gene therapy - arpita mishra

8/3/2019 Gene Therapy - Arpita Mishra

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Bio- Technology Project Work

Submitted By:Arpita Mishra

Class:- Xi th ARoll- 50

Subject Teacher:Miss Nabaneeta DasPgt Bio-Technology


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GENETHERAPY


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A Chromosome


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Gene therapy is the insertion, alteration, or removal of genes within anindividual's cells and biological tissues to treat disease.

It is a technique for correcting defective genes that are responsible for disease

development.

There are four approaches to gene therapy:

1. A normal gene inserted to compensate for a nonfunctional gene.

2. An abnormal gene traded for a normal gene.

3. An abnormal gene repaired through selective reverse mutation.

4. Change the regulation of gene pairs.

Although the technology is still in its infancy, it has been used with somesuccess.


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The First Approach

In the 1980s, Scientists began to look into genetherapy.They would insert human genes into a bacteria cell.Then the bacteria cell would transcribe and translatethe information into a proteinThen they would introduce the protein into human

cells.However today, most gene therapy studies are aimedat cancer and hereditary diseases linked to a geneticdefect.


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The First Time Ever

The first gene therapy was performed on September 14th,1990Ashanti DeSilva was treated for SCID(Sever combined

immunodeficiency ) .Doctors removed her white blood cells, inserted themissing gene into the WBC, and then put them back

into her blood stream.This strengthened her immune system.This only worked for a few months .


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How It Works

A vector delivers the therapeutic gene intoa patients target cell.The target cells become infected with theviral vector.The vectors genetic material is insertedinto the target cell.Functional proteins are created from thetherapeutic gene causing the cell to returnto a normal state.


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How it actually happens


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Types Of Gene Therapy

Germ line gene therapy :In the case of germ line genetherapy, Germ cells are modified by the introduction offunctional genes, which are integrated into their genomes.Therefore, the change due to therapy would be heritable and

would be passed on to later generations. Somatic gene therapy :In the case of somatic gene therapy,

the therapeutic genes are transferred into the somatic cells of

a patient. Any modifications and effects will be restricted tothe individual patient only, and will not be inherited by the

patient's offspring or later generations.


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Viruses: A Vector

These replicate by inserting their DNA into a host

cell.Gene therapy can use this to insert genes that

encode for a desired protein to create the desiredtrait.

Four different types:

- Retroviruses

- Adenoviruses

-Adeno-associated Viruses

-Herpes Simplex Viruses

R i


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Retroviruses

o The genetic material in retroviruses is in the form ofRNA molecules, while thegenetic material of their hosts is in the form ofDNA.

o

When a retrovirus infects a host cell, it will introduce itsRNA together withsome enzymes, namely reverse transcriptase and integrase, into the cell. The process of producing aDNA copy from anRNA molecule is

termed reverse transcription. Reverse transcriptase is the enzyme carried withthe virus ,through which this process is carried out.

Integrase is the enzyme carried in the virus which helps in the insertion ofDNA copy produced in the chromosomes.o Now the host cell has been modified to contain new genes. If this host cell

divides later, its descendants will all contain the new genes.o One of the problems is that Integrase inserts the gene anywhere randomly

because it has no specific site. .o If genetic material happens to be inserted in the middle of one of the original

genes of the host cell, this gene will be disrupted (insertional mutagenesis). Ifthe gene happens to be one regulating cell division, uncontrolled cell division

(i.e., cancer) can occur.

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Adenoviruses

Carries their genetic material in the form of double stranded DNAgenome that cause respiratory, intestinal, and eye infections inhumans. The inserted DNA is not incorporated into the host cell's geneticmaterial. The extra genes are not replicated when the cell is about to undergocell division so the descendants of that cell will not have the extragene. So it again has to be reinserted when more cells divide. This vector system has been promoted for treating cancer and indeedthe first gene therapy product to be licensed to treat cancer.
http://en.wikipedia.org/wiki/File:Gene_therapy.jpg


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http://en.wikipedia.org/wiki/File:Gene_therapy.jpg


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Adeno-associated Viruses

Adeno-associated Virus is small, has single strandedDNA that insert genetic material at a specific point onchromosome 19.

It belongs to parvovirus family. It causes no knowndisease and doesn't trigger patient immune response. It has got low information capacity The gene is always "on" so the protein is always beingexpressed, possibly even in instances when it isn'tneeded. It includes hemophilia treatments. For example, a gene-carrying vector could be injected into a muscle,prompting the muscle cells to produce Factor IX and thusprevent bleeding.

H S l V


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Herpes Simplex Viruses

The Herpes simplex virus is mostly examined for gene

transfer in the nervous system. The double stranded DNA

viruses that infect neurons.

For Example: Herpes simplex virus type 1.

Antibodies to HSV-1 are common in humans, howevercomplications due to herpes infection are somewhat rare.

