Gene therapy

Gene therapy

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Gene therapy is the insertion, alteration, or removal of gene
Gene
A gene is a molecular unit of heredity of a living organism. It is a name given to some stretches of DNA and RNA that code for a type of protein or for an RNA chain that has a function in the organism. Living beings depend on genes, as they specify all proteins and functional RNA chains...

s within an individual's cells
Cell (biology)
The cell is the basic structural and functional unit of all known living organisms. It is the smallest unit of life that is classified as a living thing, and is often called the building block of life. The Alberts text discusses how the "cellular building blocks" move to shape developing embryos....

 and biological tissue
Biological tissue
Tissue is a cellular organizational level intermediate between cells and a complete organism. A tissue is an ensemble of cells, not necessarily identical, but from the same origin, that together carry out a specific function. These are called tissues because of their identical functioning...

s to treat disease
Disease
A disease is an abnormal condition affecting the body of an organism. It is often construed to be a medical condition associated with specific symptoms and signs. It may be caused by external factors, such as infectious disease, or it may be caused by internal dysfunctions, such as autoimmune...

. It is a technique for correcting defective genes that are responsible for disease development. The most common form of gene therapy involves the insertion of functional genes into an unspecified genomic location in order to replace a mutated gene, but other forms involve directly correcting the mutation or modifying normal gene that enables a viral infection. Although the technology is still in its infancy, it has been used with some success.

Approach


Scientists have taken the logical step of trying to introduce genes directly into human cells, focusing on diseases caused by single-gene defects, such as cystic fibrosis
Cystic fibrosis
Cystic fibrosis is a recessive genetic disease affecting most critically the lungs, and also the pancreas, liver, and intestine...

, haemophilia
Haemophilia
Haemophilia is a group of hereditary genetic disorders that impair the body's ability to control blood clotting or coagulation, which is used to stop bleeding when a blood vessel is broken. Haemophilia A is the most common form of the disorder, present in about 1 in 5,000–10,000 male births...

, muscular dystrophy
Muscular dystrophy
Muscular dystrophy is a group of muscle diseases that weaken the musculoskeletal system and hamper locomotion. Muscular dystrophies are characterized by progressive skeletal muscle weakness, defects in muscle proteins, and the death of muscle cells and tissue.In the 1860s, descriptions of boys who...

 and sickle cell anemia. However, this has proven more difficult than modifying bacteria, primarily because of the problems involved in carrying large sections of DNA and delivering them to the correct site on the gene. Today, most gene therapy studies are aimed at cancer and hereditary diseases linked to a genetic defect. Antisense therapy
Antisense therapy
Antisense therapy is a form of treatment for genetic disorders or infections.When the genetic sequence of a particular gene is known to be causative of a particular disease, it is possible to synthesize a strand of nuc acid that will bind to the messenger RNA produced by that gene and inactivate...

 is not strictly a form of gene therapy, but is a related, genetically-mediated therapy.

The most common form of genetic engineering involves the insertion of a functional gene at an unspecified location in the host genome.This is accomplished by isolating and copying the gene of interest, generating a construct containing all the genetic elements for correct expression
Gene expression
Gene expression is the process by which information from a gene is used in the synthesis of a functional gene product. These products are often proteins, but in non-protein coding genes such as ribosomal RNA , transfer RNA or small nuclear RNA genes, the product is a functional RNA...

, and then inserting this construct into a random location in the host organism. Other forms of genetic engineering include gene targeting
Gene targeting
Gene targeting is a genetic technique that uses homologous recombination to change an endogenous gene. The method can be used to delete a gene, remove exons, add a gene, and introduce point mutations. Gene targeting can be permanent or conditional...

 and knocking out specific genes via engineered nucleases such as zinc finger nucleases, engineered I-CreI
I-CreI
I-CreI is a homing endonuclease whose gene was first discovered in the chloroplast genome of Chlamydomonas reinhardtii, a species of unicellular green algae. It is named for the facts that: it resides in an Intron; it was isolated from Clamydomonas reinhardtii; it was the first such gene isolated...

 homing endonucleases, or nucleases generated from TAL effector
TAL effector
TAL effectors are proteins secreted by Xanthomonas bacteria via their type III secretion system when they infect various plant species...

s. An example of gene-knockout mediated gene therapy is the knockout of the human CCR5
CCR5
C-C chemokine receptor type 5, also known as CCR5, is a protein that in humans is encoded by the CCR5 gene. CCR5 is a member of the beta chemokine receptors family of integral membrane proteins...

 gene in T-cells in order to control HIV
HIV
Human immunodeficiency virus is a lentivirus that causes acquired immunodeficiency syndrome , a condition in humans in which progressive failure of the immune system allows life-threatening opportunistic infections and cancers to thrive...

 infection. This approach is currently being used in several human clinical trials.

The biology of human gene therapy remains complex and many techniques need further development. Many diseases and their strict genetic link need to be understood more fully before gene therapy can be used appropriately. The public policy debate surrounding the possible use of genetically engineered material in human subjects has been equally complex. Major participants in the debate have come from the fields of biology, government, law, medicine, philosophy, politics, and religion, each bringing different views to the discussion.

Germ line gene therapy


In the case of germ line gene therapy, Germ cell
Germ cell
A germ cell is any biological cell that gives rise to the gametes of an organism that reproduces sexually. In many animals, the germ cells originate near the gut of an embryo and migrate to the developing gonads. There, they undergo cell division of two types, mitosis and meiosis, followed by...

s, i.e., sperm or eggs, are modified by the introduction of functional genes, which are integrated into their genomes. Therefore, the change due to therapy would be heritable and would be passed on to later generations. This new approach, theoretically, should be highly effective in counteracting genetic disorders and hereditary diseases. However, many jurisdictions prohibit this for application in human beings, at least for the present, for a variety of technical and ethical reasons.

Somatic gene therapy


In the case of somatic gene therapy, the therapeutic genes are transferred into the somatic cell
Somatic cell
A somatic cell is any biological cell forming the body of an organism; that is, in a multicellular organism, any cell other than a gamete, germ cell, gametocyte or undifferentiated stem cell...

s of a patient. Any modifications and effects will be restricted to the individual patient only, and will not be inherited by the patient's offspring or later generations.

Viruses



All virus
Virus
A virus is a small infectious agent that can replicate only inside the living cells of organisms. Viruses infect all types of organisms, from animals and plants to bacteria and archaea...

es bind to their hosts and introduce their genetic material into the host cell as part of their replication cycle. This genetic material contains basic 'instructions' of how to produce more copies of these viruses, hacking the body's normal production machinery to serve the needs of the virus. The host cell will carry out these instructions and produce additional copies of the virus, leading to more and more cells becoming infected. Some types of viruses insert their genes into the host's genome, but do not actually enter the cell. Others penetrate the cell membrane disguised as protein molecules and enter the cell.

There are two main types of virus infection: lytic
Lytic cycle
The lytic cycle is one of the two cycles of viral reproduction, the other being the lysogenic cycle. The lytic cycle is typically considered the main method of viral replication, since it results in the destruction of the infected cell...

 and lysogenic. Shortly after inserting its DNA, viruses of the lytic cycle quickly produce more viruses, burst from the cell and infect more cells. Lysogenic viruses integrate their DNA into the DNA of the host cell and may live in the body for many years before responding to a trigger. The virus reproduces as the cell does and does not inflict bodily harm until it is triggered. The trigger releases the DNA from that of the host and employs it to create new viruses.

