Gene therapy is the use of DNS as a pharmaceutical agent to treat disease. It derives its name from the idea that DNA can be used to supplement or alter genes within an individual's cells as a therapy to treat disease. The most common form of gene therapy involves using DNA that encodes a functional, therapeutic gene to replace a mutated gene. Other forms involve directly correcting a mutation, or using DNA that encodes a therapeutic protein drug (rather than a natural human gene) to provide treatment. In gene therapy, DNA that encodes a therapeutic protein is packaged within a "vector", which is used to get the DNA inside cells within the body. Once inside, the DNA becomes expressed by the cell machinery, resulting in the production of therapeutic protein, which in turn treats the patient's disease.
Gene therapy using an Adenovirus vector. A new gene is inserted into a cell using an adenovirus. If the treatment is successful, the new gene will make functional protein to treat a disease.
Gene therapy is still in its infancy, with obvious challenges around targeting damaged cells and creating corrective genes. An equally important challenge, addressed by this research, is finding ways to transport the corrective genes into the cell. This is a problem, because of the poor permeability of cell membranes. There are several barriers to gene delivery, The genetic material must be protected during transit to a cell, it must pass into a cell, it must survive the cell's defense mechanisms, and it must enter into the cell's guarded nucleus.
If all of these barriers can be overcome, gene therapy would be a valuable technique with profound clinical implications. It has the potential to correct a number of human diseases that result from specific genes in a person's DNA makeup not functioning properly - or not at all. Gene therapy would provide a mechanism to replace these specific genes, swapping out the bad for the good. Safety is a primary concern when working with gene therapy. Some of the first attempts at gene therapy used viruses to insert DNA into cells. Viruses can be dangerously toxic, however, and this fact was tragically demonstrated a decade ago when an 18-year-old boy enrolled in a gene therapy study had a massive immune reaction to the viruses used. He died just a few days into the treatment from multiple organ failure, precipitating an immediate halt to the trial.
Several patients in gene therapy clinical trials have developed leukemia as a result of their treatment. The underlying cause of leukemia is thought to be that the viral vectors used to carry the therapeutic gene into cells (gamma-RVs) integrate into the genome of the cells disrupting the natural control of cancer-associated genes (a process known as insertional mutagenesis).
In gene therapy, the viruses are often equipped with additional genes, such as for immune mediators or for proteins inducing programmed cell death. However, these gene products can harm the body if they are released at the wrong moment or at excessive levels. It would be advantageous, if these switches could be turned on and off at a specific time. In order to construct such a switch, the researchers inserted synthetic segments of DNA into the viral genetic material in the direct vicinity of the transferred gene. In the infected cell, this construct is transcribed together with the transferred gene into a single messenger RNA (mRNA) molecule. The switch is operated using an agent which is added to cells infected with the virus. The substance is precisely fitted to bind to the RNA molecule and induces it to cut itself up. Thus, the potentially dangerous protein cannot be produced. The researchers copied this regulation mechanism from bacteria which use RNA switches to regulate production of numerous proteins.
Since then, many alternatives to viruses have emerged for use in gene therapy, including synthetic molecules like "dendrimers," a word that derives from the Greek word for "tree." Similar to trees, dendrimers are branching molecules that are slightly positively charged. This allows them to be loaded with DNA (which is slightly negative charged) for insertion into a cell. Dendrimers seem to offer many advantages over viruses. They may be much less toxic, and they may offer other advantages in terms of cost, ease of production, and the ability to transport very long genes. If they can be designed to efficiently -- and safely -- shuttle genes into human cells, then they may be a more practical solution to gene therapy than viruses. So far, laboratory experiments with different types of dendrimers have shown that they can insert genes into cells, but only with very low efficiency.
Using the principles of evolution and natural selection, that were initially conceived by Charles Darwin, they have now developped an efficient and safe gene delivery approach based on non-viral genetic elements, called transposons. Transposons are mobile DNA elements that can integrate into 'foreign' DNA via a 'cut-and-paste' mechanism. In a way they are natural gene delivery vehicles. The researchers constructed the transposons in such a way that they can carry the therapeutic gene into the target cell DNA. Doing so, they obviate the need to rely on viral vectors
This research describes a model peptide sequence, dubbed GeT (gene transporter), which wraps around genes, transports them through cell membranes and helps their escape from intracellular degradation traps. The process mimics the mechanisms viruses use to infect human cells. GeT was designed to undergo differential membrane-induced folding -- a process whereby the peptide changes its structure in response to only one type of membranes. This enables the peptide, and viruses, to carry genes into the cell. Interestingly, the property also makes it antibacterial and so capable of gene transfer even in bacteria-challenged environments. To prove the concept, the researchers used GeT to transfer a synthetic gene encoding for a green fluorescent protein -- a protein whose fluorescence in cells can be seen and monitored using fluorescence microscopy. The design can serve as a potential template for non-viral delivery systems and specialist treatments of genetic disorders.
