Gene Therapy

Gene therapy is a rapidly growing field of medicine in which genes are introduced into the body to treat diseases. Genes control heredity and provide the basic biological code for determining a cell's specific functions. Gene therapy seeks to provide genes that correct or supplant the disease-controlling functions of cells that are not, in essence, doing their job. Somatic gene therapy introduces therapeutic genes at the tissue or cellular level to treat a specific individual. Germ-line gene therapy inserts genes into reproductive cells or possibly into embryos to correct genetic defects that could be passed on to future generations. Initially conceived as an approach for treating inherited diseases, like cystic fibrosis and Huntington's disease, the scope of potential gene therapies has grown to include treatments for cancers, arthritis, and infectious diseases. Although gene therapy testing in humans has advanced rapidly, many questions surround its use. For example, some scientists are concerned that the therapeutic genes themselves may cause disease. Others fear that germ-line gene therapy may be used to control human development in ways not connected with disease, like intelligence or appearance.

The biological basis of gene therapy

Gene therapy has grown out of the science of genetics or how heredity works. Scientists know that life begins in a cell, the basic building block of all multicellular organisms. Humans, for instance, are made up of trillions of cells, each performing a specific function. Within the cell's nucleus (the center part of a cell that regulates its chemical functions) are pairs of chromosomes. These threadlike structures are made up of a single molecule of DNA (deoxyribonucleic acid), which carries the blueprint of life in the form of codes, or genes, that determine inherited characteristics.

A DNA molecule looks like two ladders with one of the sides taken off both and then twisted around each other. The rungs of these ladders meet (resulting in a spiral staircase-like structure) and are called base pairs. Base pairs are made up of nitrogen molecules and arranged in specific sequences. Millions of these base pairs, or sequences, can make up a single gene, specifically defined as a segment of the chromosome and DNA that contains certain hereditary information. The gene, or combination of genes formed by these base pairs ultimately direct an organism's growth and characteristics through the production of certain chemicals, primarily proteins, which carry out most of the body's chemical functions and biological reactions.

Scientists have long known that alterations in genes present within cells can cause inherited diseases like cystic fibrosis, sickle-cell anemia, and hemophilia. Similarly, errors in the total number of chromosomes can cause conditions such as Down syndrome or Turner's syndrome. As the study of genetics advanced, however, scientists learned that an altered genetic sequence also can make people more susceptible to diseases, like atherosclerosis, cancer, and even schizophrenia. These diseases have a genetic component, but also are influenced by environmental factors (like diet and lifestyle). The objective of gene therapy is to treat diseases by introducing functional genes into the body to alter the cells involved in the disease process by either replacing missing genes or providing copies of functioning genes to replace nonfunctioning ones. The inserted genes can be naturally-occurring genes that produce the desired effect or may be genetically engineered (or altered) genes.

Scientists have known how to manipulate a gene's structure in the laboratory since the early 1970s through a process called gene splicing. The process involves removing a fragment of DNA containing the specific genetic sequence desired, then inserting it into the DNA of another gene. The resultant product is called recombinant DNA and the process is genetic engineering.

There are basically two types of gene therapy. Germ-line gene therapy introduces genes into reproductive cells (sperm and eggs) or someday possibly into embryos in hopes of correcting genetic abnormalities that could be passed on to future generations. Most of the current work in applying gene therapy, however, has been in the realm of somatic gene therapy. In this type of gene therapy, therapeutic genes are inserted into tissue or cells to produce a naturally occurring protein or substance that is lacking or not functioning correctly in an individual patient.

Viral vectors

In both types of therapy, scientists need something to transport either the entire gene or a recombinant DNA to the cell's nucleus, where the chromosomes and DNA reside. In essence, vectors are molecular delivery trucks. One of the first and most popular vectors developed were viruses because they invade cells as part of the natural infection process. Viruses have the potential to be excellent vectors because they have a specific relationship with the host in that they colonize certain cell types and tissues in specific organs. As a result, vectors are chosen according to their attraction to certain cells and areas of the body.

One of the first vectors used was retroviruses. Because these viruses are easily cloned (artificially reproduced) in the laboratory, scientists have studied them extensively and learned a great deal about their biological action. They also have learned how to remove the genetic information that governs viral replication, thus reducing the chances of infection.

Retroviruses work best in actively dividing cells, but cells in the body are relatively stable and do not divide often. As a result, these cells are used primarily for ex vivo (outside the body) manipulation. First, the cells are removed from the patient's body, and the virus, or vector, carrying the gene is inserted into them. Next, the cells are placed into a nutrient culture where they grow and replicate. Once enough cells are gathered, they are returned to the body, usually by injection into the blood stream. Theoretically, as long as these cells survive, they will provide the desired therapy.

Another class of viruses, called the adenoviruses, also may prove to be good gene vectors. These viruses can effectively infect nondividing cells in the body, where the desired gene product then is expressed naturally. In addition to being a more efficient approach to gene transportation, these viruses, which cause respiratory infections, are more easily purified and made stable than retroviruses, resulting in less chance of an unwanted viral infection. However, these viruses live for several days in the body, and some concern surrounds the possibility of infecting others with the viruses through sneezing or coughing. Other viral vectors include influenza viruses, Sindbis virus, and a herpes virus that infects nerve cells.

Scientists also have delved into nonviral vectors. These vectors rely on the natural biological process in which cells uptake (or gather) macromolecules. One approach is to use liposomes, globules of fat produced by the body and taken up by cells. Scientists also are investigating the introduction of raw recombinant DNA by injecting it into the bloodstream or placing it on microscopic beads of gold shot into the skin with a "gene-gun." Another possible vector under development is based on dendrimer molecules. A class of polymers (naturally occurring or artificial substances that have a high molecular weight and formed by smaller molecules of the same or similar substances), is "constructed" in the laboratory by combining these smaller molecules. They have been used in manufacturing Styrofoam, polyethylene cartons, and Plexiglass. In the laboratory, dendrimers have shown the ability to transport genetic material into human cells. They also can be designed to form an affinity for particular cell membranes by attaching to certain sugars and protein groups.

The history of gene therapy

In the early 1970s, scientists proposed "gene surgery" for treating inherited diseases caused by faulty genes. The idea was to take out the disease-causing gene and surgically implant a gene that functioned properly. Although sound in theory, scientists, then and now, lack the biological knowledge or technical expertise needed to perform such a precise surgery in the human body.

However, in 1983, a group of scientists from Baylor College of Medicine in Houston, Texas, proposed that gene therapy could one day be a viable approach for treating Lesch-Nyhan disease, a rare neurological disorder. The scientists conducted experiments in which an enzyme-producing gene (a specific type of protein) for correcting the disease was injected into a group of cells for replication. The scientists theorized the cells could then be injected into people with Lesch-Nyhan disease, thus correcting the genetic defect that caused the disease.

As the science of genetics advanced throughout the 1980s, gene therapy gained an established foothold in the minds of medical scientists as a promising approach to treatments for specific diseases. One of the major reasons for the growth of gene therapy was scientists' increasing ability to identify the specific genetic malfunctions that caused inherited diseases. Interest grew as further studies of DNA and chromosomes (where genes reside) showed that specific genetic abnormalities in one or more genes occurred in successive generations of certain family members who suffered from diseases like intestinal cancer, bipolar disorder, Alzheimer's disease, heart disease, diabetes, and many more. Although the genes may not be the only cause of the disease in all cases, they may make certain individuals more susceptible to developing the disease because of environmental influences, like smoking, pollution, and stress. In fact, some scientists theorize that all diseases may have a genetic component.

On September 14, 1990, a four-year old girl suffering from a genetic disorder that prevented her body from producing a crucial enzyme became the first person to undergo gene therapy in the United States. Because her body could not produce adenosine deaminase (ADA), she had a weakened immune system, making her extremely susceptible to severe, life-threatening infections. W. French Anderson and colleagues at the National Institutes of Health's Clinical Center in Bethesda, Maryland, took white blood cells (which are crucial to proper immune system functioning) from the girl, inserted ADA producing genes into them, and then transfused the cells back into the patient. Although the young girl continued to show an increased ability to produce ADA, debate arose as to whether the improvement resulted from the gene therapy or from an additional drug treatment she received.

Nevertheless, a new era of gene therapy began as more and more scientists sought to conduct clinical trial (testing in humans) research in this area. In that same year, gene therapy was tested on patients suffering from melanoma (skin cancer). The goal was to help them produce antibodies (disease fighting substances in the immune system) to battle the cancer.

These experiments have spawned an ever growing number of attempts at gene therapies designed to perform a variety of functions in the body. For example, a gene therapy for cystic fibrosis aims to supply a gene that alters cells, enabling them to produce a specific protein to battle the disease. Another approach was used for brain cancer patients, in which the inserted gene was designed to make the cancer cells more likely to respond to drug treatment. Another gene therapy approach for patients suffering from artery blockage, which can lead to strokes, induces the growth of new blood vessels near clogged arteries, thus ensuring normal blood circulation.

Currently, there are a host of new gene therapy agents in clinical trials. In the United States, both nucleic acid based (in vivo ) treatments and cell-based (ex vivo ) treatments are being investigated. Nucleic acid based gene therapy uses vectors (like viruses) to deliver modified genes to target cells. Cell-based gene therapy techniques remove cells from the patient in order to genetically alter them then reintroduce them to the patient's body. Presently, gene therapies for the following diseases are being developed: cystic fibrosis (using adenoviral vector), HIV infection (cell-based), malignant melanoma (cell-based), Duchenne muscular dystrophy (cell-based), hemophilia B (cell-based), kidney cancer (cell-based), Gaucher's Disease (retroviral vector), breast cancer (retroviral vector), and lung cancer (retroviral vector). When a cell or individual is treated using gene therapy and successful incorporation of engineered genes has occurred, the cell or individual is said to be transgenic.

The medical establishment's contribution to transgenic research has been supported by increased government funding. In 1991, the U.S. government provided $58 million for gene therapy research, with increases in funding of $15-40 million dollars a year over the following four years. With fierce competition over the promise of societal benefit in addition to huge profits, large pharmaceutical corporations have moved to the forefront of transgenic research. In an effort to be first in developing new therapies, and armed with billions of dollars of research funds, such corporations are making impressive strides toward making gene therapy a viable reality in the treatment of once elusive diseases.

