity Institute of Virology and Immunology

11. Amity Institute of Virology and Immunology
ADVANTAGES DISADVANTAGES
• Target specific types of cells. • They can cause immune responses in
patients, which leads to organ failure.
• They're very good at targeting and
entering cells
• They can carry alimited amount of
genetic material. Therefore, some genes
may be too big to fit into someviruses.
• They can be modified so that they can't
replicate and destroy cells.
• They target multiple cell type, which
leads to illness such as Cancer.

12. Method of Gene Delivery
 PHYSICAL METHODS
 GENEGUN
 Employs ahigh-pressure delivery system to shoot tissue with gold or tungsten particles that are coated with DNA.
 MICROINJECTION
 Process of using aglassmicropipette toinsert microscopic substancesinto a single living cell.
 Normally performed under a specialized optical microscope setup called amicromanipulator.
 CHEMICAL METHODS
 USING DETERGENT
 Certain charged chemical compounds like Calcium phosphates are mixed with functional cDNAof desired function.
 The mixture isintroduced near the vicinity o
f
recipient cells.
 The chemicals disturbs the cell membrane, widens the pore sizeand allows cDNAto pass through the cell.
 LIPOFECTION
 I t isatechnique usedto inject genetic materials into acell by meansof liposomes.
 Liposomes areartificial phospholipid vesiclesusedto deliver avariety of molecules including DNA into the cells.
Amity Institute of Virology and Immunology

13. Amity Institute of Virology and Immunology
ADVANTAGES DISADVANTAGES
 Gene therapy hasthe potential to eliminate and
prevent hereditary diseasessuch as cystic fibrosis,
ADA- SCIDetc.
 Long lasting therapy is not achieved by gene
therapy; Due to rapid dividing of cells benefits of
gene therapy isshort lived.
 I t is apossible cure for heart disease, AIDSand cancer.  Immune response to the transferred gene stimulates a
potential risk to gene therapy.
 I t gives someone born with agenetic diseaseachance
to life..
 Disorders caused by defects in multiple genes
cannot be treated effectively using gene therapy.
 I t can be used to eradicate diseasesfrom the future
generations
 Viruses used asvectors for gene transfer may cause
toxicity, immune responses, and inflammatory
reactions in the host.

14. CONCLUSION
• Theoretically, gene therapy is the permanent solution for geneticdiseases.
• But it hasseveral complexities. At its current stage, it is not accessible to most people due to its hugecost.
• Abreakthrough may come anytime and a
day may come when almost every disease will have agene therapy
• Genetherapy have the potential to revolutionize the practice of medicine.
Amity Institute of Virology and Immunology

15. THANK YOU