Retroviruses


The genetic material in retrovirus
Retrovirus
A retrovirus is an RNA virus that is duplicated in a host cell using the reverse transcriptase enzyme to produce DNA from its RNA genome. The DNA is then incorporated into the host's genome by an integrase enzyme. The virus thereafter replicates as part of the host cell's DNA...

es is in the form of RNA
RNA
Ribonucleic acid , or RNA, is one of the three major macromolecules that are essential for all known forms of life....

 molecules, while the genetic material of their hosts is in the form of DNA. When a retrovirus infects a host cell, it will introduce its RNA together with some enzymes, namely reverse transcriptase and integrase, into the cell. This RNA molecule from the retrovirus must produce a DNA copy from its RNA molecule before it can be integrated into the genetic material of the host cell. The process of producing a DNA copy from an RNA molecule is termed reverse transcription. It is carried out by one of the enzymes carried in the virus, called reverse transcriptase
Reverse transcriptase
In the fields of molecular biology and biochemistry, a reverse transcriptase, also known as RNA-dependent DNA polymerase, is a DNA polymerase enzyme that transcribes single-stranded RNA into single-stranded DNA. It also helps in the formation of a double helix DNA once the RNA has been reverse...

. After this DNA copy is produced and is free in the nucleus
Cell nucleus
In cell biology, the nucleus is a membrane-enclosed organelle found in eukaryotic cells. It contains most of the cell's genetic material, organized as multiple long linear DNA molecules in complex with a large variety of proteins, such as histones, to form chromosomes. The genes within these...

 of the host cell, it must be incorporated into the genome of the host cell. That is, it must be inserted into the large DNA molecules in the cell (the chromosomes). This process is done by another enzyme carried in the virus called integrase
Integrase
Retroviral integrase is an enzyme produced by a retrovirus that enables its genetic material to be integrated into the DNA of the infected cell...

.

Now that the genetic material of the virus has been inserted, it can be said that the host cell has been modified to contain new genes. If this host cell divides later, its descendants will all contain the new genes. Sometimes the genes of the retrovirus do not express their information immediately.

One of the problems of gene therapy using retroviruses is that the integrase enzyme can insert the genetic material of the virus into any arbitrary position in the genome of the host; it randomly inserts the genetic material into a chromosome. If genetic material happens to be inserted in the middle of one of the original genes of the host cell, this gene will be disrupted (insertional mutagenesis
Insertional mutagenesis
Insertional mutagenesis is mutagenesis of DNA by the insertion of one or more bases.Insertional mutations can occur naturally, mediated by virus or transposon, or can be artificially created for research purposes in the lab.- Signature tagged mutagenesis :...

). If the gene happens to be one regulating cell division, uncontrolled cell division (i.e., cancer
Cancer
Cancer , known medically as a malignant neoplasm, is a large group of different diseases, all involving unregulated cell growth. In cancer, cells divide and grow uncontrollably, forming malignant tumors, and invade nearby parts of the body. The cancer may also spread to more distant parts of the...

) can occur. This problem has recently begun to be addressed by utilizing zinc finger nucleases or by including certain sequences such as the beta-globin locus control region to direct the site of integration to specific chromosomal sites.

Gene therapy trials using retroviral vectors to treat X-linked severe combined immunodeficiency
Severe combined immunodeficiency
Severe combined immunodeficiency , is a genetic disorder in which both "arms" of the adaptive immune system are impaired due to a defect in one of several possible genes. SCID is a severe form of heritable immunodeficiency...

 (X-SCID) represent the most successful application of gene therapy to date. More than twenty patients have been treated in France and Britain, with a high rate of immune system reconstitution observed. Similar trials were restricted or halted in the USA when leukemia was reported in patients treated in the French X-SCID gene therapy trial. To date, four children in the French trial and one in the British trial have developed leukemia as a result of insertional mutagenesis by the retroviral vector. All but one of these children responded well to conventional anti-leukemia treatment. Gene therapy trials to treat SCID due to deficiency of the Adenosine Deaminase (ADA
Adenosine deaminase
Adenosine deaminase is an enzyme involved in purine metabolism. It is needed for the breakdown of adenosine from food and for the turnover of nucleic acids in tissues.-Reactions:...

) enzyme (one form of SCID) continue with relative success in the USA, Britain, Ireland, Italy and Japan.

Adenoviruses


Adenoviruses are viruses that carry their genetic material in the form of double-stranded DNA. They cause respiratory, intestinal, and eye infections in humans (especially the common cold). When these viruses infect a host cell, they introduce their DNA molecule into the host. The genetic material of the adenoviruses is not incorporated (transient) into the host cell's genetic material. The DNA molecule is left free in the nucleus of the host cell, and the instructions in this extra DNA molecule are transcribed
Transcription (genetics)
Transcription is the process of creating a complementary RNA copy of a sequence of DNA. Both RNA and DNA are nucleic acids, which use base pairs of nucleotides as a complementary language that can be converted back and forth from DNA to RNA by the action of the correct enzymes...

 just like any other gene. The only difference is that these extra genes are not replicated when the cell is about to undergo cell division so the descendants of that cell will not have the extra gene.
As a result, treatment with the adenovirus will require readministration in a growing cell population although the absence of integration into the host cell's genome should prevent the type of cancer seen in the SCID trials. This vector system has been promoted for treating cancer and indeed the first gene therapy product to be licensed to treat cancer, Gendicine
Gendicine
Gendicine is a recombinant adenovirus engineered to express wildtype-p53 . This virus is designed to treat patients with tumors which have mutated p53 genes....

, is an adenovirus. Gendicine, an adenoviral p53-based
P53
p53 , is a tumor suppressor protein that in humans is encoded by the TP53 gene. p53 is crucial in multicellular organisms, where it regulates the cell cycle and, thus, functions as a tumor suppressor that is involved in preventing cancer...

 gene therapy was approved by the Chinese food and drug regulators in 2003 for treatment of head and neck cancer. Advexin, a similar gene therapy approach from Introgen, was turned down by the US Food and Drug Administration
Food and Drug Administration
The Food and Drug Administration is an agency of the United States Department of Health and Human Services, one of the United States federal executive departments...

 (FDA) in 2008.

Concerns about the safety of adenovirus vectors were raised after the 1999 death of Jesse Gelsinger
Jesse Gelsinger
Jesse Gelsinger was the first person publicly identified as having died in a clinical trial for gene therapy....

 while participating in a gene therapy trial. Since then, work using adenovirus vectors has focused on genetically crippled versions of the virus.

Envelope protein pseudotyping of viral vectors


The viral vectors described above have natural host cell populations that they infect most efficiently. Retroviruses have limited natural host cell ranges, and although adenovirus and adeno-associated virus
Adeno-associated virus
Adeno-associated virus is a small virus which infects humans and some other primate species. AAV is not currently known to cause disease and consequently the virus causes a very mild immune response. AAV can infect both dividing and non-dividing cells and may incorporate its genome into that of...

 are able to infect a relatively broader range of cells efficiently, some cell types are refractory to infection by these viruses as well. Attachment to and entry into a susceptible cell is mediated by the protein envelope on the surface of a virus. Retroviruses and adeno-associated viruses have a single protein coating their membrane, while adenoviruses are coated with both an envelope protein and fibers that extend away from the surface of the virus. The envelope proteins on each of these viruses bind to cell-surface molecules such as heparin sulfate, which localizes them upon the surface of the potential host, as well as with the specific protein receptor that either induces entry-promoting structural changes in the viral protein, or localizes the virus in endosomes wherein acidification of the lumen
Lumen (anatomy)
A lumen in biology is the inside space of a tubular structure, such as an artery or intestine...

 induces this refolding of the viral coat. In either case, entry into potential host cells requires a favorable interaction between a protein on the surface of the virus and a protein on the surface of the cell.
For the purposes of gene therapy, one might either want to limit or expand the range of cells susceptible to transduction by a gene therapy vector. To this end, many vectors have been developed in which the endogenous viral envelope proteins have been replaced by either envelope proteins from other viruses, or by chimeric proteins. Such chimera
Chimera (virus)
A chimera virus is defined by the Center for Veterinary Biologics as a "new hybrid microorganism created by joining nucleic acid fragments from two or more different microorganisms in which each of at least two of the fragments contain essential genes necessary for replication." The term chimera...

 would consist of those parts of the viral protein necessary for incorporation into the virion as well as sequences meant to interact with specific host cell proteins. Viruses in which the envelope proteins have been replaced as described are referred to as pseudotyped viruses. For example, the most popular retroviral vector for use in gene therapy trials has been the lentivirus
Lentivirus
Lentivirus is a genus of slow viruses of the Retroviridae family, characterized by a long incubation period...