Diseases targeted for treatment by gene therapy

The potential scope of gene therapy is enormous. More than 4,200 diseases have been identified as resulting directly from abnormal genes, and countless others that may be partially influenced by a person's genetic makeup. Initial research has concentrated on developing gene therapies for diseases whose genetic origins have been established and for other diseases that can be cured or improved by substances genes produce.

The following are examples of potential gene therapies. People suffering from cystic fibrosis lack a gene needed to produce a salt-regulating protein. This protein regulates the flow of chloride into epithelial cells, (the cells that line the inner and outer skin layers) that cover the air passages of the nose and lungs. Without this regulation, patients with cystic fibrosis build up a thick mucus that makes them prone to lung infections. A gene therapy technique to correct this abnormality might employ an adenovirus to transfer a normal copy of what scientists call the cystic fibrosis transmembrane conductance regulator, or CTRF, gene. The gene is introduced into the patient by spraying it into the nose or lungs. Researchers announced in 2004 that they had, for the first time, treated a dominant neurogenerative disease called Spinocerebella ataxia type 1, with gene therapy. This could lead to treating similar diseases such as Huntingtons disease. They also announced a single intravenous injection could deliver therapy to all muscles, perhaps providing hope to people with muscular dystrophy.

Familial hypercholesterolemia (FH) also is an inherited disease, resulting in the inability to process cholesterol properly, which leads to high levels of artery-clogging fat in the blood stream. Patients with FH often suffer heart attacks and strokes because of blocked arteries. A gene therapy approach used to battle FH is much more intricate than most gene therapies because it involves partial surgical removal of patients' livers (ex vivo transgene therapy). Corrected copies of a gene that serve to reduce cholesterol build-up are inserted into the liver sections, which then are transplanted back into the patients.

Gene therapy also has been tested on patients with AIDS. AIDS is caused by the human immunodeficiency virus (HIV), which weakens the body's immune system to the point that sufferers are unable to fight off diseases like pneumonias and cancer. In one approach, genes that produce specific HIV proteins have been altered to stimulate immune system functioning without causing the negative effects that a complete HIV molecule has on the immune system. These genes are then injected in the patient's blood stream. Another approach to treating AIDS is to insert, via white blood cells, genes that have been genetically engineered to produce a receptor that would attract HIV and reduce its chances of replicating. In 2004, researchers reported that had developed a new vaccine concept for HIV, but the details were still in development.

Several cancers also have the potential to be treated with gene therapy. A therapy tested for melanoma, or skin cancer, involves introducing a gene with an anticancer protein called tumor necrosis factor (TNF) into test tube samples of the patient's own cancer cells, which are then reintroduced into the patient. In brain cancer, the approach is to insert a specific gene that increases the cancer cells' susceptibility to a common drug used in fighting the disease. In 2003, researchers reported that they had harnessed the cell killing properties of adenoviruses to treat prostate cancer. A 2004 report said that researchers had developed a new DNA vaccine that targeted the proteins expressed in cervical cancer cells.

Gaucher disease is an inherited disease caused by a mutant gene that inhibits the production of an enzyme called glucocerebrosidase. Patients with Gaucher disease have enlarged livers and spleens and eventually their bones deteriorate. Clinical gene therapy trials focus on inserting the gene for producing this enzyme.

Gene therapy also is being considered as an approach to solving a problem associated with a surgical procedure known as balloon angioplasty. In this procedure, a stent (in this case, a type of tubular scaffolding) is used to open the clogged artery. However, in response to the trauma of the stent insertion, the body initiates a natural healing process that produces too many cells in the artery and results in restenosis, or reclosing of the artery. The gene therapy approach to preventing this unwanted side effect is to cover the outside of the stents with a soluble gel. This gel contains vectors for genes that reduce this overactive healing response.

Regularly throughout the past decade, and no doubt over future years, scientists have and will come up with new possible ways for gene therapy to help treat human disease. Recent advancements include the possibility of reversing hearing loss in humans with experimental growing of new sensory cells in adult guinea pigs, and avoiding amputation in patients with severe circulatory problems in their legs with angiogenic growth factors.

The Human Genome Project

Although great strides have been made in gene therapy in a relatively short time, its potential usefulness has been limited by lack of scientific data concerning the multitude of functions that genes control in the human body. For instance, it is now known that the vast majority of genetic material does not store information for the creation of proteins, but rather is involved in the control and regulation of gene expression, and is, thus, much more difficult to interpret. Even so, each individual cell in the body carries thousands of genes coding for proteins, with some estimates as high as 150,000 genes. For gene therapy to advance to its full potential, scientists must discover the biological role of each of these individual genes and where the base pairs that make them up are located on DNA.

To address this issue, the National Institutes of Health initiated the Human Genome Project in 1990. Led by James D. Watson (one of the co-discoverers of the chemical makeup of DNA) the project's 15-year goal is to map the entire human genome (a combination of the words gene and chromosomes). A genome map would clearly identify the location of all genes as well as the more than three billion base pairs that make them up. With a precise knowledge of gene locations and functions, scientists may one day be able to conquer or control diseases that have plagued humanity for centuries.

Scientists participating in the Human Genome Project identified an average of one new gene a day, but many expected this rate of discovery to increase. By the year 2005, their goal was to determine the exact location of all the genes on human DNA and the exact sequence of the base pairs that make them up. Some of the genes identified through this project include a gene that predisposes people to obesity, one associated with programmed cell death (apoptosis), a gene that guides HIV viral reproduction, and the genes of inherited disorders like Huntington's disease, Lou Gehrig's disease, and some colon and breast cancers. In April 2003, the finished sequence was announced, with 99% of the human genome's gene-containing regions mapped to an accuracy of 99.9%.

The future of gene therapy

Gene therapy seems elegantly simple in its concept: supply the human body with a gene that can correct a biological malfunction that causes a disease. However, there are many obstacles and some distinct questions concerning the viability of gene therapy. For example, viral vectors must be carefully controlled lest they infect the patient with a viral disease. Some vectors, like retroviruses, also can enter cells functioning properly and interfere with the natural biological processes, possibly leading to other diseases. Other viral vectors, like the adenoviruses, often are recognized and destroyed by the immune system so their therapeutic effects are short-lived. Maintaining gene expression so it performs its role properly after vector delivery is difficult. As a result, some therapies need to be repeated often to provide long-lasting benefits.

One of the most pressing issues, however, is gene regulation. Genes work in concert to regulate their functioning. In other words, several genes may play a part in turning other genes on and off. For example, certain genes work together to stimulate cell division and growth, but if these are not regulated, the inserted genes could cause tumor formation and cancer. Another difficulty is learning how to make the gene go into action only when needed. For the best and safest therapeutic effort, a specific gene should turn on, for example, when certain levels of a protein or enzyme are low and must be replaced. But the gene also should remain dormant when not needed to ensure it doesn't oversupply a substance and disturb the body's delicate chemical makeup.

One approach to gene regulation is to attach other genes that detect certain biological activities and then react as a type of automatic off-and-on switch that regulates the activity of the other genes according to biological cues. Although still in the rudimentary stages, researchers are making headway in inhibiting some gene functioning by using a synthetic DNA to block gene transcriptions (the copying of genetic information). This approach may have implications for gene therapy.

The ethics of gene therapy

While gene therapy holds promise as a revolutionary approach to treating disease, ethical concerns over its use and ramifications have been expressed by scientists and lay people alike. For example, since much needs to be learned about how these genes actually work and their long-term effect, is it ethical to test these therapies on humans, where they could have a disastrous result? As with most clinical trials concerning new therapies, including many drugs, the patients participating in these studies usually have not responded to more established therapies and often are so ill the novel therapy is their only hope for long-term survival.

Another questionable outgrowth of gene therapy is that scientists could possibly manipulate genes to genetically control traits in human offspring that are not health related. For example, perhaps a gene could be inserted to ensure that a child would not be bald, a seemingly harmless goal. However, what if genetic manipulation was used to alter skin color, prevent homosexuality, or ensure good looks? If a gene is found that can enhance intelligence of children who are not yet born, will everyone in society, the rich and the poor, have access to the technology or will it be so expensive only the elite can afford it?

The Human Genome Project, which plays such an integral role for the future of gene therapy, also has social repercussions. If individual genetic codes can be determined, will such information be used against people? For example, will someone more susceptible to a disease have to pay higher insurance premiums or be denied health insurance altogether? Will employers discriminate between two potential employees, one with a "healthy" genome and the other with genetic abnormalities?

Some of these concerns can be traced back to the eugenics movement popular in the first half of the twentieth century. This genetic "philosophy" was a societal movement that encouraged people with "positive" traits to reproduce while those with less desirable traits were sanctioned from having children. Eugenics was used to pass strict immigration laws in the United States, barring less suitable people from entering the country lest they reduce the quality of the country's collective gene pool. Probably the most notorious example of eugenics in action was the rise of Nazism in Germany, which resulted in the Eugenic Sterilization Law of 1933. The law required sterilization for those suffering from certain disabilities and even for some who were simply deemed "ugly." To ensure that this novel science is not abused, many governments have established organizations specifically for overseeing the development of gene therapy. In the United States, the Food and Drug Administration (FDA) and the National Institutes of Health require scientists to take a precise series of steps and meet stringent requirements before proceeding with clinical trials. As of mid-2004, more than 300 companies were carrying out gene medicine developments and 500 clinical trials were underway. How to deliver the therapy is the key to unlocking many of the researchers discoveries.

In fact, gene therapy has been immersed in more controversy and surrounded by more scrutiny in both the health and ethical arena than most other technologies (except, perhaps, for cloning) that promise to substantially change society. Despite the health and ethical questions surrounding gene therapy, the field will continue to grow and is likely to change medicine faster than any previous medical advancement.

KEY TERMS

Cell— The smallest living unit of the body that groups together to form tissues and help the body perform specific functions.

Chromosome— A microscopic thread-like structure found within each cell of the body, consisting of a complex of proteins and DNA. Humans have 46 chromosomes arranged into 23 pairs. Changes in either the total number of chromosomes or their shape and size (structure) may lead to physical or mental abnormalities.

Clinical trial— The testing of a drug or some other type of therapy in a specific population of patients.

