GENE  THERAPY

•  It is  a technique  for  correcting  defective  gene  responsible  for  disease  development. It implies to correct the basic genetic abnormality.
• Gene therapy is a procedure involving inserting (or sometimes deleting) portions of gene in diseased patients for the purpose of cure.
• Antisense gene therapy or gene silencing selectively inactivate (or turn off) a mutated gene by RNA interference using short synthetic double stranded siRNA (small interfacing RNA).

Divided into
1. Somatic cell gene therapy-Gene is introduced into somatic cells.
2. Germ cell gene therapy (Transgenic animal).

several  approaches maybe  used:

1. Replacement:-  Mutant (abnormal)  gene  is  removed  and replaced  by  normal  gene.
2.  Augmentation:– Introduction  of foreign  gene  into  a cell to  compensate  for  the  defective  mutated gene.
3. correction:–  Pathological  change  in  the  nucleotide sequence  is  repaired.

Gene  of  interest is administered  in two vectors:-

i)   Viral vedors:- Retrovirus,adenovirus, adeno-associated virus, lentivirus.
ii)  Non-viral plasmid liposome, (Cationic-lipid) comptex

Transfer of genes into cells are :-

i) Site directed recombination
ii) Intranuclear microinjection
iii) Transduction by using viruses
iv) Transfection
v) Electroporation

Severe combined immunodeficiency disease (SCID) was the first disease treated successfully by gene therapy.

Methods of  Gene  Delivery

A- Chemical  methods:

DEAE-dextran:- Diethylaminoethyl-dextran (DEAE Dextran) is a poly cationic derivative of the carbohydrate polymer, dextran.

Calcium  Phosphate: Mixing DNA with calcium chloride, then adding this mixture to a phosphate buffered saline solution followed by incubation at room temperature.

Cationic  Lipids (Lipofection):- When liposomes were added to cells growing in vitro, some of the liposomes(lipids  mixed  with DNA in  water) would fuse with cellular plasma membranes and be taken up into the cells via endocytosis.

Polymers:  most popular polymer is the polycation, polyethylenimine (PEI). PEI is an organic macromolecule that possesses a high cationic charge density,(proton  spong

B- Physical  Methods 

Microinjection: It entails the direct iniection of DNA into the nuclei of target cells using fine glass needles under microscopy.

Electroporation: method of introducing nucleic acids into cells by exposing the cells to a rapid pulse of high-voltage current, causing pores in the cell membrane to open temporarily.

• the gene transfer efficiency is relatively low, and electroporation frequently results in a high incidence of cell death.

Gene  Gun:  Plasmid DNA is coated onto metal microparticles and then blasted into cells using either electrostatic force or gas pressure.

• This technique is fast, simple and safe, and it can transfer genes to a wide variety of tissues.

Gene therapy clinical trials addressed in:

1. Cancer (67%)-Most common
2. Vascular Disease (8.9%)
3. Monogenic Disorders (8.6 %)

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