 Simian immunodeficiency virus
Simian immunodeficiency virus
Simian immunodeficiency virus , also known as African Green Monkey virus and also as Monkey AIDS is a retrovirus able to infect at least 33 species of African primates...

 coated with the envelope proteins, G-protein, from Vesicular stomatitis virus
Vesicular stomatitis virus
Vesicular stomatitis Indiana virus is a virus in the family Rhabdoviridae; the well-known Rabies virus belongs to the same family. VSIV can infect insects, cattle, horses, pigs and humans. It has particular importance to farmers in certain regions of the world where it can infect cattle...

. This vector is referred to as VSV G-pseudotyped lentivirus, and infects an almost universal set of cells. This tropism is characteristic of the VSV G-protein with which this vector is coated. Many attempts have been made to limit the tropism of viral vectors to one or a few host cell populations. This advance would allow for the systemic administration of a relatively small amount of vector. The potential for off-target cell modification would be limited, and many concerns from the medical community would be alleviated. Most attempts to limit tropism have used chimeric envelope proteins bearing antibody
Antibody
An antibody, also known as an immunoglobulin, is a large Y-shaped protein used by the immune system to identify and neutralize foreign objects such as bacteria and viruses. The antibody recognizes a unique part of the foreign target, termed an antigen...

 fragments. These vectors show great promise for the development of "magic bullet" gene therapies.

Replication-Competent Vectors


A replication-competent vector called ONYX-015 is used in replicating tumor cells. It was found that in the absence of the E1B-55Kd viral protein, adenovirus caused very rapid apoptosis of infected, p53(+) cells, and this results in dramatically reduced virus progeny and no subsequent spread. Apoptosis was mainly the result of the ability of EIA to inactivate p300. In p53(-) cells, deletion of E1B 55kd has no consequence in terms of apoptosis, and viral replication is similar to that of wild-type virus, resulting in massive killing of cells.

A replication-defective vector deletes some essential genes. These deleted genes are still necessary in the body so they are replaced with either a helper virus or a DNA molecule.

Cis and trans-acting elements


Replication-defective vectors always contain a “transfer construct”. The transfer construct carries the gene to be transduced or “transgene”. The transfer construct also carries the sequences which are necessary for the general functioning of the viral genome: packaging sequence, repeats for replication and, when needed, priming of reverse transcription. These are denominated cis-acting elements, because they need to be on the same piece of DNA as the viral genome and the gene of interest. Trans-acting elements are viral elements, which can be encoded on a different DNA molecule. For example, the viral structural proteins can be expressed from a different genetic element than the viral genome.

Herpes Simplex Virus


The Herpes simplex virus
Herpes simplex virus
Herpes simplex virus 1 and 2 , also known as Human herpes virus 1 and 2 , are two members of the herpes virus family, Herpesviridae, that infect humans. Both HSV-1 and HSV-2 are ubiquitous and contagious...

 is a human neurotropic virus. This is mostly examined for gene transfer in the nervous system. The wild type HSV-1 virus is able to infect neurons. Infected neurons are not rejected by the immune system. Though the latent virus is not transcriptionally apparent, it does possess neuron specific promoters that can continue to function normally. Antibodies to HSV-1 are common in humans, however complications due to herpes infection are somewhat rare.

Non-viral methods


Non-viral methods present certain advantages over viral methods, with simple large scale production and low host immunogenicity being just two. Previously, low levels of transfection
Transfection
Transfection is the process of deliberately introducing nucleic acids into cells. The term is used notably for non-viral methods in eukaryotic cells...

 and expression of the gene
Gene expression
Gene expression is the process by which information from a gene is used in the synthesis of a functional gene product. These products are often proteins, but in non-protein coding genes such as ribosomal RNA , transfer RNA or small nuclear RNA genes, the product is a functional RNA...

 held non-viral methods at a disadvantage; however, recent advances in vector technology have yielded molecules and techniques with transfection efficiencies similar to those of viruses.

Injection of Naked DNA


This is the simplest method of non-viral transfection. Clinical trials carried out of intramuscular injection of a naked DNA
Naked DNA
Naked DNA is histone-free DNA that is passed from cell to cell during a gene transfer process called transformation or transfection. In transformation , purified or naked DNA is taken up by the recipient cell which will give the recipient cell a new characteristic or phenotype...

 plasmid have occurred with some success; however, the expression has been very low in comparison to other methods of transfection. In addition to trials with plasmids, there have been trials with naked PCR
Polymerase chain reaction
The polymerase chain reaction is a scientific technique in molecular biology to amplify a single or a few copies of a piece of DNA across several orders of magnitude, generating thousands to millions of copies of a particular DNA sequence....

 product, which have had similar or greater success. Cellular uptake of naked DNA is generally inefficient. Research efforts focusing on improving the efficiency of naked DNA uptake have yielded several novel methods, such as electroporation
Electroporation
Electroporation, or electropermeabilization, is a significant increase in the electrical conductivity and permeability of the cell plasma membrane caused by an externally applied electrical field...

, sonoporation
Sonoporation
Sonoporation, or cellular sonication, is the use of sound for modifying the permeability of the cell plasma membrane. This technique is usually used in molecular biology and non-viral gene therapy in order to allow uptake of large molecules such as DNA into the cell, in a cell disruption process...

, and the use of a "gene gun"
Gene gun
A gene gun or a biolistic particle delivery system, originally designed for plant transformation, is a device for injecting cells with genetic information. The payload is an elemental particle of a heavy metal coated with plasmid DNA...

, which shoots DNA coated gold particles into the cell using high pressure gas.
Electroporation

Electroporation
Electroporation
Electroporation, or electropermeabilization, is a significant increase in the electrical conductivity and permeability of the cell plasma membrane caused by an externally applied electrical field...

 is a method that uses short pulses of high voltage to carry DNA across the cell membrane. This shock is thought to cause temporary formation of pores in the cell membrane, allowing DNA molecules to pass through. Electroporation is generally efficient and works across a broad range of cell types. However, a high rate of cell death following electroporation has limited its use, including clinical applications.

More recently a newer method of electroporation, termed electron-avalanche transfection, has been used in gene therapy experiments. By using a high-voltage plasma discharge, DNA was efficiently delivered following very short (microsecond) pulses. Compared to electroporation, the technique resulted in greatly increased efficiency and less cellular damage.
Gene Gun

The use of particle bombardment, or the gene gun
Gene gun
A gene gun or a biolistic particle delivery system, originally designed for plant transformation, is a device for injecting cells with genetic information. The payload is an elemental particle of a heavy metal coated with plasmid DNA...

, is another physical method of DNA transfection. In this technique, DNA is coated with gold particles and loaded into a device which generates a force to achieve penetration of DNA/gold into the cells.
Sonoporation

Sonoporation
Sonoporation
Sonoporation, or cellular sonication, is the use of sound for modifying the permeability of the cell plasma membrane. This technique is usually used in molecular biology and non-viral gene therapy in order to allow uptake of large molecules such as DNA into the cell, in a cell disruption process...

 uses ultrasonic frequencies to deliver DNA into cells. The process of acoustic cavitation is thought to disrupt the cell membrane and allow DNA to move into cells.
Magnetofection

In a method termed magnetofection
Magnetofection
Magnetofection is a simple and highly efficient transfection method that uses magnetic fields to concentrate particles containing nucleic acid into the target cells...