Clone— A cell or organism derived through asexual (without sex) reproduction containing the identical genetic information of the parent cell or organism.

Deoxyribonucleic acid (DNA)— The genetic material in cells that holds the inherited instructions for growth, development, and cellular functioning.

Embryo— The earliest stage of development of a human infant, usually used to refer to the first eight weeks of pregnancy. The term fetus is used from roughly the third month of pregnancy until delivery.

Enzyme— A protein that causes a biochemical reaction or change without changing its own structure or function.

Eugenics— A social movement in which the population of a society, country, or the world is to be improved by controlling the passing on of hereditary information through mating.

Gene— A building block of inheritance, which contains the instructions for the production of a particular protein, and is made up of a molecular sequence found on a section of DNA. Each gene is found on a precise location on a chromosome.

Gene transcription— The process by which genetic information is copied from DNA to RNA, resulting in a specific protein formation.

Genetic engineering— The manipulation of genetic material to produce specific results in an organism.

Genetics— The study of hereditary traits passed on through the genes.

Germ-line gene therapy— The introduction of genes into reproductive cells or embryos to correct inherited genetic defects that can cause disease.

Liposome— Fat molecule made up of layers of lipids.

Macromolecules— A large molecule composed of thousands of atoms.

Nitrogen— A gaseous element that makes up the base pairs in DNA.

Nucleus— The central part of a cell that contains most of its genetic material, including chromosomes and DNA.

Protein— Important building blocks of the body, composed of amino acids, involved in the formation of body structures and controlling the basic functions of the human body.

Somatic gene therapy— The introduction of genes into tissue or cells to treat a genetic related disease in an individual.

Vectors— Something used to transport genetic information to a cell.

Resources

PERIODICALS

Abella, Harold. "Gene Therapy May Save Limbs." Diagnostic Imaging (May 1, 2003): 16.

Christensen R. "Cutaneous Gene Therapy—An Update." Histochemical Cell Biology (January 2001): 73-82.

"Gene Therapy Important Part of Cancer Research." Cancer Gene Therapy Week (June 30, 2003): 12.

"Initial Sequencing and Analysis of the Human Genome." Nature (February 15, 2001): 860-921.

Kingsman, Alan. "Gene Therapy Moves On." SCRIP World Pharmaceutical News (July 7, 2004): 19:ndash;21.

Nevin, Norman. "What Has Happened to Gene Therapy?" European Journal of Pediatrics (2000): S240-S242.

"New DNA Vaccine Targets Proteins Expressed in Cervical Cancer Cells." Gene Therapy Weekly (September 9, 2004): 14.

"New Research on the Progress of Gene Therapy Presented at Meeting." Obesity, Fitness & Wellness Week (July 3, 2004): 405.

Pekkanen, John. "Genetics: Medicine's Amazing Leap." Readers Digest (September 1991): 23-32.

Silverman, Jennifer, and Steve Perlstein. "Genome Project Completed." Family Practice News (May 15, 2003): 50-51.

"Study Highlights Potential Danger of Gene Therapy." Drug Week (June 20, 2003): 495.

"Study May Help Scientists Develop Safer Mthods for Gene Therapy." AIDS Weekly (June 30, 2003): 32.

Trabis, J. "With Gene Therapy, Ears Grow New Sensory Cells." Science News (June 7, 2003): 355.

ORGANIZATIONS

National Human Genome Research Institute. The National Institutes of Health. 9000 Rockville Pike, Bethesda, MD 20892. (301) 496-2433. 〈http://www.nhgri.nih.gov〉.

OTHER

Online Mendelian Inheritance in Man. Online genetic testing information sponsored by National Center for Biotechnology Information. 〈http://www.ncbi.nlm.nih.gov/Omim/〉.

Gene Therapy

Definition

Gene therapy is a rapidly growing field of medicine in which genes are introduced into the body to treat diseases. Genomics is the DNA which is found in an organism's total set of genes and is passed on to the offspring as information necessary for survival. Genetics is the study of the patterns of inheritance of specific traits. Genes control heredity and provide the basic biological code for determining a cell's specific functions. Gene therapy seeks to provide genes that correct or supplant the disease-controlling functions of cells that are not performing in a normal manner.

Somatic gene therapy introduces therapeutic genes at the tissue or cellular level to treat a specific individual. Germ-line gene therapy inserts genes into reproductive cells or possibly into embryos to correct genetic abnormalities that could be passed on to future generations. Initially conceived as an approach for treating inherited diseases such as cystic fibrosis and Huntington's disease, the scope of potential gene therapies has grown to include treatments for cancer, arthritis, and infectious diseases.

Description

The history of gene therapy

In the early 1970s, scientists proposed "gene surgery" for treating inherited diseases caused by abnormally functioning genes. The idea was to take out the disease-causing gene and surgically implant a gene that functioned correctly. Although sound in theory, and after some advances in science, this technique has not yet been successful.

However, in 1983, a group of scientists from Baylor College of Medicine in Houston, Texas, proposed that gene therapy could one day be a viable approach for treating Lesch-Nyhan disease, a rare neurological disorder. The scientists conducted experiments in which an enzyme-producing gene (a specific type of protein) for correcting the disease was injected into a group of cells for replication. The scientists theorized the cells could then be injected into people with Lesch-Nyhan disease, thus correcting the genetic abnormality that caused the disease.

As the science of genetics advanced throughout the 1980s, gene therapy grew in the estimation of medical scientists as a promising approach to treatments for specific diseases. One of the major reasons for the growth of gene therapy was the increasing body of knowledge available to assist in identifying the specific genetic malfunctions that caused inherited diseases. Interest grew as further studies of DNA and chromosomes (where genes reside) showed that specific genetic abnormalities in one or more genes occurred in successive generations of certain family members who experienced diseases like intestinal cancer, manic-depression (bipolar disorder), Alzheimer's disease, heart disease, diabetes, and many more. Although genes may not be the only cause of the disease in all cases, they may make certain individuals more susceptible to developing a particular condition due to environmental influences such as smoking, pollution, and stress. In fact, some scientists theorize that all diseases may have a genetic component.

The biological basis of gene therapy

Gene therapy has grown out of the science of genetics or how heredity functions. Scientists know that life begins in a cell, the basic building block of all multicellular organisms. Humans, for instance, are made up of trillions of cells, each performing a specific function. Within each cell's nucleus (the center part of a cell that regulates its chemical functions) are pairs of chromosomes. These threadlike structures are made up of deoxyribonucleic acid (DNA), which carries the blueprint of life in the form of codes, or genes, that determine dominant or recessive inherited characteristics.

A DNA molecule looks like two ladders with one of the sides taken off both and then twisted around each other—a formation known as the double helix. The rungs of these ladders meet (resulting in a spiral staircase-like structure) and are called base pairs. Base pairs are made up of nitrogen-containing molecules and arranged in specific sequences. Millions of these base pairs, or sequences, constitute a single gene, specifically defined as a segment of the chromosome and DNA that contains certain hereditary information. The gene, or combination of genes formed by these base pairs, ultimately directs an organism's growth and characteristics through the production of certain chemicals—primarily proteins that carry out most of the body's chemical functions and biological reactions.

Scientists have long known that alterations in the genes present within cells may cause inherited diseases such as cystic fibrosis, sickle cell disease, and hemophilia. Similarly, errors in entire chromosomes may cause conditions such as Down syndrome or Turner syndrome. As the study of genetics advanced, however, scientists learned that altered genetic sequences may also make people more susceptible to diseases such as atherosclerosis, cancer, and schizophrenia. These diseases have a genetic component, but are also influenced by environmental factors such as diet and lifestyle. The objective of gene therapy is to treat diseases by introducing functional genes into the body to alter the cells involved in the disease process, either by replacing missing genes or by providing copies of functioning genes to replace nonfunctioning ones. The inserted genes may be naturally occurring genes that produce the desired effect or may be engineered (or altered) genes.

Scientists have known how to manipulate a gene's structure in the laboratory since the early 1970s through a process called gene splicing. The process involves removing a fragment of DNA containing a specific desired genetic sequence and then inserting it into the DNA of another gene. The resultant product is called recombinant DNA, and the process is called genetic engineering. This technique is used in preparing some new therapies (monoclonal antibodies, blood component replacements for hemophilia, anti-inflammatory therapy for collagen diseases).

There are two types of gene therapy. Germ-line gene therapy introduces genes into reproductive cells (sperm and eggs) to participate in germination. Some scientists hope that it may eventually be possible to insert genes into embryos in hopes of correcting genetic abnormalities that can then be passed on to future generations. Most of the current work in applied gene therapy, however, has been in the realm of somatic therapy. In this type of gene therapy, therapeutic genes are inserted into tissue or cells to produce a naturally occurring protein or substance that is lacking or not functioning correctly in an individual.

Viral vectors

In both types of therapy, scientists need a mechanism to transport either an entire gene or a recombinant DNA to a cell's nucleus, where the chromosomes and DNA reside. In essence, vectors are molecular delivery trucks. One of the first and most widely used vectors to be developed were viruses, because they invade cells as part of their natural infection process. Viruses have the potential to be excellent vectors because they have a specific relationship with a host in that they colonize certain cell types and tissues in specific organs. As a result, vectors are chosen according to their attraction to certain cells and areas of the body.

One of the first classes of vectors used were retroviruses. Because these viruses are easily cloned (artificially reproduced) in the laboratory, scientists have studied them extensively and learned a great deal about their biologic action. They have also learned how to remove the genetic information that governs viral replication, thus reducing the chances of infection from the host vector.

Retroviruses work best in actively dividing cells, but most cells in a human body are relatively stable and do not often divide. As a result, these cells are used primarily for ex vivo (outside the body) manipulation. First, the cells are removed from a person's body, and the vector, or virus carrying the gene, is inserted into them. Next, the cells are placed into a nutrient culture where they grow and replicate. Once enough cells are gathered, they are returned to the body, usually by injection into the blood stream. Theoretically, as long as these cells survive, they will provide the desired therapy.

