Gene therapy- Methods, Status and Limitations

Methods of gene delivery (therapeutic constructs)

It Includes two methods:

•Nonviral gene-delivery systems •Virus-Mediated gene-delivery systems

Non viral gene-delivery systems

• Naked DNA injection

• Gene gun

• Lipofection

Naked DNA gene therapy

•This is the simplest method i.e. the direct introduction of therapeutic DNA into target cells.

•Very cheap.

Microprojectile gene transfer (gene guns) therapy

Gene transfer through Liposomes

Therapeutic drugs

• Positively charged lipid droplets can interact with negatively charged DNA to wrap it up and deliver to cells.

• Inside liposomes DNA is resistant to degradation and is capable of passing the DNA through the target cell's membrane.

• Used in cystic fibrosis, cancer, parkinson’s disease.

Non viral gene-delivery systems

Advantages:• DNA can be of any size • Non-infectious and • Can’t replicate; no inflammatory response Disadvantages:

• low efficiency• non-specific

Virus-mediated gene-delivery systems

• Include Biological vehicles (vectors) such as viruses and bacteria.

• Viruses are currently the most efficient means of gene transfer.

• Viruses attack their hosts and introduce their genetic material containing genetic material into the host cell as part of their replication cycle.

Some viral vectors used in gene therapy are:

• Retroviruses

• Adenoviruses

• Adeno-associated viruses

• Herpes simplex virus

Single most optimal vector for all gene therapies

•No optimal gene therapy vector.•Different sites on the body requires different vectors different durations of treatment .•For example treatment of cancer requires short-term efficiency,

just enough to kill the cancer cells; whereas, treatment of Huntington's disease would likely

require life-long gene expression from a single injection.

Genetic Defects that are Candidates for Gene Therapy

• SCID caused by a number of defects: Example: ADA (adenosine deaminase) gene

defect. X – linked SCID.• Lack of functional lymphocytes.•No T-cell dependent antibody response.•Block the DNA Synthesis. •No cell mediated immune response.

Successful Gene Therapy for Severe Combine Immunodeficiency

SCID: Ex vivo gene therapy

The First Case

•The first gene therapy was performed on September 14th, 1990▫Ashanti DeSilva was treated for SCID

Sever combined immunodeficiency ▫Doctors removed her white blood cells,

inserted the missing gene into the WBC, and then put them back into her blood stream.

▫This strengthened her immune system▫Only worked for a few months

Gene Therapy Successes

Ashanti de Silva successfully treated for ADA deficiency - 1990

Ryes Evans successfully treated for SCID - 2001

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Current status in GT

•August 30,2006, Researchers at the National Cancer Institute (NCI), part of the National Institutes of Health, successfully reengineer immune cells, called lymphocytes, to target and attack cancer cells in patients with advanced metastatic melanoma.

• May 1, 2007): A team of British doctors from Moorfields Eye Hospital and University College in London conduct first human gene therapy trials to treat Leber's congenital amaurosis, a type of inherited childhood blindness caused by a single abnormal gene.

• 28 April 2008, UK researchers from the UCL

institute of Opthalmology and Moorfields Eye Hospital NIHR Biomedical research centre have announced results of World’s first gene therapy for inherited bilndness .

Current status in GT in India

•Very little gene therapy work being done in India

•Metro heart Institute in Noida is used the GT in production of vascular endothelial growth protein.

•Indian dept. of Bio-tech. has been given the permission to use the GT for treat renal cell carcinoma, colon, breast, & lung cancer by country’s regulators

Hurdles In Gene Therapy•Short lived nature of gene therapy

• Immune response

•Problems with viral vectors

• Insertional mutagenesis.

Major Problems that Scientists Must Overcome •Identify more efficient ways to deliver the

genes to the patients’ genetic material •Develop vectors that can specifically focus

on the targeted cells •Ensure that vectors will successfully

insert the desired genes into each of these target cells

•Deliver genes to a precise location in the patient’s DNA

•Ensure that transplanted genes are

precisely controlled by the body’s normal physiologic signals