, DNA is complexed to magnetic particles, and a magnet is placed underneath the tissue culture dish to bring DNA complexes into contact with a cell monolayer.
Oligonucleotides

The use of synthetic oligonucleotides in gene therapy is to deactivate the genes involved in the disease process. There are several methods by which this is achieved. One strategy uses antisense specific to the target gene to disrupt the transcription of the faulty gene. Another uses small molecules of RNA called siRNA
Sírna
Sírna Sáeglach , son of Dian mac Demal, son of Demal mac Rothechtaid, son of Rothechtaid mac Main, was, according to medieval Irish legend and historical tradition, a High King of Ireland...

 to signal the cell to cleave specific unique sequences in the mRNA transcript of the faulty gene, disrupting translation of the faulty mRNA, and therefore expression of the gene. A further strategy uses double stranded oligodeoxynucleotides as a decoy for the transcription factors that are required to activate the transcription of the target gene. The transcription factors bind to the decoys instead of the promoter of the faulty gene, which reduces the transcription of the target gene, lowering expression. Additionally, single stranded DNA oligonucleotides have been used to direct a single base change within a mutant gene. The oligonucleotide is designed to anneal with complementarity to the target gene with the exception of a central base, the target base, which serves as the template base for repair. This technique is referred to as oligonucleotide mediated gene repair, targeted gene repair, or targeted nucleotide alteration.
Lipoplexes and polyplexes

To improve the delivery of the new DNA into the cell, the DNA must be protected from damage and (positively charged). Initially, anionic and neutral lipids were used for the construction of lipoplexes for synthetic vectors. However, in spite of the facts that there is little toxicity associated with them, that they are compatible with body fluids and that there was a possibility of adapting them to be tissue specific; they are complicated and time consuming to produce so attention was turned to the cationic versions.

Cationic lipids
Cationic liposome
- life :Cationic liposomes are structures that are made of positively charged lipids and are increasingly being researched for use in gene therapy due to their favourable interactions with negatively charged DNA and cell membranes. Cationic liposomes are also known as cationic lipoplexes....

, due to their positive charge, were first used to condense negatively charged DNA molecules so as to facilitate the encapsulation of DNA into liposomes. Later it was found that the use of cationic lipids significantly enhanced the stability of lipoplexes. Also as a result of their charge, cationic liposomes interact with the cell membrane, endocytosis
Endocytosis
Endocytosis is a process by which cells absorb molecules by engulfing them. It is used by all cells of the body because most substances important to them are large polar molecules that cannot pass through the hydrophobic plasma or cell membrane...

 was widely believed as the major route by which cells uptake lipoplexes. Endosomes are formed as the results of endocytosis, however, if genes can not be released into cytoplasm by breaking the membrane of endosome, they will be sent to lysosomes where all DNA will be destroyed before they could achieve their functions. It was also found that although cationic lipids themselves could condense and encapsulate DNA into liposomes, the transfection efficiency is very low due to the lack of ability in terms of “endosomal escaping”. However, when helper lipids (usually electroneutral lipids, such as DOPE) were added to form lipoplexes, much higher transfection efficiency was observed. Later on, it was figured out that certain lipids have the ability to destabilize endosomal membranes so as to facilitate the escape of DNA from endosome, therefore those lipids are called fusogenic lipids. Although cationic liposomes have been widely used as an alternative for gene delivery vectors, a dose dependent toxicity of cationic lipids were also observed which could limit their therapeutic usages.

The most common use of lipoplexes has been in gene transfer into cancer cells, where the supplied genes have activated tumor suppressor control genes in the cell and decrease the activity of oncogenes. Recent studies have shown lipoplexes to be useful in transfecting respiratory epithelial cells
Epithelium
Epithelium is one of the four basic types of animal tissue, along with connective tissue, muscle tissue and nervous tissue. Epithelial tissues line the cavities and surfaces of structures throughout the body, and also form many glands. Functions of epithelial cells include secretion, selective...

, so they may be used for treatment of genetic respiratory diseases such as cystic fibrosis.

Complexes of polymers with DNA are called polyplexes. Most polyplexes consist of cationic polymers and their production is regulated by ionic interactions. One large difference between the methods of action of polyplexes and lipoplexes is that polyplexes cannot release their DNA load into the cytoplasm, so to this end, co-transfection with endosome-lytic agents (to lyse the endosome that is made during endocytosis, the process by which the polyplex enters the cell) such as inactivated adenovirus must occur. However, this isn't always the case, polymers such as polyethylenimine
Polyethylenimine
Linear polyethyleneimines contain all secondary amines, in contrast to branched PEIs which contain primary, secondary and tertiary amino groups. The linear PEIs are solids at room temperature where branched PEIs are liquids at all molecular weights. Linear polyethyleneimines are soluble in hot...

 have their own method of endosome disruption as does chitosan
Chitosan
Chitosan is a linear polysaccharide composed of randomly distributed β--linked D-glucosamine and N-acetyl-D-glucosamine...

 and trimethylchitosan.
Dendrimers

A dendrimer
Dendrimer
Dendrimers are repetitively branched molecules. The name comes from the Greek word "δένδρον" , which translates to "tree". Synonymous terms for dendrimer include arborols and cascade molecules. However, dendrimer is currently the internationally accepted term. A dendrimer is typically symmetric...

 is a highly branched macromolecule
Macromolecule
A macromolecule is a very large molecule commonly created by some form of polymerization. In biochemistry, the term is applied to the four conventional biopolymers , as well as non-polymeric molecules with large molecular mass such as macrocycles...

 with a spherical shape. The surface of the particle may be functionalized in many ways and many of the properties of the resulting construct are determined by its surface.

In particular it is possible to construct a cationic dendrimer, i.e. one with a positive surface charge. When in the presence of genetic material such as DNA or RNA, charge complimentarity leads to a temporary association of the nucleic acid with the cationic dendrimer. On reaching its destination the dendrimer-nucleic acid complex is then taken into the cell via endocytosis.

In recent years the benchmark for transfection agents has been cationic lipids. Limitations of these competing reagents have been reported to include: the lack of ability to transfect a number of cell types, the lack of robust active targeting capabilities, incompatibility with animal models, and toxicity. Dendrimers offer robust covalent construction and extreme control over molecule structure, and therefore size. Together these give compelling advantages compared to existing approaches.

Producing dendrimers has historically been a slow and expensive process consisting of numerous slow reactions, an obstacle that severely curtailed their commercial development. The Michigan based company Dendritic Nanotechnologies discovered a method to produce dendrimers using kinetically driven chemistry, a process that not only reduced cost by a magnitude of three, but also cut reaction time from over a month to several days. These new "Priostar" dendrimers can be specifically constructed to carry a DNA or RNA payload that transfects cells at a high efficiency with little or no toxicity.
Inorganic Nanoparticles

Inorganic nanoparticles, such as gold
Gold
Gold is a chemical element with the symbol Au and an atomic number of 79. Gold is a dense, soft, shiny, malleable and ductile metal. Pure gold has a bright yellow color and luster traditionally considered attractive, which it maintains without oxidizing in air or water. Chemically, gold is a...

, silica, iron oxide (ex. magnetofection
Magnetofection
Magnetofection is a simple and highly efficient transfection method that uses magnetic fields to concentrate particles containing nucleic acid into the target cells...

) and calcium phosphates have been shown to be capable of gene delivery. Some of the benefits of inorganic vectors is in their storage stability, low manufacturing cost and often time, low immunogenicity, and resistance to microbial attack. Nanosized materials less than 100nm have been shown to efficiently trap the DNA
DNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...

 or RNA
RNA
Ribonucleic acid , or RNA, is one of the three major macromolecules that are essential for all known forms of life....

 and allows its escape from the endosome
Endosome
In biology, an endosome is a membrane-bound compartment inside eukaryotic cells. It is a compartment of the endocytic membrane transport pathway from the plasma membrane to the lysosome. Molecules internalized from the plasma membrane can follow this pathway all the way to lysosomes for...

 without degradation. Inorganics have also been shown to exhibit improved in vitro transfection
Transfection
Transfection is the process of deliberately introducing nucleic acids into cells. The term is used notably for non-viral methods in eukaryotic cells...

 for attached cell lines due to their increased density and preferential location on the base of the culture dish. Quantum dots have also been used successfully and permits the coupling of gene therapy with a stable fluorescence marker.