Another class of viruses, called adenoviruses, may also prove to be good gene vectors. These viruses effectively infect non-dividing cells in the body, where the desired gene product is then expressed naturally. In addition to being a more efficient approach to the problem of gene transportation, these viruses, which are known to cause respiratory infections, are more easily purified and stabilized than are retroviruses. The result is less liklihood of unintended viral infection. However, these viruses live for several days in the body, and there is some concern about the possibility of infecting other people with the viruses through sneezing or coughing. Other viral vectors include influenza viruses, Sindbis virus, and a herpes virus that infects nerve cells.

Scientists have also studied nonviral vectors. These vectors rely on the natural biologic process in which cells take up (or gather) macromolecules. One approach is to use liposomes, globules of fat produced by the body and taken up by cells. Scientists are also investigating the introduction of raw recombinant DNA by injecting it into the bloodstream or placing it on microscopic beads of gold injected into the skin using air pressure. Another possible vector under development is based on dendrimer molecules. A class of polymers (naturally occurring or artificial substances that have a high molecular weight and are formed by smaller molecules of the same or similar substances) is constructed in a laboratory by combining these smaller molecules. They have been used in manufacturing styrofoam, polyethylene cartons, and Plexiglas. In the laboratory, dendrimers have shown the ability to transport genetic material into human cells. They can also be designed to form an affinity for particular cell membranes by attaching to certain sugars and protein groups. Much additional research must be conducted before dendrimers can be used on a routine basis.

Viewpoints

On September 14, 1990, a four-year old girl who had a genetic disorder that prevented her body from producing a crucial enzyme became the first person to undergo gene therapy in the United States. Because her body could not produce adenosine deaminase (ADA), she had a weakened immune system, making her extremely susceptible to severe, life-threatening infections. W. French Anderson and colleagues at the National Institutes of Health's Clinical Center in Bethesda, Maryland, took white blood cells (which are crucial to proper immune system functioning) from the girl, inserted ADA-producing genes into them, and then transfused the cells back into the girl. Although the young girl continued to show an increased ability to produce ADA, debate arose as to whether the improvement resulted from the gene therapy or from an additional drug treatment she received.

Although gene therapy testing in humans has advanced rapidly, many questions surround its use. For example, some scientists are concerned that the therapeutic genes themselves may cause disease. Others fear that germ-line gene therapy may be used to control human development in ways not connected with disease, such as intelligence or physical appearance.

Nevertheless, a new era of gene therapy began as more and more scientists sought to conduct clinical trial (testing in humans) research in this area. In that same year, gene therapy was tested on persons with melanoma (skin cancer). The goal was to help them produce antibodies (disease fighting substances in the immune system) to battle the cancer.

The relative success of these experiments prompted a growing number of attempts at gene therapies designed to perform a variety of functions in the body. For example, a gene therapy for cystic fibrosis aims to supply a gene that alters cells, enabling people with cystic fibrosis to produce a specific protein to battle the disease. Another approach was used for people with brain cancer, in which the inserted gene was designed to make the cancer cells more likely to respond to drug treatment. A third gene therapeutic approach for people experiencing artery blockage, which can lead to strokes, induces the growth of new blood vessels (collateral circulation) near clogged arteries, thus ensuring relatively normal blood circulation.

In the United States, both nucleic acid-based (in vivo) treatments and cell-based (ex vivo) treatments are being investigated. Nucleic acid-based gene therapy uses vectors (such as viruses) to deliver modified genes to target cells. Cell-based gene therapy requires removal of cells from a person, genetically altering the cells and then reintroducing them into the body of the person being treated. Presently, gene therapies for the following diseases are being studied: cystic fibrosis (using adenoviral vector), HIV infection (cell-based), malignant melanoma (cell-based), Duchenne muscular dystrophy (cell-based), hemophilia B (cell-based), kidney cancer (cell-based), Gaucher disease (retroviral vector), breast cancer (retroviral vector), and lung cancer (retroviral vector). When a cell or individual is treated using gene therapy and successful incorporation of engineered genes has occurred, the cell or individual is said to be transgenic.

The medical establishment's contribution to transgenic research has been supported by increased government funding. In 1991, the U.S. government provided $58 million for gene therapy research, with increases in funding of $15-40 million dollars a year over the following four years. With fierce competition over the promise of societal benefits in addition to huge profits, large pharmaceutic corporations have moved to the forefront of transgenic research. In an effort to be first in developing new therapies, and armed with billions of dollars of research funds, such corporations are making impressive progress toward making gene therapy a viable reality in the treatment of once elusive diseases.

The Human Genome Project

Although great strides have been made in gene therapy in a relatively short time, its potential usefulness has been limited by lack of scientific data concerning the multitude of functions that genes control in the human body. For instance, it is now known that much genetic material is contained in non-coding regions. That is, they merely store information that may be used at different times in a cell's life cycle. Some of these large portions of the genome are involved in control and regulation of gene expression. Each individual cell in the body carries thousands of genes that have coding for proteins. Some experts estimate this number to be 150,000 genes. For gene therapy to advance to its full potential, scientists must discover the biologic role for each of these individual genes and identify the location on the DNA helix for each of the base pairs that comprise them.

To address this issue, the National Institutes of Health initiated the Human Genome Project in 1990. Led by Dr. James Watson (one of the co-discoverers of the chemical makeup of DNA) the project's 15-year goal is to map the entire human genome (a combination of the words gene and chromosome). A genome map would clearly identify the location of all genes as well as the more than three billion base pairs that comprise them. With a precise knowledge of gene locations and functions, scientists may one day be able to conquer or control diseases that have plagued humanity for centuries.

Scientists participating in the Human Genome Project have identified an average of one new gene a day, but many expect this rate of discovery to increase. In February of 2001, scientists published a rough draft of the complete human genome; the final complete sequence was published in 2003.

Some of the genes identified through this project include a gene that predisposes people to obesity; one associated with programmed cell death (apoptosis); a gene that guides HIV viral reproduction; and the genes of inherited disorders like Huntington's disease, amyotrophic lateral aclerosis (Lou Gehrig's disease), and some colon and breast cancers.

Professional implications

Diseases targeted for treatment by gene therapy

The potential scope of gene therapy is enormous. More than 4,200 diseases have been identified as resulting directly from non-functioning or abnormal genes, and countless others that may be partially influenced by a person's genetic makeup. Initial research has concentrated on developing gene therapies for diseases whose genetic origins have been established and for other diseases that can be cured or ameliorated by substances genes produce.

The following are examples of potential gene therapies. People with cystic fibrosis lack a gene needed to produce a salt-regulating protein. This protein regulates the flow of chloride into epithelial cells (the cells that line the inner and outer skin layers), that cover the air passages of the nose and lungs. Without this regulation, people with cystic fibrosis have a buildup of thick mucus in their lungs. In turn, this mucus makes these patients prone to lung infections and respiratory problems, and usually leads to death within the first 29 years of life. A gene therapy technique to correct this abnormality might employ an adenovirus to transfer a normal copy of what scientists call the cystic fibrosis transmembrane conductance regulator (CTRF) gene. The gene is introduced into a person by spraying it into the nose or lungs.

Familial hypercholesterolemia (FH) is also an inherited disease, resulting in the inability to process cholesterol properly, which leads to high levels of artery-clogging fat in the bloodstream of even the youngest family members. Persons with FH often suffer heart attacks and strokes because of blocked arteries. A gene therapy approach used to address FH is much more intricate than most gene therapies because it involves partial surgical removal of persons' livers (ex vivo transgene therapy). Corrected copies of a gene that acts to reduce cholesterol buildup are inserted into the liver sections, which are then transplanted back into the people.

Gene therapy has also been tested on persons with acquired immune difficiency syndrome (AIDS ). AIDS is caused by the human immunodeficiency virus (HIV), which weakens the body's immune system to the point that people with the condition are unable to fight off diseases such as pneumonia and cancer. In one approach, genes that produce specific HIV proteins have been altered to stimulate immune system functioning without causing the negative effects that a complete HIV molecule has on the immune system. These genes are then injected in a person's blood stream. Another approach to treating AIDS is to insert, via white blood cells, genes that have been genetically engineered to produce a receptor that would attract HIV and reduce its chances of replicating. These approaches are still primarily experimental.

Several cancers also have the potential to be treated with gene therapy. A therapy tested for melanoma, a progressive, agressive skin cancer, would introduce a gene with an anticancer protein called tumor necrosis factor (TNF) into test tube samples of a person's own cancer cells, which are then reintroduced into the person's body. In brain cancer, the approach is to insert a specific gene that increases the cancer cells' susceptibility to a common drug used in fighting the disease.

Gaucher disease is an inherited disease caused by a mutant gene that inhibits the production of an enzyme called glucocerebrosidase. Persons with Gaucher disease have enlarged livers (hepatomegaly) and spleens (splenomegaly). Clinical gene therapy trials will focus on inserting the gene for producing the missing enzyme.

Gene therapy is also being considered as an approach to solving a problem associated with a surgical procedure known as balloon angioplasty. In this procedure, a stent (a piece of tubular material resembling a straw) is used to open the clogged artery. However, in a "fail-safe" response to the trauma of the stent insertion, the body initiates a natural healing process that produces too many cells in the artery and results in restenosis or reclosing of the artery. The gene therapy approach to preventing this unwanted side effect is to cover the outside surfaces of an inserted stent with a soluble gel containing vectors for genes that may reduce an overactive healing response.

The future of gene therapy

Gene therapy seems elegantly simple in its concept: supply the human body with a gene that can correct a biologic malfunction causing a disease. However, there are many obstacles and some distinct questions concerning the viability of gene therapy. For example, viral vectors must be carefully controlled lest they infect a person with a viral disease. Some vectors, like retroviruses, can also enter normally functioning cells and interfere with natural biologic processes, possibly leading to other diseases. Other viral vectors, such as adenoviruses, are often recognized and destroyed by the immune system so their therapeutic effects are short-lived. Maintaining gene expression so that it performs its role properly after vector delivery is difficult. As a result, some therapies need to be repeated often to provide long-lasting benefits.

One of the most pressing issues, however, is gene regulation. Genes work in concert to regulate their functioning. In other words, several genes may play a part in turning other genes on and off. For example, certain genes work together to stimulate cell division and growth; but if these are not regulated, the inserted genes could cause tumor formation and cancer. Another difficulty is learning how to make the gene go into action only when needed. For the best and safest therapeutic effort, a specific gene should turn on, for example, when certain levels of a protein or enzyme are low and must be replaced. But the gene should also remain dormant when not needed to ensure that it does not oversupply a substance and disturb the body's delicate chemical balance.

