Structural Biochemistry/DNA recombinant techniques/Gene Therapy

What is gene therapy?Edit

Gene therapy is an experimental technique that uses genes to treat or prevent diseases. Genes are specific sequences of bases that encode instructions on how to make proteins. When genes are altered so that the encoded proteins are unable to carry out their normal functions, genetic disorders can result. Gene therapy is used for correcting defective genes responsible for disease development. Researchers may use one of several approaches for correcting faulty genes. Although gene therapy is a promising treatment which helps successfully treat and prevent various diseases including inherited disorders, some types of cancer, and certain viral infections, it is still at experimental stage. Gene therapy is currently only being tested for the treatment of diseases that have no other cures.


There are many methods utilized to try and correct these altered genes:

• A normal gene can be inserted into a nonspecific location within the genome to replace a nonfunctional gene. This is the most common method.

• An abnormal gene could be swapped for a normal gene through homologous recombination.

• The abnormal gene could be repaired through selective reverse mutation, which returns the gene to its normal function.

• The regulation (act of turning the gene on or off) of a particular gene could be altered.

Using the method of inserting a normal gene to replace the disease-causing gene, a vector is used to deliver the new gene to the target cells. The most commonly used vector is a genetically altered virus that can carry normal human DNA. Target cells are then infected with the viral vector as the vector unload the genetic information into the target cell. This restores the target cell to the normal state.

EX VIVO method: cells are genetically-altered outside of the body and then reintroduced.

IN VIVO method: cells are genetically-altered inside the body after a genetically-altered vector containing therapeutic DNA is injected into the body.




In order to insert a “normal” gene to replace the abnormal/defective one, a carrier molecule called a vector must be used to deliver the new gene to the target cells. The most common vector used is a virus that has been genetically altered to carry normal human DNA. Because viruses have evolved a way of encapsulating and delivering their genes to human cells in a pathogenic manner, they make excellent vectors, provided they are manipulated to remove disease-causing genes and instead insert therapeutic genes. Target cells are infected with the viral vector and the vector unloads its genetic material containing the “normal” gene into the target cell. The main types of viruses used as gene therapy vectors include: Retroviruses, Adenoviruses, Adeno-associated viruses and Herpes simplex viruses. Retroviruses are a unique class of viruses that make double-stranded DNA copies of their RNA genomes which are integrated into the chromosomes of host cells. Adenoviruses is another class of viruses with the double-stranded DNA genome property that causes respiratory, intestinal, and eye inflections. Adeno-association viruses is a class of viruses that use single-stranded DNA to insert their genetic material at a specific sit. Herpes simplex viruses is a class of double-stranded DNA viruses that infect neurons.

There are also several non-viral options for gene delivery, the simplest being direct introduction of therapeutic DNA into target cells. However, this approach is limited in its application because it can be used only with certain tissues and requires large amounts of DNA. Another non-viral approach is the creation of an artificial lipid sphere with an aqueous core, (i.e. - a liposome), which is capable of carrying the DNA and then passing it through the target cell's membrane. Another way to get the DNA inside target cells is by chemically linking it to a molecule that will bind to special cell receptors. Once the molecule is bound to the receptors, the DNA is engulfed by the cell membrane and passed into the interior of the target cell. However, this delivery system tends to be less effective than other options.

Many of the challenges that make non-viral techniques ineffective is the challenge of getting genetic material past the plasma membrane without the help of viruses and avoiding the host's immune response. Methods using liposomes and plasmids are usually coupled with techniques to help facilitate the process of delivering genetic material past the plasma membrane. Electroporation is the use of electric shock from a high-voltage source to induce the formation of pores along the cell membrane to allow genetic material to enter. Sonoporation is similar, but uses acoustic sound waves to create these openings. While these techniques are used to make the delivery of genetic material by liposomes and plasmids more effective, they run the risk of causing cell death by rupturing the membrane.

In the future scientists hope to apply particle bombardment which has been successfully used to introduce genetic material in plants in the form of a gene gun. The concept would involve using small particles of gold along with liposomes/plasmids in "bullets" to punch pores in the cell membrane and deliver the genes to the interior. Although this is commonly used in the laboratory for research, it is a technique limited to be used ex vitro.

Short-comings of Gene Therapy:Edit

Short-lived nature of gene therapy – There are many problems with integrating the new DNA into the genome, the new DNA introduced into target cells must remain functional and the cells must live long and remain stable. However, rapidly dividing nature of many cells prevents gene therapy from achieving any long-term benefits; patients would have to undergo multiple rounds of gene therapy.

Immune response - Anytime a foreign object is introduced into human tissues, the immune system is designed to attack the invader, making it difficult to deliver the “normal” gene effectively. Furthermore, the immune system's enhanced response to invaders it has seen before makes it difficult for gene therapy to be repeated in patients.

Problems with viral vectors – While viral vectors are the carrier of choice in most gene therapy studies, they also present a variety of potential problems: toxicity, immune and inflammatory responses, gene control and targeting issues and the possibility that the virus may recover its ability to cause disease while inside the patient.

Multi-gene disorders- Conditions or disorders that arise from mutations in a single gene are the best/most effectively treated using gene therapy. Unfortunately, many of most common disorders, such as heart disease, high blood pressure, Alzheimer's disease, arthritis, and diabetes, are caused by the combined effects of variations in multiple genes. These disorders would be especially difficult to treat effectively using gene therapy.

However there have been many recent developments/advancements in gene therapy research, making it safer, and more effective, the field of gene therapy holds great potential for the future of medicine.