Hybrid methods


Due to every method of gene transfer having shortcomings, there have been some hybrid methods developed that combine two or more techniques. Virosomes are one example; they combine liposomes with an inactivated HIV
HIV
Human immunodeficiency virus is a lentivirus that causes acquired immunodeficiency syndrome , a condition in humans in which progressive failure of the immune system allows life-threatening opportunistic infections and cancers to thrive...

 or influenza virus. This has been shown to have more efficient gene transfer in respiratory epithelial cells than either viral or liposomal methods alone. Other methods involve mixing other viral vectors with cationic lipids or hybridising viruses.

1970s and earlier


In 1972 Friedmann and Roblin authored a paper in Science titled "Gene therapy for human genetic disease?" They cite Rogers S for proposing "that exogenous 'good'" DNA be used to replace the defective DNA in those who suffer from genetic defects. They also cite the first attempt to perform gene therapy as [New York Times, 20 September 1970].

1990s


The first approved gene therapy case in the United States took place on September 14, 1990, at the National Institute of Health. It was performed on a four year old girl named Ashanti DaSilva. It was a treatment for a genetic defect that left her with an immune system deficiency. The effects were only temporary, but successful (Boylan 313).

New gene therapy approach repairs errors in messenger RNA derived from defective genes. This technique has the potential to treat the blood disorder thalassaemia, cystic fibrosis
Cystic fibrosis
Cystic fibrosis is a recessive genetic disease affecting most critically the lungs, and also the pancreas, liver, and intestine...

, and some cancers.
Researchers at Case Western Reserve University
Case Western Reserve University
Case Western Reserve University is a private research university located in Cleveland, Ohio, USA...

 and Copernicus Therapeutics are able to create tiny liposomes 25 nanometers across that can carry therapeutic DNA through pores in the nuclear membrane.

Sickle cell disease is successfully treated in mice.

in 1992 Doctor Claudio Bordignon working at the Vita-Salute San Raffaele University, Milan, Italy performed the first procedure of gene therapy using hematopoietic stem cells as vectors to deliver genes intended to correct hereditary diseases. In 2002 this work led to the publication of the first successful gene therapy treatment for adenosine deaminase-deficiency (SCID).
The success of a multi-center trial for treating children with SCID (severe combined immune deficiency or "bubble boy" disease) held from 2000 and 2002 was questioned when two of the ten children treated at the trial's Paris center developed a leukemia-like condition. Clinical trials were halted temporarily in 2002, but resumed after regulatory review of the protocol in the United States, the United Kingdom, France, Italy, and Germany.

In 1993 Andrew Gobea was born with severe combined immunodeficiency
Severe combined immunodeficiency
Severe combined immunodeficiency , is a genetic disorder in which both "arms" of the adaptive immune system are impaired due to a defect in one of several possible genes. SCID is a severe form of heritable immunodeficiency...

 (SCID). Genetic screening before birth showed that he had SCID. Blood was removed from Andrew's placenta and umbilical cord immediately after birth, containing stem cells. The allele that codes for ADA
Adenosine deaminase
Adenosine deaminase is an enzyme involved in purine metabolism. It is needed for the breakdown of adenosine from food and for the turnover of nucleic acids in tissues.-Reactions:...

 was obtained and was inserted into a retrovirus. Retroviruses and stem cells were mixed, after which they entered and inserted the gene into the stem cells' chromosomes. Stem cells containing the working ADA gene were injected into Andrew's blood system via a vein. Injections of the ADA enzyme were also given weekly. For four years T-cells (white blood cells), produced by stem cells, made ADA enzymes using the ADA gene. After four years more treatment was needed.

The 1999 death of Jesse Gelsinger
Jesse Gelsinger
Jesse Gelsinger was the first person publicly identified as having died in a clinical trial for gene therapy....

 in a gene-therapy experiment resulted in a significant setback to gene therapy research in the United States. As a result, the U.S. FDA suspended several clinical trials pending the re-evaluation of ethical and procedural practices in the field.

2003


In 2003 a University of California, Los Angeles
University of California, Los Angeles
The University of California, Los Angeles is a public research university located in the Westwood neighborhood of Los Angeles, California, USA. It was founded in 1919 as the "Southern Branch" of the University of California and is the second oldest of the ten campuses...

 research team inserted genes into the brain using liposomes coated in a polymer
Polymer
A polymer is a large molecule composed of repeating structural units. These subunits are typically connected by covalent chemical bonds...

 called polyethylene glycol
Polyethylene glycol
Polyethylene glycol is a polyether compound with many applications from industrial manufacturing to medicine. It has also been known as polyethylene oxide or polyoxyethylene , depending on its molecular weight, and under the tradename Carbowax.-Available forms:PEG, PEO, or POE refers to an...

. The transfer of genes into the brain is a significant achievement because viral vectors are too big to get across the blood-brain barrier
Blood-brain barrier
The blood–brain barrier is a separation of circulating blood and the brain extracellular fluid in the central nervous system . It occurs along all capillaries and consists of tight junctions around the capillaries that do not exist in normal circulation. Endothelial cells restrict the diffusion...

. This method has potential for treating Parkinson's disease
Parkinson's disease
Parkinson's disease is a degenerative disorder of the central nervous system...

.

RNA interference
RNA interference
RNA interference is a process within living cells that moderates the activity of their genes. Historically, it was known by other names, including co-suppression, post transcriptional gene silencing , and quelling. Only after these apparently unrelated processes were fully understood did it become...

 or gene silencing
Gene silencing
Gene silencing is a general term describing epigenetic processes of gene regulation. The term gene silencing is generally used to describe the "switching off" of a gene by a mechanism other than genetic modification...

 may be a new way to treat Huntington's disease
Huntington's disease
Huntington's disease, chorea, or disorder , is a neurodegenerative genetic disorder that affects muscle coordination and leads to cognitive decline and dementia. It typically becomes noticeable in middle age. HD is the most common genetic cause of abnormal involuntary writhing movements called chorea...

. Short pieces of double-stranded RNA (short, interfering RNAs or siRNAs) are used by cells to degrade RNA of a particular sequence. If a siRNA
Sírna
Sírna Sáeglach , son of Dian mac Demal, son of Demal mac Rothechtaid, son of Rothechtaid mac Main, was, according to medieval Irish legend and historical tradition, a High King of Ireland...

 is designed to match the RNA copied from a faulty gene, then the abnormal protein product of that gene will not be produced.

2006


Scientists at the National Institutes of Health
National Institutes of Health
The National Institutes of Health are an agency of the United States Department of Health and Human Services and are the primary agency of the United States government responsible for biomedical and health-related research. Its science and engineering counterpart is the National Science Foundation...

 (Bethesda, Maryland
Bethesda, Maryland
Bethesda is a census designated place in southern Montgomery County, Maryland, United States, just northwest of Washington, D.C. It takes its name from a local church, the Bethesda Meeting House , which in turn took its name from Jerusalem's Pool of Bethesda...

) have successfully treated metastatic melanoma in two patients using killer T cells genetically retargeted to attack the cancer cells. This study constitutes one of the first demonstrations that gene therapy can be effective in treating cancer
Cancer
Cancer , known medically as a malignant neoplasm, is a large group of different diseases, all involving unregulated cell growth. In cancer, cells divide and grow uncontrollably, forming malignant tumors, and invade nearby parts of the body. The cancer may also spread to more distant parts of the...