One approach to gene regulation is to attach other genes that detect certain biologic activities and then react as a type of automatic off-and-on switch, regulating the activity of other genes according to biologic cues. Although still in the rudimentary stages, researchers are making progress in inhibiting some gene functioning by using a synthetic DNA to block gene transcriptions (the copying of genetic information). This approach may have applications for gene therapy.

The ethics of gene therapy

While gene therapy holds promise as a revolutionary approach for treating disease, ethical concerns over its use and ramifications have been expressed by scientists and lay people alike. For example, since much needs to be learned about how these genes actually work and their long-term effects, is it ethical to test these therapies on humans, in whom they could have a disastrous result? As with most clinical trials concerning new therapies, including many drugs, the people participating in these studies have usually not responded to more established therapies and are often so ill that the novel therapy is their only hope for long-term survival.

Another questionable outgrowth of gene therapy is that scientists could potentially manipulate genes to control traits in human offspring that are not related to health. For example, perhaps a gene could be inserted to ensure that a child would not be bald, a seemingly harmless goal. However, what if genetic manipulation was used to alter skin color, prevent homosexuality, or ensure good looks? If a gene is found that can enhance intelligence of children who are not yet born, will all members of society have access to the technology, or will it be so expensive that only the elite can afford it?

The Human Genome Project, which plays such an integral role for the future of gene therapy, also has social repercussions. If individual genetic codes can be determined, will such information be used against people? For example, will someone more susceptible to a disease have to pay higher insurance premiums or be denied health insurance altogether? Will employers discriminate between two potential employees, one with a healthy genome and the other with genetic abnormalities?

KEY TERMS

Cells— The smallest living units of the body that carry a full complement of the DNA, and which group together to form tissues and help the body perform specific functions.

Chromosome— Threadlike structures in a cell that carry most of the genetic material in the form of DNA and genes.

Clinical trial— The testing of a drug or some other type of therapy in a specific human population.

Clone— A cell or organism derived through asexual (without sex) reproduction, and which contains the identical genetic information of the parent cell or organism.

DNA (deoxyribonucleic acid)— The specific molecules that comprise chromosomes and genes.

Embryo— The earliest stage of development of the zygote before the human or animal is considered a fetus (which is usually the point at which the embryo takes on the basic physical form of its species). Embryos are formed in vivo (in utero) or in vitro (in a laboratory) in preparation for implantation.

Enzyme— Atypeofmoleculemadebycells that,when released, facilitates chemical reactions in the body.

Eugenics— A social movement in which the population of a society, country, or the world is to be improved by selective mating, controlling the passage of hereditary information.

Gene— A specific biologic component found in the cell nucleus that carries the instructions for the formation of an organism and its specific traits, such as eye or hair color.

Gene transcription— The process by which genetic information is copied from DNA to RNA, resulting in a specific protein formation.

Genetic engineering— The manipulation of genetic material to produce specific results in an organism.

Genetics— The study of hereditary traits passed on through genes.

Genome— The total set of genes carried by an individual or cell.

Genomics— The DNA which is found in the organism's total set of genes carried by an individual or cell and is passed on to offspring as information necessary for survival.

Germ-line gene therapy— The introduction of genes (natural or engineered) into reproductive cells or embryos to correct inherited genetic abnormalities that can cause disease by replication.

Liposome— Fat organelle made up of layers of lipids.

Macromolecule— A large molecule composed of thousands of atoms.

Nitrogen— An element that is a component of the base pairs in DNA.

Nucleus— The central part of a cell that contains most of its genetic material, including chromosomes and DNA.

Protein— Macromolecule made up of long sequences of amino acids. Proteins comprise the dry weight of most cells and are involved in structures, hormones, and enzymes in muscle contraction, immunological response, and many other functions essential to life.

Somatic gene therapy— The introduction of genes into tissue or cells to treat a genetic disease in an individual.

Vector— Something used to transport genetic information to a cell.

Some of these concerns can be traced back to the eugenics movement that was popular in the first half of the twentieth century. This genetic philosophy was a societal movement that encouraged people with so-called positive traits to reproduce while those with less desirable traits were sanctioned from having children. Eugenics was used to pass strict immigration laws in the United States, barring less suitable people from entering the country lest they reduce the quality of the country's collective gene pool. Probably the most notorious example of eugenics in action was the rise of Nazism in Germany, which fostered the Eugenic Sterilization Law of 1933. The law required sterilization for those with certain disabilities and even for some persons who were simply deemed to be unattractive. To ensure that this novel science is not abused, many governments have established organizations specifically for overseeing the development of gene therapy. In the United States, the Food and Drug Administration and the National Institutes of Health require scientists to take a precise series of steps and meet stringent requirements before approving clinical trials.

In fact, gene therapy has been immersed in more controversy and is surrounded by more scrutiny from both the health care and ethics communities than most other technologies (except, perhaps, for cloning) that have the potential to substantially change society. Despite the health and ethical questions surrounding gene therapy, the field will continue to grow and is likely to change medicine more quickly than any previous medical advancement.

Resources

BOOKS

Burdette, Walter J. The Basis for Gene Therapy. Springfield, IL, Charles C Thomas, 2001.

Gomez-Navarro, Jesus, Guadalupe Bilbao, and David T. Curiel. "Gene therapy." In Cecil Textbook of Medicine, 21st ed., edited by Goldman, Lee and Bennett, J. Claude. Philadelphia: W.B. Saunders, 2000, 140-143.

Hengge, Ulrich R., and Beatrix Volc-Platzer. The Skin and Gene Therapy. New York: Springer Verlag, 2000.

Huard, Johnny, and Freddie Fu. Gene Therapy and Tissue Engineering in Orthopaedic and Sports Medicine. Boston: Birkhauser, 2000.

Lemoine, Nicholas R., and Richard G. Vile. Understanding Gene Therapy. New York: Springer Verlag, 2000.

Needleman, Robert D. "Fetal growth and development." In Nelson Textbook of Pediatrics, 16th ed. edited by Richard E. Behrman et al., Philadelphia: Saunders, 2000, 27-30.

Valle, David. "Treatment and prevention of genetic disease." In Harrison's Principles of Internal Medicine, 14th ed. edited by Anthony S. Fauci, et al. New York: McGraw-Hill, 1998, 403-411.

Walther, Wolfgang, and Ulrike Stein. Gene Therapy of Cancer: Methods and Protocols. Totowa, NJ: Humana Press, 2000.

PERIODICALS

Gottlieb, S. "Gene therapy shows promise for hemophilia." British Medical Journal 322 no.7300 (2001): 1442A-1443.

Gray, S.G. "Pill-based gene therapy." Trends in Genetics 17 no.7 (2001): 380-384.

McKay, D. "Restoring sight by gene therapy." Trends in Biotechnology, 19 no.7 (2001): 243-246.

Newman, C.M., Lawrie, A., Brisken, A.F., Cumberland, D.C. "Ultrasound gene therapy: on the road from concept to reality." Echocardiography 18 no.4 (2001): 339-347.

Savulescu, J. "Harm, ethics committees and the gene therapy death." Journal of Medical Ethics 27 no.3 (2001): 148-150.

Verma, I.M. "Ombudsman or Hotline for Gene Therapy Clinical Trials?" Molecular Therapeutics 3 no.6 (2001): 817-818.

ORGANIZATIONS

American Academy of Family Physicians. 11400 Tomahawk Creek Parkway, Leawood, KS 66211-2672, (913) 906-6000, 〈http://www.aafp.org〉.

American Society of Gene Therapy. 611 East Wells Street, Milwaukee, WI 53202, (414) 278-1341, (414) 276-3349. 〈http://www.asgt.org〉.

World Health Organization. 20 Avenue Appia, 1211 Geneva 27, Switzerland, +41 (22) 791 4140, +41 (22) 791 4268. 〈http://www.who.int/gtb〉.

OTHER

American Civil Liberties Union. 〈http://www.aclu.org/issues/aids/docket98.html〉.

Association of American Medical Colleges. 〈http://www.aamc.org/newsroom/reporter/june2000/view.htm〉.

Human Genome Project Information. 〈http://www.ornl.gov/hgmis/medicine/genetherapy.html〉.

National Cancer Institute. 〈http://cancernet.nci.nih.gov/clinpdq/therapy/Questions_and_Answers_About_Gene_Therapy.html〉.

Public Broadcasting System (animation). 〈http://www.pbs.org/wnet/innovation/show1/html/animation2.html〉.

University of Pennsylvania. 〈http://www.med.upenn.edu/ihgt/info/whatisgt.html〉.

U.S. Food and Drug Administration. 〈http://www.fda.gov/fdac/features/2000/gene.html〉.

Vanderbilt University. 〈http://www.mc.vanderbilt.edu/gcrc/gene〉.

Gene Therapy

The biological basis of gene therapy

Viral vectors

The history of gene therapy

Diseases targeted for treatment by gene therapy

The future of gene therapy

Ethical considerations in gene therapy

Resources

Gene therapy refers to the deliberate introduction of genes into an organism. The intent of gene therapy is to correct a genetic defect or alleviate the symptoms of a genetically-determined disease when the introduced gene is expressed and its product is produced.

Gene therapy is the name applied to the treatment of inherited diseases by corrective genetic engineering of the dysfunctional genes. It is part of a broader field called genetic medicine, which involves the screening, diagnosis, prevention and treatment of hereditary conditions in humans. The results of genetic screening can pinpoint a potential problem to which gene therapy can sometimes offer a solution.

Genes represent the genetic material that organisms pass on from one generation to the next. Therefore, genes are responsible for controlling hereditary traits and provide the basic biological code or blueprint for living organisms. Genes produce protein such as hair and skin as well as proteins that are important for the proper functioning of organs. Mutated or defective genes often cause disease. The purpose of gene therapy is to replace a defective gene with a normal copy of the same gene in attempt to restore function.

