GENE THERAPY

Agustina Setiawati, M.Sc., Apt

DEFINITION

1. Gene therapy is a technique for correcting “defective” genes responsible for disease development.

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CYSTIC FIBRIOSIS Cystic fibrosis (CF) is one of the most fatal heredity It is caused by more than 500 different mutations in the

cystic fibrosis transmembrane conductance regulator (CFTR) gene

A thick mucus which is a results of: Alignate produced by bacteria DNA from lysed cells Leucocytes which accumulate due to the infection

“ When genes are altered so that the encoded proteins are unable to carry out their normal functions, genetic disorders can result”

Several approaches of Gene Therapy

A normal gene may be inserted into a nonspecific location

An abnormal gene could be swapped The abnormal gene could be repaired through

selective reverse mutation

What is reverse mutation?

+-WT, active

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+

Mutate - to + Mutant, inactive

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WT, activeSame site revertant

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Mutate + to -

WT ? activedifferent site revertant

1. Gene therapy studies, a "normal" gene is inserted into the genome to replace an "abnormal," disease-causing gene.

2. A carrier molecule called a vector must be used to deliver the therapeutic gene to the patient's target cells.

3. The most common vector is a virus that has been genetically altered to carry normal human DNA

How Does Gene Therapy Work?

1. Enabling people to have children where natural conception is impossible (a more effective treatment of infertility).

2. The potential for discovering cures for terrible diseases - leading to less pain and suffering.

3. Sex selection to prevent genetic diseases.

4. Increased availability of organs for transplant.

5. Increased procreative autonomy (choice over some of the genetic characteristics that one's future child will possess).

Advantages of Gene Therapy

Disadvantages Gene Therapy

Safety concerns - the risk of detrimental genetic effects (bad biological consequences)

Dangers associated with 'genetic reductionism' - reducing human difficulties to genetic causes and ignoring their social context (EG, disability)

Other potential harms - e.g., the perpetuation of present inequalities/ stereotypes/ power relations, both within countries and between countries, could result,

Type of Human Gene Therapy

1. Somatic Gene TherapyIn vivo & Ex vivo

2. Germline Gene Therapy

SOMATIC GENE THERAPY“ Transfer material genetic to the body cells

except germline”

GERMLINE GENE THERAPY“ Involve genetic modification of germ cells”

Ex vivo manipulation techniques Electroporation Liposomes Gold bullets (fired within helium pressurized gun) Retrotransposons (jumping genes – early days)

In vivo techniques usually utilize viral vectors Virus = carrier of desired gene Virus is usually “crippled” to disable its ability to cause

disease Viral methods have proved to be the most efficient to

date Many viral vectors can stable integrate the desired

gene into the target cell’s genome

Problem: Replication defective viruses adversely affect the virus’ normal ability to spread genes in the body

Reliant on diffusion and spread Hampered by small intercellular spaces for transport Restricted by viral-binding ligands on cell surface therefore

cannot advance far.

VECTOR

An adequate carrying capacity To be undetectable by immune system Safe to patient High efficiency

VIRUS as VECTOR Retrovirus Adenovirus Adeno-associated virus Herpes simplex virus

RETROVIRUS

RNA virus, so it need reverse transcription and integration when enter to the cell

Only divide in rapid proliferation cell (hair, intestine )

Insert their material genetic in the middle of important gene

Response is reduced by immune response

group-specific antigen (gag) codes for core and structural proteins of the viruspolymerase (pol) codes for reverse transcriptase, protease and integraseenvelope (env) codes for the retroviral coat proteins.

ADENOVIRUS DNA virus Can infect slow proliferating cell, ie: blood,

lung, skin Material genetic can not be integrated to host

DNA Response is reduced by immune response

ADENO-ASSOCIATE VIRUS (AAV)

Occur naturally in human body DNA virus Harmless, non pathogenic, do not cause

immune response Insert their material genetic into chromosome

19 (spesific site) Only 2 genes can be inserted to these vector Trial to treat muscle and eye disease

HERPES SIMPLEKS

Non VIRAL VECTOR

Artificial liposome Human artificial chromosomes—introducing 47th

chromosome to the cell Chemical linking therapeutic DNA that can bind

to specific receptor of cell Cationic Liposome Naked DNA (electroporation, gene gun, DNA

injection)

Cationik liposome forms complex with DNA (negative charge)

Net charge of the complexes is positive so it can be interact to cell membrane

Human Artificial Chromosome

47th chromosome Microchromosome, has only 6- 10 Mb size Developed after bacterial and yeast artificial

chromosome Grown in HT1080 cell

Therapy Gene for Cancer

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DNA

mRNA

Protein

Transcription

Translation

AntisenseOligonucleotides

Any question?

THANK YOU