.

In March 2006 an international group of scientists announced the successful use of gene therapy to treat two adult patients for a disease affecting myeloid
Myeloid
The term myeloid suggests an origin in the bone marrow or spinal cord, or a resemblance to the marrow or spinal cord.In hematopoiesis, the term "myeloid cell" is used to describe any leukocyte that is not a lymphocyte...

 cells. The study, published in Nature Medicine, is believed to be the first to show that gene therapy can cure diseases of the myeloid
Myeloid
The term myeloid suggests an origin in the bone marrow or spinal cord, or a resemblance to the marrow or spinal cord.In hematopoiesis, the term "myeloid cell" is used to describe any leukocyte that is not a lymphocyte...

 system.

In May 2006 a team of scientists led by Dr. Luigi Naldini and Dr. Brian Brown from the San Raffaele Telethon Institute for Gene Therapy (HSR-TIGET) in Milan
Milan
Milan is the second-largest city in Italy and the capital city of the region of Lombardy and of the province of Milan. The city proper has a population of about 1.3 million, while its urban area, roughly coinciding with its administrative province and the bordering Province of Monza and Brianza ,...

, Italy
Italy
Italy , officially the Italian Republic languages]] under the European Charter for Regional or Minority Languages. In each of these, Italy's official name is as follows:;;;;;;;;), is a unitary parliamentary republic in South-Central Europe. To the north it borders France, Switzerland, Austria and...

 reported a breakthrough for gene therapy in which they developed a way to prevent the immune system from rejecting a newly delivered gene. Similar to organ transplant
Organ transplant
Organ transplantation is the moving of an organ from one body to another or from a donor site on the patient's own body, for the purpose of replacing the recipient's damaged or absent organ. The emerging field of regenerative medicine is allowing scientists and engineers to create organs to be...

ation, gene therapy has been plagued by the problem of immune rejection. So far, delivery of the 'normal' gene has been difficult because the immune system
Immune system
An immune system is a system of biological structures and processes within an organism that protects against disease by identifying and killing pathogens and tumor cells. It detects a wide variety of agents, from viruses to parasitic worms, and needs to distinguish them from the organism's own...

 recognizes the new gene as foreign and rejects the cells carrying it. To overcome this problem, the HSR-TIGET group utilized a newly uncovered network of genes regulated by molecules known as microRNAs. Dr. Naldini's group reasoned that they could use this natural function of microRNA to selectively turn off the identity of their therapeutic gene in cells of the immune system and prevent the gene from being found and destroyed. The researchers injected mice with the gene containing an immune-cell microRNA target sequence, and the mice did not reject the gene, as previously occurred when vectors without the microRNA target sequence were used. This work will have important implications for the treatment of hemophilia and other genetic diseases by gene therapy.

In November 2006 Preston Nix from the University of Pennsylvania School of Medicine
University of Pennsylvania School of Medicine
The Perelman School of Medicine , formerly the University of Pennsylvania School of Medicine, was founded in 1765, making it the oldest American medical school. As part of the University of Pennsylvania, it is located in the University City section of Philadelphia, Pennsylvania. It is widely...

 reported on VRX496, a gene-based immunotherapy
Immunotherapy
Immunotherapy is a medical term defined as the "treatment of disease by inducing, enhancing, or suppressing an immune response". Immunotherapies designed to elicit or amplify an immune response are classified as activation immunotherapies. While immunotherapies that reduce or suppress are...

 for the treatment of human immunodeficiency virus (HIV) that uses a lentiviral
Lentivirus
Lentivirus is a genus of slow viruses of the Retroviridae family, characterized by a long incubation period...

 vector
Viral vector
Viral vectors are a tool commonly used by molecular biologists to deliver genetic material into cells. This process can be performed inside a living organism or in cell culture . Viruses have evolved specialized molecular mechanisms to efficiently transport their genomes inside the cells they infect...

 for delivery of an antisense
Sense (molecular biology)
In molecular biology and genetics, sense is a concept used to compare the polarity of nucleic acid molecules, such as DNA or RNA, to other nucleic acid molecules...

 gene against the HIV envelope. In the Phase I trial enrolling five subjects with chronic HIV infection who had failed to respond to at least two antiretroviral regimens, a single intravenous infusion of autologous CD4
CD4
CD4 is a glycoprotein expressed on the surface of T helper cells, monocytes, macrophages, and dendritic cells. It was discovered in the late 1970s and was originally known as leu-3 and T4 before being named CD4 in 1984...

 T cells genetically modified with VRX496 was safe and well tolerated. All patients had stable or decreased viral load; four of the five patients had stable or increased CD4 T cell counts. In addition, all five patients had stable or increased immune response to HIV antigen
Antigen
An antigen is a foreign molecule that, when introduced into the body, triggers the production of an antibody by the immune system. The immune system will then kill or neutralize the antigen that is recognized as a foreign and potentially harmful invader. These invaders can be molecules such as...

s and other pathogen
Pathogen
A pathogen gignomai "I give birth to") or infectious agent — colloquially, a germ — is a microbe or microorganism such as a virus, bacterium, prion, or fungus that causes disease in its animal or plant host...

s. This was the first evaluation of a lentiviral vector administered in U.S. Food and Drug Administration-approved human clinical trials for any disease. Data from an ongoing Phase I/II clinical trial were presented at CROI 2009.

2007


On 1 May 2007 Moorfields Eye Hospital
Moorfields Eye Hospital
Moorfields Eye Hospital NHS Foundation Trust is an NHS eye hospital located in London, United Kingdom. It is the oldest and largest eye hospital in the world and is internationally renowned for its comprehensive clinical and research activities...

 and University College London
University College London
University College London is a public research university located in London, United Kingdom and the oldest and largest constituent college of the federal University of London...

's Institute of Ophthalmology announced the world's first gene therapy trial for inherited retinal disease. The first operation was carried out on a 23 year-old British
United Kingdom
The United Kingdom of Great Britain and Northern IrelandIn the United Kingdom and Dependencies, other languages have been officially recognised as legitimate autochthonous languages under the European Charter for Regional or Minority Languages...

 male, Robert Johnson
Robert Simon Johnson
Robert Johnson is the first person in the world to undergo gene therapy treatment on the eye, in early 2007. Born with an inherited retinal dystrophy Robert was operated on by a team at world renowned Moorfields Eye Hospital in London, led by Mr James Bainbridge, as part of a gene therapy clinical...

, in early 2007. Leber's congenital amaurosis
Leber's congenital amaurosis
Leber's congenital amaurosis is a rare inherited eye disease that appears at birth or in the first few months of life, and affects around 1 in 80,000 of the population.It was first described by Theodor Leber in the 19th century...

 is an inherited blinding disease caused by mutations in the RPE65
RPE65
Retinal pigment epithelium-specific 65 kDa protein is a protein that in humans is encoded by the RPE65 gene.-Function:The retinal pigment epithelium-specific 65 kDa protein is located in the retinal pigment epithelium and is involved in the conversion of all-trans retinol to 11-cis retinal during...

 gene. The results of the Moorfields/UCL trial were published in New England Journal of Medicine
New England Journal of Medicine
The New England Journal of Medicine is an English-language peer-reviewed medical journal published by the Massachusetts Medical Society. It describes itself as the oldest continuously published medical journal in the world.-History:...

 in April 2008. They researched the safety of the subretinal delivery of recombinant adeno associated virus (AAV) carrying RPE65 gene, and found it yielded positive results, with patients having modest increase in vision, and, perhaps more importantly, no apparent side-effects.