Somatic gene therapy introduces a normal gene into tissues or cells to treat an individual that has an abnormal gene. Germline gene therapy inserts genes into reproductive cells (the egg or the sperm) or into embryos to correct genetic defects that could be passed on to future generations. Germline gene therapy differs from somatic gene therapy in that germline integration of a gene will ideally correct every progenitor cell that differentiates from the germ cell. Somatic gene therapy involves integrating corrected genes into cell and tissues that are fully differentiated or mature. An example of the latter is the efforts to treat cystic fibrosis in children or adults by the introduction of the normal gene that specifies a protein that forms a chloride transport channel through the epithelial cell membrane (in cystic fibrosis, the impaired transport of chloride causes the buildup of sticky mucus in the lungs, which can lead to bacterial infection).

Initially conceived as an approach for treating inherited diseases like cystic fibrosis and Huntington disease, the scope of potential gene therapies has grown to include treatments for cancers, arthritis, and infectious diseases. Although gene therapy testing in humans has rapidly advanced, in general, the field of gene therapy has proven to be problematic and complicated by a variety of ethical issues. For example, some scientists are concerned that the integrating genes into the human genome may cause disease. This concern has arisen because of evidence that randomly integrating corrected genes might disrupt other genes in the genome and if the disrupted gene is a tumor suppressor gene, cancer may develop. Others fear that germline gene therapy may be used to control human development in ways not connected with disease, like intelligence or appearance.

The biological basis of gene therapy

Gene therapy has grown out of the field of molecular biology. Life begins with a single cell, the basic building block of all multicellular organisms. Humans, for instance, are made up of trillions of cells, that make up tissues that form into organs. Each cell type can perform a specific function. Within the cells nucleus (the center part of a cell that regulates its chemical functions) are pairs of chromosomes. These threadlike structures are made up of

DNA (deoxyribonucleic acid), which carries the blueprint of life in the form of codes, or genes, that are interspersed throughout the DNA sequence.

A DNA molecule looks like a twisted ladder. The rungs of these represent bonds between each letter of the DNA sequence called base pairs. Base pairs are made up of nitrogenous molecules. Thousands of these base pairs, or DNA sequences, can make up a single gene, specifically defined as a segment of the chromosome. The gene, or combination of genes formed by these base pairs ultimately direct an organisms growth and characteristics through the production of certain proteins, which are important for many biochemical functions.

Scientists have long known that defects in genes present within cells can cause inherited diseases such as cystic fibrosis, sickle-cell anemia, and hemophilia. Similarly, a gain or a loss of an entire chromosome can cause diseases such as Down syndrome or Turners syndrome. As the study of genetics advanced, however, scientists learned that an altered genetic sequence can also make people more susceptible to develop diseases making these individuals predisposed to having atherosclerosis, cancer, or schizophrenia. These diseases have a genetic component, but are also influenced by environmental factors (like diet and lifestyle). The objective of gene therapy is to treat diseases by introducing corrected genes into the body to replace a missing or dysfunctional protein. The inserted genes can be naturally-occurring genes that produce the desired effect or may be genetically engineered (or altered) genes.

Scientists have known how to manipulate the structure of a gene in the laboratory since the early 1970’s through a process called gene splicing. The process involves cutting a sequence of the genome with restriction enzymes, or proteins that act like molecular scissors. The ends where the DNA has been cut are sticky in the sense that they will easily bind to another sequence of DNA that was cut with the same enzyme. A DNA sequence and a gene sequence to be integrated in the DNA sequence can both be cut with the same type of enzyme and their ends will stick together. The new DNA sequence will now have the gene inserted into it. The resulting product is called genetic engineered recombinant DNA.

There are basically two types of gene therapy. Germ-line gene therapy introduces genes into reproductive cells (sperm and eggs) or into embryos in order to correct genetic defects that could be passed on to future generations. Most of the current research, however, has been in the applications of somatic cell gene therapy. In this type of gene therapy, therapeutic genes are inserted into tissue or cells to produce a naturally occurring protein or substance that is lacking or not functioning correctly in an individual patient. The main downside to this approach is that as each corrected cell dies, the therapeutic effects from gene therapy are lessened.

Viral vectors

In both types of therapy, scientists need something to transport either the entire gene or a recombinant DNA to the cells nucleus, where the DNA is located. In essence, vectors are molecular delivery trucks. One of the first and most popular vectors developed was viral vectors, or vectors made of viruses because they invade cells as part of a natural infection process. Viruses were originally considered the most ideal vector because they have a specific relationship with the host in that they can infect specific cell types or tissues. As a result, vectors are chosen according to their affinity for certain cells and areas of the body.

One of the first viral vectors used was the retro-virus. Because these viruses are easily cloned (artificially reproduced) in the laboratory, scientists have studied them extensively and learned a great deal about their biological characteristics. They have also learned how to remove the genetic information that governs viral replication, thus reducing the chances of multiple rounds of infection. Additionally, many of the proteins from these viruses that can cause an immune response can be removed.

Retroviruses work best in actively dividing cells, but most of the cells in the body particularly those that are fully differentiated are relatively stable and do not divide often. As a result, these cells are used primarily for ex vivo (outside the body) manipulation. First, the cells are removed from the patient’s body, and the virus, or vector, carrying the gene is inserted into them. Next, the cells are placed into a nutrient culture where they grow and replicate. Once enough cells are gathered, they are returned to the body, usually by injection into the blood stream. Theoretically, as long as these cells survive, they can have therapeutic potential.

Another class of viruses, called the adenoviruses, have proven to be good gene vectors in certain cases. These cells can effectively infect nondividing cells in the body, where the desired gene product is then expressed. These viruses, which cause respiratory tract infections, are more easily purified and more stable than retroviruses, resulting in less chance of an unwanted viral infection. These viruses live for several days in the body and can have potentially life-threatening complications related to immune cell responses. Other viral vectors include influenza viruses (that causes the flu), Sindbis virus, and a herpes virus that infects nerve cells. Each of these vectors can be modified to minimize the risk of causing disease or immune cell responses.

Scientists have also developed nonviral vectors. These vectors rely on the natural biological process in which cells uptake (or gather) macromolecules (large molecules). One approach is to use liposomes, or globules of fat produced by the body and taken up by cells. Scientists are also investigating the introduction of recombinant DNA by directly injecting it into the bloodstream or placing it on microscopic beads of gold shot into the skin with a gene gun. Another possible vector under development is based on den-drimer molecules. This is a class of polymers or naturally occurring or artificial substances that have a high molecular weight and are formed by smaller molecules of the same or similar substances. They have been used in manufacturing Styrofoam, polyethylene cartons, and Plexiglass.

In the laboratory, dendrimers have shown the ability to transport genetic material into human cells. They can also be designed with a high affinity for the membrane of a cell by attaching sugars and protein groups to it. The use of liposomes (spherical bags whose membrane is comprise of lipids) as a carrier of DNA has shown promise, especially since other molecules can be incorporated into the lipid membrane that help target to liposome to a specific site in the body. As of 2006, liposome use is still at the experimental stage. However, the research has so far been very promising.

Another approach to gene therapy introduces DNA that can bind to a target stretch of DNA. The binding prevents the expression of the target sequence, which typically produces a product that contributes to the disease. This strategy is known as antisense therapy or gene silencing. As of 2006, this tact has shown great experimental promise.

The history of gene therapy

In the early 1970s, scientists proposed what they called “gene surgery” for treating inherited diseases caused by defective genes. In 1983, a group of scientists from Baylor College of Medicine in Houston, Texas, proposed that gene therapy could one day be a viable approach for treating Lesch-Nyhan disease, a rare neurological disorder. The scientists conducted experiments in which an enzyme-producing gene for correcting the disease was injected into a group of cells. The scientists theorized the cells could then be injected into people with Lesch-Nyhan disease.

As the science of genetics advanced throughout the 1980s, gene therapy gained an established foothold in the minds of medical scientists as a viable approach to treatments for specific diseases. However, its promises were more than what it could deliver. One of the major impetuses in the growth of gene therapy was an increasing ability to identify the genetic abnormalities that cause inherited diseases. Interest grew as further studies showed that specific genetic defects in one or more genes occurred in successive generations of certain family members who suffered from diseases like intestinal cancer, manic-depression, Alzheimer disease, heart disease, diabetes, and many more. Although the genes may not be the sole cause of the disease in all cases, they may make certain individuals more susceptible to developing the disease because of environmental influences, such as smoking, pollution, and stress. In fact, many scientists believe that all diseases have a genetic component.

On September 14, 1990, a four-year old girl suffering from a genetic disorder that prevented her body from producing a crucial enzyme became the first person to undergo gene therapy in the United States. Since her body could not produce adenosine deaminase (ADA), she had a weakened immune system, making her extremely susceptible to severe, life-threatening infections. W. French Anderson and colleagues at the National Institutes of Health’s Clinical Center in Bethesda, Maryland, took white blood cells (which are crucial for proper immune system functioning) from the girl, inserted ADA producing genes into them, and then transferred the cells back into the patient. Although the young girl continued to show an increased ability to produce ADA, debate arose as to whether the improvement resulted from the gene therapy or from an additional drug treatment she received.

Nevertheless, a new era of gene therapy began as more and more scientists sought to conduct clinical trials in this area. In that same year, gene therapy was tested on patients suffering from melanoma (skin cancer). The goal was to help them produce antibodies (disease fighting substances in the immune system) to battle the cancer.

These experiments have spawned a growing number of attempts to refine develop new gene therapies. For example, gene therapy for cystic fibrosis, a disease that affects the airways, is being developed. However, due to the complications involved in penetrating the natural barriers that impedes viral entry into the airways, it is unlikely that currently used vectors for cystic fibrosis gene therapy represent a plausible approach. Modifications of these vectors by adding compounds that naturally bind to areas on the outermost membranes of the lung and gain entrance into these tissues are currently being investigated. Another approach was developed for treating brain cancer patients, in which the inserted gene was designed to make the cancer cells more likely to respond to drug treatment. Additionally, gene therapy for patients suffering from artery blockage, which can lead to strokes, that induces the growth of new blood vessels near clogged arteries improving normal blood circulation is also being investigated.

In the United States, both DNA-based (in vivo ) treatments and cell-based (ex vivo ) treatments are being investigated. DNA-based gene therapy uses vectors (like viruses) to deliver modified genes to target cells. Cell-based gene therapy techniques remove cells from the patient, which are genetically altered and then reintroduce them to the patient’s body. Presently, gene therapies for the following diseases are being developed: cystic fibrosis (using adeno-viral vector), HIV infection (cell-based), malignant melanoma (cell-based), kidney cancer (cell-based), Gaucher’s disease (retroviral vector), breast cancer (retroviral vector), and lung cancer (retroviral vector).