N i l O i
http://www.ucmp.berkeley.edu/alllife/virus.gif


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Non-viral Options

Direct injection of DNA simplest method of non-viral transfection. Requires a lot of DNA. Cellular uptake is inefficient. Physical Methods to Enhance Delivery Electroporation : - uses short pulses of high voltage to carry DNA

across the cell membrane.- temporary formation of pores in the cell membrane is caused,

allowing DNA molecules to pass through.- efficient and works across a broad range of cell types ,but usage

has been limited.-More recently a newer method of electroporation, termed electron-

avalanche transfection, has been used in gene therapy experiments.

Gene Gun: -DNA is coated with gold particles and loaded into a devicewhich generates a force to achieve penetration of DNA/gold into thecells.

Sonoporation: -uses ultrasonic frequencies to deliver DNA into cells. Magnetofection: -DNA is complexed to magnetic particles, and a

magnet is placed underneath the tissue culture dish to bring DNAcomplexes into contact with a cell monolayer.

Ch i l M h d E h D li


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Chemical Methods to Enhance Delivery

1. Oligonucleotides: it inactivates the genes involved in thedisease process.

It includes several strategies like, use of antisense specific tothe target gene to disrupt the transcription of the faulty geneor use of small molecules of RNA to signal the cell to cleavespecific unique sequences in the mRNA transcript of thefaulty gene, disrupting translation of the faulty mRNA.

2. Lipoplexes and polyplexes: Its most common use has been ingene transfer into cancer cells.

lipoplexes are useful in transfecting respiratory epithelial cells,so they may be used for treatment of genetic respiratorydiseases such as cystic fibrosis.

Complexes of polymers with DNA are called polyplexes.

consist of cationic polymers and production is regulated byionic interactions.

Polyplexes cannot release their DNA load into the cytoplasm.

H b id h d


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Hybrid methods

Hybrid methods developed combined two or more techniques.Virosomes are one example; they combine liposomes with aninactivated HIV or influenza virus.

M j d l t i th


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Major developments in gene therapy

1972: Friedmann and Roblin authored a paper in Sciencetitled "Gene therapy for human genetic disease?

1990: The first approved gene therapy case in the United

States took place on September 14, 1990 on a four year

old girl named Ashanti DeSilva.

1992: Doctor Claudio Bordignon performed the firstprocedure of gene therapy using hematopoietic stem cells

as vectors to deliver genes intended to correct hereditary

diseases.

1999: Death of Jesse Gelsinger in a gene-therapy

experiment resulted in a significant setback to gene

therapy research in the United States.

2003 U f C l f h d h b


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2003 :University of California research team inserted genes into the brainusing liposomes coated in a polymer called polyethylene glycol. It is a

significant achievement because viral vectors are too big to get across the blood-

brain barrier. 2006 : Scientists at the National Institutes of Health have successfully treated

metastatic melanoma in two patients using killer T cells which demonstrated

that gene therapy can be effective in treating cancer.

In March ,an international group of scientists announced the successful use of

gene therapy to treat two adult patients for a disease affecting myeloid cells.

In May, a team of scientists led by Dr. Luigi Naldini and Dr. Brian Brown from

Telethon Institute for Gene Therapy (HSR-TIGET) developed a way to preventthe immune system from rejecting a newly delivered gene.


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In November ,Preston Nix from the University of Pennsylvania School ofMedicine reported, a gene-based immunotherapy for the treatmentof HIV that uses a lentiviral vector for delivery of an antisense geneagainst the HIV envelope. The insertion of T cells (geneticallymodified) was safe and well tolerated. 2007 :On 1 May, Moorfields Eye Hospital and Institute of

Ophthalmology in London, announced the world's first gene therapytrial for inherited retinal disease which was carried out on a 23 year-old British male. 2009 :In September, researchers at the University ofWashington and University of Florida were able to give trichromaticvision to squirrel monkeys using gene therapy, a hopeful precursor to a

treatment for color blindness in humans.

HIV


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2011: In 2007 and 2008, a man was cured of HIV by repeated Hematopoieticstem cell transplantation with double-delta-32 mutation this cure was not completelyaccepted by the medical community until 2011.

P bl d hi


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Problems and ethics

Short-lived nature of gene therapy

Immune response

Problems with viral vectors

Multigene disorders

Chance of inducing a tumor (insertional

mutagenesis)

Deaths have occurred due to gene therapy

Recent Developments


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Recent Developments

Genes get into brain using liposomes coated in polymer call polyethylene glycol.

Creation of tiny liposomes that can carry therapeutic DNA through pores of

nuclear membrane

Sickle cell successfully treated in mice


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Thank You for the patience and


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Thank You for the patience andcooperation

Special References:

WIKIPEDIA

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gene therapy - arpita mishra

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