2009


In September 2009, the journal Nature
Nature (journal)
Nature, first published on 4 November 1869, is ranked the world's most cited interdisciplinary scientific journal by the Science Edition of the 2010 Journal Citation Reports...

 reported that researchers at the University of Washington
University of Washington
University of Washington is a public research university, founded in 1861 in Seattle, Washington, United States. The UW is the largest university in the Northwest and the oldest public university on the West Coast. The university has three campuses, with its largest campus in the University...

 and University of Florida
University of Florida
The University of Florida is an American public land-grant, sea-grant, and space-grant research university located on a campus in Gainesville, Florida. The university traces its historical origins to 1853, and has operated continuously on its present Gainesville campus since September 1906...

 were able to give trichromatic vision to squirrel monkeys using gene therapy, a hopeful precursor to a treatment for color blindness
Color blindness
Color blindness or color vision deficiency is the inability or decreased ability to see color, or perceive color differences, under lighting conditions when color vision is not normally impaired...

 in humans. In November 2009, the journal Science
Science (journal)
Science is the academic journal of the American Association for the Advancement of Science and is one of the world's top scientific journals....

 reported that researchers succeeded at halting a fatal brain disease, adrenoleukodystrophy
Adrenoleukodystrophy
Adrenoleukodystrophy is a rare, inherited disorder that leads to progressive brain damage, failure of the adrenal glands and eventually death. ALD is a disease in a group of genetic disorders called leukodystrophies, whose chief feature is damage to myelin...

, using a vector derived from HIV
HIV
Human immunodeficiency virus is a lentivirus that causes acquired immunodeficiency syndrome , a condition in humans in which progressive failure of the immune system allows life-threatening opportunistic infections and cancers to thrive...

 to deliver the gene for the missing enzyme.

2010


A paper by Komáromy et al. published in April 2010, deals with gene therapy for a form of achromatopsia
Achromatopsia
Achromatopsia , is a medical syndrome that exhibits symptoms relating to at least five separate individual disorders. Although the term may refer to acquired disorders such as color agnosia and cerebral achromatopsia, it typically refers to an autosomal recessive congenital color vision disorder,...

 in dogs. Achromatopsia, or complete color blindness, is presented as an ideal model to develop gene therapy directed to cone photoreceptors. Cone function and day vision have been restored for at least 33 months in two young dogs with achromatopsia. However, the therapy was less efficient for older dogs.

2011


In 2007 and 2008, a man being treated by Gero Hütter
Gero Hütter
Gero Hütter is a German hematologist. Huetter and his medical team transplanted bone marrow deficient in a key HIV receptor to a leukemia patient, Timothy Ray Brown, who was also infected with human immunodeficiency virus . Subsequently, the patient's circulating HIV dropped to undetectable levels...

 was cured of HIV
HIV
Human immunodeficiency virus is a lentivirus that causes acquired immunodeficiency syndrome , a condition in humans in which progressive failure of the immune system allows life-threatening opportunistic infections and cancers to thrive...

 by repeated Hematopoietic stem cell transplantation (see also Allogeneic stem cell transplantation
Allogeneic stem cell transplantation
Allogeneic stem cell transplantation is a procedure in which a person receives blood-forming stem cells from a genetically similar, but not identical, donor. This is often a sister or brother, but could be an unrelated donor...

, Allogeneic bone marrow transplantation
Allogeneic bone marrow transplantation
Allogeneic bone marrow transplantation is a procedure in which a person receives stem cells from a genetically similar, but not identical, donor.- External links :...

, Allotransplantation
Allotransplantation
Allotransplantation is the transplantation of cells, tissues, or organs, sourced from a genetically non-identical member of the same species as the recipient. The transplant is called an allograft or allogeneic transplant or homograft...

) with double-delta-32 mutation which disables the CCR5
CCR5
C-C chemokine receptor type 5, also known as CCR5, is a protein that in humans is encoded by the CCR5 gene. CCR5 is a member of the beta chemokine receptors family of integral membrane proteins...

 receptor; this cure was not completely accepted by the medical community until 2011. This cure required complete ablation of existing bone marrow
Bone marrow
Bone marrow is the flexible tissue found in the interior of bones. In humans, bone marrow in large bones produces new blood cells. On average, bone marrow constitutes 4% of the total body mass of humans; in adults weighing 65 kg , bone marrow accounts for approximately 2.6 kg...

 which is very debilitating.

Problems and ethics


For the safety of gene therapy, the Weismann barrier
Weismann barrier
The Weismann barrier is the principle that hereditary information moves only from genes to body cells, and never in reverse. In more precise terminology hereditary information moves only from germline cells to somatic cells .This does not refer to the central dogma of molecular biology which...

 is fundamental in the current thinking. Soma-to-germline feedback should therefore be impossible. However, there are indications that the Weismann barrier can be breached. One way it might possibly be breached is if the treatment were somehow misapplied and spread to the testes and therefore would infect the germline against the intentions of the therapy.

Some of the problems of gene therapy include:
  • Short-lived nature of gene therapy – Before gene therapy can become a permanent cure for any condition, the therapeutic DNA introduced into target cells must remain functional and the cells containing the therapeutic DNA must be long-lived and stable. Problems with integrating therapeutic DNA into the genome and the rapidly dividing nature of many cells prevent gene therapy from achieving any long-term benefits. Patients will have to undergo multiple rounds of gene therapy.
  • Immune response – Anytime a foreign object is introduced into human tissues, the immune system has evolved to attack the invader. The risk of stimulating the immune system in a way that reduces gene therapy effectiveness is always a possibility. Furthermore, the immune system
    Immune system
    An immune system is a system of biological structures and processes within an organism that protects against disease by identifying and killing pathogens and tumor cells. It detects a wide variety of agents, from viruses to parasitic worms, and needs to distinguish them from the organism's own...

    's enhanced response to invaders that it has seen before makes it difficult for gene therapy to be repeated in patients.
  • Problems with viral vectors – Viruses, the carrier of choice in most gene therapy studies, present a variety of potential problems to the patient —toxicity, immune and inflammatory responses, and gene control and targeting issues. In addition, there is always the fear that the viral vector, once inside the patient, may recover its ability to cause disease.
  • Multigene disorders – Conditions or disorders that arise from mutations in a single gene are the best candidates for gene therapy. Unfortunately, some of the most commonly occurring disorders, such as heart disease
    Heart disease
    Heart disease, cardiac disease or cardiopathy is an umbrella term for a variety of diseases affecting the heart. , it is the leading cause of death in the United States, England, Canada and Wales, accounting for 25.4% of the total deaths in the United States.-Types:-Coronary heart disease:Coronary...

    , high blood pressure, Alzheimer's disease
    Alzheimer's disease
    Alzheimer's disease also known in medical literature as Alzheimer disease is the most common form of dementia. There is no cure for the disease, which worsens as it progresses, and eventually leads to death...

    , arthritis
    Arthritis
    Arthritis is a form of joint disorder that involves inflammation of one or more joints....

    , and diabetes, are caused by the combined effects of variations in many genes. Multigene or multifactorial disorders such as these would be especially difficult to treat effectively using gene therapy.
  • Chance of inducing a tumor
    Tumor
    A tumor or tumour is commonly used as a synonym for a neoplasm that appears enlarged in size. Tumor is not synonymous with cancer...

     (insertional mutagenesis) - If the DNA
    DNA
    Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...

     is integrated in the wrong place in the genome
    Genome
    In modern molecular biology and genetics, the genome is the entirety of an organism's hereditary information. It is encoded either in DNA or, for many types of virus, in RNA. The genome includes both the genes and the non-coding sequences of the DNA/RNA....

    , for example in a tumor suppressor gene
    Tumor suppressor gene
    A tumor suppressor gene, or anti-oncogene, is a gene that protects a cell from one step on the path to cancer. When this gene is mutated to cause a loss or reduction in its function, the cell can progress to cancer, usually in combination with other genetic changes.-Two-hit hypothesis:Unlike...