The medical has contributed to transgenic research that is supported by government funding. In 1991, the U.S. government provided $58 million for gene therapy research, with increases of $15-40 million dollars a year over the following four years. With fierce competition over the promise of major medical benefit in addition to huge profits, large pharmaceutical corporations moved to the forefront of transgenic research.

Diseases targeted for treatment by gene therapy

The potential scope of gene therapy is enormous. More than 4,200 diseases have been identified that result directly from defective genes. People suffering from cystic fibrosis lack a gene needed to produce a salt-regulating protein. This protein regulates the flow of chloride into epithelial cells, which cover the air passages of the nose and lungs. Without this regulation, cystic fibrosis patients suffer from a buildup of thick mucus, which can cause lung infections and respiratory problems, which usually leads to death within the first 30 years of life. A gene therapy technique to correct this defect might employ an adenovirus to transfer a normal copy of what scientists call the cystic fibrosis transmembrane conductance regulator, or CTRF, gene. The gene is introduced into the patient by spraying it into the nose or lungs.

Familial hypercholesterolemia is also an inherited disease, resulting in the inability to process cholesterol properly, which leads to high levels of artery-clogging fat in the blood stream. Patients often suffer heart attacks and strokes because of blocked arteries. A gene therapy approach that was still being investigated as of 2006 involves partially and surgically removing the patient’s liver (ex vivo transgene therapy). Corrected copies of a gene that serve to reduce cholesterol build-up are inserted into the liver sections, which are then transplanted back into the patient.

Gene therapy has also been tested on AIDS patients. AIDS is caused by the human immunodeficiency virus (HIV), which weakens the body’s immune system to the point that sufferers are unable to fight off diseases such as pneumonia. An approach to treat AIDS is to insert genes into a patient’s bloodstream that have been genetically engineered to produce a receptor that would attract HIV and reduce its chances of replicating.

Several cancers also have the potential to be treated with gene therapy. A therapy that is currently being tested for the treatment of melanoma (a form of skin cancer), involves introducing a gene with an anticancer protein called tumor necrosis factor (TNF) into test tube samples of the patient’s own cancer cells, which are then reintroduced into the patient. In brain cancer, the approach is to insert a specific gene that increases the cancer cells susceptibility to a common drug used in fighting the disease. Gene therapy can also be used to treat diseases that involve dysfunctional enzymes. For example, Gaucher’s disease is an inherited disease caused by a mutant gene that inhibits the production of an enzyme called glucocerebrosi-dase. Gaucher patients have enlarged livers and spleens and eventually their bones fall apart. Clinical gene therapy trials focus on inserting the gene for producing this enzyme.

Gene therapy is also being considered as an approach to solving a problem associated with a surgical procedure known as angioplasty. In this procedure, a type of tubular scaffolding is used to open a clogged artery. However, in response to the trauma of the scaffold insertion, the body often initiates a natural healing process resulting in restenosis, or reclosing of the artery. The gene therapy approach to preventing this unwanted side effect is to cover the outside of the stents with a soluble gel. This gel is designed to contain vectors for genes that would reduce restenosis.

The future of gene therapy

There are many obstacles and ethical questions concerning gene therapy. For example, some retrovir-usal vectors, can also enter normal cells and interfere with the natural biological processes, possibly leading to other diseases. Other viral vectors, like the adeno-viruses, are often recognized and destroyed by the immune system so their therapeutic effects are shortlived. One of the primary limitations in gene therapy is that delivering a gene using a viral vector that can only undergo one round of infection (making it safer) may provide only temporary therapeutic value that lasts only as long as the corrected gene is expressed. As a result, some therapies need to be repeated often to provide long-lasting benefits.

One of the most pressing issues, however, involves gene regulation. Several genes may play a role in turning other genes on and off. For example, certain genes work together to stimulate cell division and growth, but if these are not regulated, the inserted genes could cause unregulated cell growth leading to the formation of a tumor. Another difficulty is learning how to make the gene be expressed in a regulated way. A specific gene should turn on, for example, when certain levels of a protein or enzyme are not sufficiently meeting cellular demands. This type of controlled regulation of gene expression for these delivered genes is very difficult to achieve.

Ethical considerations in gene therapy

While gene therapy holds promise as a revolutionary approach to treating disease, ethical concerns over its use and ramifications have been expressed. For example, it is difficult to determine the long-term effect of exposure to viral vectors and the effects these engineered viruses have on the human genome.

As the technology develops and more mainstream applications become possible, it is likely that medically unrelated genetic traits might be the target of manipulation. For example, perhaps a gene could be introduced that prevents balding in males. Or what if

KEY TERMS

Cells —The smallest living units of the body which together form tissues.

Chromosomes —he structures that carry genetic information in the form of DNA. Chromosomes are located within every cell and are responsible for directing the development and functioning of all the cells in the body.

Clinical trial —The testing of a drug or some other type of therapy in a specific population of patients.

Clone —A cell or organism derived through asexual reproduction, which contains the identical genetic information of the parent cell or organism.

DNA —Deoxyribonucleic acid; the genetic material in a cell.

Enzyme —Biological molecule, usually a protein, which promotes a biochemical reaction but is not consumed by the reaction.

Eugenics —A social movement in which the population of a society, country, or the world is to be improved by controlling the passing on of hereditary information through selective breeding.

Gene —A discrete unit of inheritance, represented by a portion of DNA located on a chromosome. The gene is a code for the production of a specific kind of protein or RNA molecule, and therefore for a specific inherited characteristic.

Gene transcription —The process by which genetic information is copied from DNA to RNA.

Genetic engineering —The manipulation of genetic material to produce specific results in an organism.

Germ-line gene therapy —The introduction of genes into reproductive cells or embryos to correct inherited genetic defects that can cause disease.

Macromolecules —A large molecule composed of thousands of atoms.

Nucleus —The central part of a cell that contains most of its genetic material, including chromosomes and DNA.

Protein —Macromolecules made up of long sequences of amino acids.

Somatic gene therapy —The introduction of genes into tissue or cells to treat a genetic related disease in an individual.

Vectors —A molecular device to transport genes or DNA sequences into a cell or organ.

genetic manipulation was used to alter skin color, prevent homosexuality, or to enhance physical attractiveness and intelligence? Will this only be available to the rich? Gene therapy has been surrounded by more controversy and scrutiny in both scientists and the general public than many other technologies.

As with every new medical technique, there are many potential dangers and unpredictable factors with gene therapy, which make its practical application risky. Even though every precaution is taken to prevent accidents, they sometimes do occur. Jesse Gelsinger, a 17 year-old boy suffering from the disease ornithine transcarbamylase (OTC) deficiency became the first tragic victim of gene therapy and died on September 17, 1999. He had volunteered to test the potential use of gene therapy in the treatment of OTC in young babies. His therapy consisted of an infusion of corrective genes, encased in a weakened adenovirus vector. Gelsinger suffered an unexpected chain reaction that resulted in his early death from multiple organ system failure. The reason for his extreme reaction to the treatment is suspected to have been an overwhelming inflammatory response to the viral vector, though the reason why is not known. Subsequent investigations revealed the deaths of six other gene therapy patients, some prior to Gelsinger, who were undergoing trials for the use of gene therapy in the treatment of heart conditions. Unlike Gelsinger, these latter six victims are thought to have died from complications stemming from their underlying illnesses rather than the gene therapy itself.

In 2006, Italian researchers were successful in preventing the immune system from rejecting introduced DNA, which had long been a problem in gene therapy. With further refinement, the advancement may enable gene therapy to become routinely successful.

Resources

BOOKS

Merkeley, Ward. A Model for Gene Therapy. Philadelphia: Xlibris Corporation, 2005.

Panno, Joseph. Gene Therapy: Treating Disease by Repairing Genes. New York: Facts on File, 2004.

PERIODICALS

Kaiser J. “RAC Hears a Plea for Resuming Trials, Despite Cancer Risk.” Science. 299:5609 (2003):991.

Kasuya H, S. Nomoto, A. Nakao. “The Potential of Gene Therapy in the Treatment of Pancreatic Cancer.” Drugs Today 38(7) (2002):457-64.

David Petechuk
Brian Cobb

Gene therapy

Gene therapy is a rapidly growing field of medicine in which genes are introduced into the body to treat diseases. Genes control heredity and provide the basic biological code for determining a cell's specific functions. Gene therapy seeks to provide genes that correct or supplant the disease-controlling functions of cells that are not, in essence, doing their job. Somatic gene therapy introduces therapeutic genes at the tissue or cellular level to treat a specific individual. Germ-line gene therapy inserts genes into reproductive cells or possibly into embryos to correct genetic defects that could be passed on to future generations. Initially conceived as an approach for treating inherited diseases, like cystic fibrosis and Huntington's disease, the scope of potential gene therapies has grown to include treatments for cancers, arthritis, and infectious diseases. Although gene therapy testing in humans has advanced rapidly, many questions surround its use. For example, some scientists are concerned that the therapeutic genes themselves may cause disease. Others fear that germ-line gene therapy may be used to control human development in ways not connected with disease, like intelligence or appearance.

The biological basis of gene therapy

Gene therapy has grown out of the science of genetics or how heredity works. Scientists know that life begins in a cell, the basic building block of all multicellular organisms. Humans, for instance, are made up of trillions of cells, each performing a specific function. Within the cell's nucleus (the center part of a cell that regulates its chemical functions) are pairs of chromosomes . These threadlike structures are made up of a single molecule of DNA (deoxyribonucleic acid), which carries the blueprint of life in the form of codes, or genes, that determine inherited characteristics.

A DNA molecule looks like two ladders with one of the sides taken off both and then twisted around each other. The rungs of these ladders meet (resulting in a spiral staircase-like structure) and are called base pairs. Base pairs are made up of nitrogen molecules and arranged in specific sequences. Millions of these base pairs, or sequences, can make up a single gene, specifically defined as a segment of the chromosome and DNA that contains certain hereditary information. The gene, or combination of genes formed by these base pairs ultimately direct an organism's growth and characteristics through the production of certain chemicals, primarily proteins, which carry out most of the body's chemical functions and biological reactions.