To create the new gene therapy vector, Jans and colleagues used pieces of different genes to create a protein called a "modular DNA carrier," which can be produced by bacteria. This protein carries therapeutic DNA and delivers it to a cell's nucleus, where it reprograms a cell to function properly. In the laboratory, these carrier proteins were combined with therapeutic DNA and attached to cell membrane receptors and the nuclear import machinery of target cells. In turn, the packaged DNA moved into the cell through the cytoplasm and into the nucleus.
Genetically-engineered spider-silk proteins represent a versatile and useful new platform polymer for non viral gene delivery. The scientists describe modifying spider silk proteins so that they attach to diseased cells and not healthy cells. They also engineered the spider silk to contain a gene that codes for the protein that makes fireflies glow in order to provide a visual signal (seen using special equipment) that the gene has reached its intended target. In lab studies using mice containing human breast cancer cells, the spider-silk proteins attached to the cancer cells and injected the DNA material into the cells without harming the mice.
For the first time ever, chitosan nanoparticles have been used as a carrier for gene therapy in the ear. Chitosan is produced from shrimp shells. researchers attempted to use chitosan as a carrier to deliver drugs and genes to the inner ear in guinea pigs. Chitosan was able to deliver drugs through the membrane that covers the tiny gap between the middle ear and inner ear. Chitosan was also able to deliver genes to the hair cells. Whether or not the results from guinea pigs can be transferred to human ears remains uncertain. However, chitosan is non-toxic and is not harmful to cells. Chitosan is therefore better than other carriers and has characteristics that mean it could potentially be used with patients. Extremely small nanoparticles in the range of 50-200 nm (nanometres) are formed spontaneously when the positively charged chitosan and negatively charged genes are mixed. Chitosan does a good job packaging up DNA and RNA's relatively large molecules. When the nanoparticles have passed through a membrane, chitosan packages up the gene molecules so they return to their normal size again. Chitosan also creates gaps between cells, which facilitate the absorption of medicine.
Researchers from Johns Hopkins and Northwestern universities have discovered how to control the shape of nanoparticles that move DNA through the body and have shown that the shapes of these carriers may make a big difference in how well they work in treating cancer and other diseases. A major advance in this work is that Mao and his colleagues reported that they were able to "tune" these particles in three shapes, resembling rods, worms and spheres, which mimic the shapes and sizes of viral particles. "We could observe these shapes in the lab, but we did not fully understand why they assumed these shapes and how to control the process well," Mao said. These questions were important because the DNA delivery system he envisions may require specific, uniform shapes. The worm-shaped particles resulted in 1,600 times more gene expression in the liver cells than the other shapes," Mao said. This means that producing nanoparticles in this particular shape could be the more efficient way to deliver gene therapy to these cells. The particle shapes used in this research are formed by packaging the DNA with polymers and exposing them to various dilutions of an organic solvent. DNA's aversion to the solvent, with the help of the team's designed polymer, causes the nanoparticles to contract into a certain shape with a "shield" around the genetic material to protect it from being cleared by immune cells.
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Researchers from Johns Hopkins and Northwestern universities have discovered how to control the shape of nanoparticles that move DNA through the body and have shown that the shapes of these carriers may make a big difference in how well they work in treating cancer and other diseases. A major advance in this work is that Mao and his colleagues reported that they were able to "tune" these particles in three shapes, resembling rods, worms and spheres, which mimic the shapes and sizes of viral particles. "We could observe these shapes in the lab, but we did not fully understand why they assumed these shapes and how to control the process well," Mao said. These questions were important because the DNA delivery system he envisions may require specific, uniform shapes. The worm-shaped particles resulted in 1,600 times more gene expression in the liver cells than the other shapes," Mao said. This means that producing nanoparticles in this particular shape could be the more efficient way to deliver gene therapy to these cells. The particle shapes used in this research are formed by packaging the DNA with polymers and exposing them to various dilutions of an organic solvent. DNA's aversion to the solvent, with the help of the team's designed polymer, causes the nanoparticles to contract into a certain shape with a "shield" around the genetic material to protect it from being cleared by immune cells.
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- Wikipedia
- VIB (the Flanders Institute for Biotechnology) (2009, May 9). Moving Gene Therapy Forward With Mobile DNA. ScienceDaily. Retrieved January 13, 2013, from http://www.sciencedaily.com/releases/2009/05/090503132615.htm
- Federation of American Societies for Experimental Biology (2009, September 2). Finding The ZIP-code For Gene Therapy: Scientists Imitate Viruses To Deliver Therapeutic Genes. ScienceDaily. Retrieved January 13, 2013, from http://www.sciencedaily.com/releases/2009/08/090831130749.htm
- Helmholtz Association of German Research Centres (2012, August 21). Viruses with integrated gene switch. ScienceDaily. Retrieved January 13, 2013, from http://www.sciencedaily.com/releases/2012/08/120821114738.htm
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- American Institute of Physics (2009, April 30). First Large-scale Computer Simulation Of Gene Therapy. ScienceDaily. Retrieved January 13, 2013, from http://www.sciencedaily.com/releases/2009/04/090429152430.htm
- Northwestern University. "Shape matters in DNA nanoparticle therapy: Particles could become a safer, more effective delivery vehicle for gene therapy." ScienceDaily, 12 Oct. 2012. Web. 13 Jan. 2013.