    , it could induce a tumor. This has occurred in clinical trials for X-linked severe combined immunodeficiency (X-SCID) patients, in which hematopoietic stem cells were transduced with a corrective transgene using a retrovirus, and this led to the development of T cell leukemia in 3 of 20 patients.


Deaths have occurred due to gene therapy, including that of Jesse Gelsinger
Jesse Gelsinger
Jesse Gelsinger was the first person publicly identified as having died in a clinical trial for gene therapy....

.

Preventive gene therapy


Preventive gene therapy is the repair of a gene with a mutation associated with a progressive disease, prior to the expression of a medical condition, in order to prevent that expression. There are a number of considerations:
  • It is hard to get U.S. FDA approval to treat a pre-symptomatic condition because it is hard to predict the complications that may arise, so it is hard to give a risk/benefit analysis. This is an obstacle to long-term therapies.
  • It is easier to gain approval for short-term therapies to treat expressed conditions rather than prevent them.
  • It is not known whether the repair of a mutation will help to treat a condition which has already progressed beyond the initial consequences of that mutation.

In popular culture

  • In the TV series Dark Angel
    Dark Angel (TV series)
    Dark Angel is an American biopunk/cyberpunk science fiction television series created by James Cameron and Charles H. Eglee. The show premiered in the United States on the Fox network on October 3, 2000, and was canceled after two seasons...

    gene therapy is mentioned as one of the practices performed on transgenics and their surrogate mothers at Manticore, and in the episode Prodigy, Dr. Tanaka uses a groundbreaking new form of gene therapy to turn Jude, a premature, vegetative baby of a crack/cocaine addict, into a boy genius.
  • Gene therapy is a crucial plot element in the video game Metal Gear Solid
    Metal Gear Solid
    is a videogame by Hideo Kojima. The game was developed by Konami Computer Entertainment Japan and first published by Konami in 1998 for the PlayStation video game console. It is the sequel to Kojimas early MSX2 computer games Metal Gear and Metal Gear 2: Solid Snake...

    , where it has been used to enhance the battle capabilities of enemy soldiers.
  • Gene therapy plays a major role in the sci-fi series Stargate Atlantis
    Stargate Atlantis
    Stargate Atlantis is a Canadian-American adventure and military science fiction television series and part of MGM's Stargate franchise. The show was created by Brad Wright and Robert C. Cooper as a spin-off series of Stargate SG-1, which was created by Wright and Jonathan Glassner and was itself...

    , as a certain type of alien technology can only be used if one has a certain gene which can be given to the members of the team through gene therapy.
  • Gene therapy also plays a major role in the plot of the James Bond
    James Bond
    James Bond, code name 007, is a fictional character created in 1953 by writer Ian Fleming, who featured him in twelve novels and two short story collections. There have been a six other authors who wrote authorised Bond novels or novelizations after Fleming's death in 1964: Kingsley Amis,...

     movie Die Another Day
    Die Another Day
    Die Another Day is the 20th spy film in the James Bond series, and the fourth and last film to star Pierce Brosnan as the fictional MI6 agent James Bond; it is also the last Bond film of the original timeline with the series being rebooted with Casino Royale...

    .
  • Gene therapy plays a recurring role in the present-time sci-fi television program ReGenesis
    ReGenesis
    ReGenesis is a Canadian television program produced by The Movie Network and Movie Central in conjunction with Shaftesbury Films. The series, which ran for four seasons, revolves around the scientists of NorBAC , a fictional organization with a lab based in Toronto...

    , where it is used to cure various diseases, enhance athletic performance and produce vast profits for bio-tech corporations. (e.g. an undetectable performance-enhancing gene therapy was used by one of the characters on himself, but to avoid copyright infringement, this gene therapy was modified from the tested-to-be-harmless original, which produced a fatal cardiovascular defect)
  • Gene therapy is the basis for the plot line of the film I Am Legend
    I Am Legend (film)
    I Am Legend is a 2007 post-apocalyptic science fiction film directed by Francis Lawrence and starring Will Smith. It is the third feature film adaptation of Richard Matheson's 1954 novel of the same name, following 1964's The Last Man on Earth and 1971's The Omega Man. Smith plays virologist Robert...

    .
  • Gene therapy is an important plot key in the game Bioshock
    Bioshock
    BioShock is a first-person shooter video game developed by 2K Boston and designed by Ken Levine. It was released for Microsoft Windows and Xbox 360 on August 21, 2007 in North America, and three days later in Europe and Australia. It became available on Steam on August 21, 2007...

     where the game contents refer to plasmids and [gene] splicers.
  • The book Next
    Next (novel)
    Next is a 2006 techno-thriller novel by Michael Crichton, the last to be published during his lifetime. Next takes place in the present world, where both the government and private investors spend billions of dollars every year on genetic research...

    by Michael Crichton
    Michael Crichton
    John Michael Crichton , best known as Michael Crichton, was an American best-selling author, producer, director, and screenwriter, best known for his work in the science fiction, medical fiction, and thriller genres. His books have sold over 200 million copies worldwide, and many have been adapted...

     unravels a story in which fictitious biotechnology companies experiment with gene therapy.
  • In the television show Alias
    Alias (TV series)
    Alias is an American action television series created by J. J. Abrams which was broadcast on ABC for five seasons, from September 30, 2001 to May 22, 2006...

    , a breakthrough in molecular gene therapy is discovered, whereby a patient's body is reshaped to identically resemble someone else. Protagonist Sydney Bristow
    Sydney Bristow
    Sydney Anne Bristow , played by Jennifer Garner, is the main character and protagonist on the television series Alias. She is an American woman with Russian-American family background who works as a spy for the CIA....

    's best friend was secretly killed and her "double" resumed her place.

Gun of God David R. Cudlip (Author)

See also

  • Antisense therapy
    Antisense therapy
    Antisense therapy is a form of treatment for genetic disorders or infections.When the genetic sequence of a particular gene is known to be causative of a particular disease, it is possible to synthesize a strand of nuc acid that will bind to the messenger RNA produced by that gene and inactivate...

  • DNA
    DNA
    Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...

  • Full Genome Sequencing
    Full genome sequencing
    Full genome sequencing , also known as whole genome sequencing , complete genome sequencing, or entire genome sequencing, is a laboratory process that determines the complete DNA sequence of an organism's genome at a single time...

  • Gene therapy for color blindness
    Gene therapy for color blindness
    Gene therapy for color blindness is an experimental gene therapy aiming to convert congenitally colorblind individuals to trichromats by introducing a photopigment gene that they lack. Though partial color blindness is considered only a mild disability and is controversial whether it is even a...

  • Genetic engineering
    Genetic engineering
    Genetic engineering, also called genetic modification, is the direct human manipulation of an organism's genome using modern DNA technology. It involves the introduction of foreign DNA or synthetic genes into the organism of interest...

  • Life extension
    Life extension
    Life extension science, also known as anti-aging medicine, experimental gerontology, and biomedical gerontology, is the study of slowing down or reversing the processes of aging to extend both the maximum and average lifespan...

  • List of life extension related topics
  • Pharmacological gene therapy
    Pharmacological gene therapy
    Pharmacological gene therapy is a new field that combines pharmacological therapy and gene therapy. It is used either to prevent a defective gene from producing its protein or to increase the concentration of normal protein produced in the body by insertion of DNA or RNA fragments...

  • Predictive Medicine
    Predictive medicine
    Predictive medicine is a rapidly emerging field of medicine that entails predicting disease and instituting preventive measures in order to either prevent the disease altogether or significantly decrease its impact upon the patient...

  • Technology assessment
    Technology assessment
    Technology assessment Technology assessment Technology assessment (TA, German Tenteractive, and communicative process that aims to contribute to the formation of public and political opinion on societal aspects of science and technology.- General description :...

  • Therapeutic gene modulation
    Therapeutic gene modulation
    Therapeutic gene modulation refers the practice of altering the expression of a gene at one of various stages, with a view to alleviate some form of ailment...



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