Scientists have long known that alterations in genes present within cells can cause inherited diseases like cystic fibrosis, sickle-cell anemia, and hemophilia . Similarly, errors in the total number of chromosomes can cause conditions such as Down syndrome or Turner syndrome . As the study of genetics advanced, however, scientists learned that an altered genetic sequence can also make people more susceptible to diseases, like atherosclerosis, cancer , and even schizophrenia . These diseases have a genetic component, but are also influenced by environmental factors (such as diet and lifestyle). The objective of gene therapy is to treat diseases by introducing functional genes into the body to alter the cells involved in the disease process by either replacing missing genes or providing copies of functioning genes to replace nonfunctioning ones. The inserted genes can be naturally occurring genes that produce the desired effect or may be genetically engineered (or altered) genes.

Scientists have known how to manipulate a gene's structure in the laboratory since the early 1970s through a process called gene splicing. The process involves removing a fragment of DNA containing the specific genetic sequence desired then inserting it into the DNA of another gene. The resultant product is called recombinant DNA and the process is genetic engineering.

There are basically two types of gene therapy. Germline gene therapy introduces genes into reproductive cells (sperm and eggs) or someday possibly into embryos in hopes of correcting genetic abnormalities that could be passed on to future generations. Most of the current work in applying gene therapy, however, has been in the realm of somatic gene therapy. In this type of gene therapy, therapeutic genes are inserted into tissue or cells to produce a naturally occurring protein or substance that is lacking or not functioning correctly in an individual patient.

Viral vectors

In both types of therapy, scientists need something to transport either the entire gene or a recombinant DNA to the cell's nucleus, where the chromosomes and DNA reside. In essence, vectors are molecular delivery trucks. One of the first and most popular vectors developed were viruses because they invade cells as part of the natural infection process. Viruses have the potential to be excellent vectors because they have a specific relationship with the host in that they colonize certain cell types and tissues in specific organs. As a result, vectors are chosen according to their attraction to certain cells and areas of the body.

One of the first vectors used was the retrovirus. Because these viruses are easily cloned (artificially reproduced) in the laboratory, scientists have studied them extensively and learned a great deal about their biological action. They have also learned how to remove the genetic information which governs viral replication, thus reducing the chances of infection.

Retroviruses work best in actively dividing cells, but cells in the body are relatively stable and do not divide often. As a result, these cells are used primarily for ex vivo (outside the body) manipulation. First, the cells are removed from the patient's body, and the virus, or vector, carrying the gene is inserted into them. Next, the cells are placed into a nutrient culture where they grow and replicate. Once enough cells are gathered, they are returned to the body, usually by injection into the blood stream. Theoretically, as long as these cells survive, they will provide the desired therapy.

Another class of viruses, called the adenoviruses, may also prove to be good gene vectors. These viruses can effectively infect nondividing cells in the body, where the desired gene product is then expressed naturally. In addition to being a more efficient approach to gene transportation, these viruses, which cause respiratory infections, are more easily purified and made stable than retroviruses, resulting in less chance of an unwanted viral infection. However, these viruses live for several days in the body, and some concern surrounds the possibility of infecting others with the viruses through sneezing or coughing. Other viral vectors include influenza viruses, Sindbis virus, and a herpes virus that infects nerve cells.

Scientists have also delved into nonviral vectors. These vectors rely on the natural biological process in which cells uptake (or gather) macromolecules. One approach is to use liposomes, globules of fat produced by the body and taken up by cells. Scientists are also investigating the introduction of raw recombinant DNA by injecting it into the bloodstream or placing it on microscopic beads of gold shot into the skin with a "gene-gun." Another possible vector under development is based on dendrimer molecules. A class of polymers (naturally occurring or artificial substances that have a high molecular weight and formed by smaller molecules of the same or similar substances), is "constructed" in the laboratory by combining these smaller molecules. They have been used in manufacturing Styrofoam, polyethylene cartons, and Plexiglass. In the laboratory, dendrimers have shown the ability to transport genetic material into human cells. They can also be designed to form an affinity for particular cell membranes by attaching to certain sugars and protein groups.

The history of gene therapy

In the early 1970s, scientists proposed "gene surgery" for treating inherited diseases caused by faulty genes. The idea was to take out the disease-causing gene and surgically implant a gene that functioned properly. Although sound in theory, scientists, then and now, lack the biological knowledge or technical expertise needed to perform such a precise surgery in the human body.

However, in 1983, a group of scientists from Baylor College of Medicine in Houston, Texas, proposed that gene therapy could one day be a viable approach for treating Lesch-Nyhan disease, a rare neurological disorder. The scientists conducted experiments in which an enzyme-producing gene (a specific type of protein) for correcting the disease was injected into a group of cells for replication. The scientists theorized the cells could then be injected into people with Lesch-Nyhan disease, thus correcting the genetic defect that caused the disease.

As the science of genetics advanced throughout the 1980s, gene therapy gained an established foothold in the minds of medical scientists as a promising approach to treatments for specific diseases. One of the major reasons for the growth of gene therapy was scientists' increasing ability to identify the specific genetic malfunctions that caused inherited diseases. Interest grew as further studies of DNA and chromosomes (where genes reside) showed that specific genetic abnormalities in one or more genes occurred in successive generations of certain family members who suffered from diseases like intestinal cancer, manic-depression, Alzheimer's disease, heart disease, diabetes, and many more. Although the genes may not be the only cause of the disease in all cases, they may make certain individuals more susceptible to developing the disease because of environmental influences, like smoking, pollution, and stress. In fact, some scientists theorize that all diseases may have a genetic component.

On September 14, 1990, a four-year-old girl with a genetic disorder that prevented her body from producing a crucial enzyme became the first person to undergo gene therapy in the United States. Because her body could not produce adenosine deaminase (ADA), she had a weakened immune system, making her extremely susceptible to severe, life-threatening infections. W. French Anderson and colleagues at the National Institutes of Health's Clinical Center in Bethesda, Maryland, took white blood cells (which are crucial to proper immune system functioning) from the girl, inserted ADA producing genes into them, and then transfused the cells back into the patient. Although the young girl continued to show an increased ability to produce ADA, debate arose as to whether the improvement resulted from the gene therapy or from an additional drug treatment she received.

Nevertheless, a new era of gene therapy began as more and more scientists sought to conduct clinical trial (testing in humans) research in this area. In that same year, gene therapy was tested on patients with melanoma (skin cancer). The goal was to help them produce antibodies (disease fighting substances in the immune system) to battle the cancer.

These experiments have spawned an ever growing number of attempts at gene therapies designed to perform a variety of functions in the body. For example, a gene therapy for cystic fibrosis aims to supply a gene that alters cells, enabling them to produce a specific protein to battle the disease. Another approach was used for brain cancer patients, in which the inserted gene was designed to make the cancer cells more likely to respond to drug treatment. Gene therapy for patients who have artery blockage, which can lead to strokes, induces the growth of new blood vessels near clogged arteries, thus ensuring normal blood circulation.

Currently, there are a host of new gene therapy agents in clinical trials. In the United States, both nucleic acid-based (in vivo) treatments and cell-based (ex vivo) treatments are being investigated. Nucleic acid-based gene therapy uses vectors (like viruses) to deliver modified genes to target cells. Cell-based gene therapy techniques remove cells from the patient in order to genetically alter them then reintroduce them to the patient's body. Presently, gene therapies for the following diseases are being developed: cystic fibrosis (using adenoviral vector), HIV infection (cell-based), malignant melanoma (cell-based), Duchenne muscular dystrophy (cell-based), hemophilia B (cell-based), kidney cancer (cell-based), Gaucher disease (retroviral vector),breast cancer (retroviral vector), and lung cancer (retroviral vector). When a cell or individual is treated using gene therapy and successful incorporation of engineered genes has occurred, the cell or individual is said to be transgenic.

The medical establishment's contribution to transgenic research has been supported by increased government funding. In 1991, the U.S. government provided $58 million for gene therapy research, with increases in funding of $15–40 million dollars a year over the following four years. With fierce competition over the promise of societal benefit in addition to huge profits, large pharmaceutical corporations have moved to the forefront of transgenic research. In an effort to be first in developing new therapies, and armed with billions of dollars of research funds, such corporations are making impressive strides toward making gene therapy a viable reality in the treatment of once elusive diseases.

Diseases targeted for treatment by gene therapy

The potential scope of gene therapy is enormous. More than 4,200 diseases have been identified as resulting directly from abnormal genes, and countless others that may be partially influenced by a person's genetic makeup. Initial research has concentrated on developing gene therapies for diseases whose genetic origins have been established and for other diseases that can be cured or ameliorated by substances genes produce.

The following are examples of potential gene therapies. People suffering from cystic fibrosis lack a gene needed to produce a salt-regulating protein. This protein regulates the flow of chloride into epithelial cells, (the cells that line the inner and outer skin layers) which cover the air passages of the nose and lungs. Without this regulation, patients with cystic fibrosis build up a thick mucus that makes them prone to lung infections. A gene therapy technique to correct this abnormality might employ an adenovirus to transfer a normal copy of what scientists call the cystic fibrosis transmembrane conductance regulator, or CTRF, gene. The gene is introduced into the patient by spraying it into the nose or lungs.

Familial hypercholesterolemia (FH) is also an inherited disease, resulting in the inability to process cholesterol properly, which leads to high levels of artery-clogging fat in the blood stream. Patients with FH often suffer heart attacks and strokes because of blocked arteries. A gene therapy approach used to battle FH is much more intricate than most gene therapies because it involves partial surgical removal of patients' livers (ex vivo transgene therapy). Corrected copies of a gene that serve to reduce cholesterol build-up are inserted into the liver sections, which are then transplanted back into the patients.

Gene therapy has also been tested on patients with AIDS. AIDS is caused by the human immunodeficiency virus (HIV), which weakens the body's immune system to the point that sufferers are unable to fight off diseases like pneumonias and cancer. In one approach, genes that produce specific HIV proteins have been altered to stimulate immune system functioning without causing the negative effects that a complete HIV molecule has on the immune system. These genes are then injected in the patient's blood stream. Another approach to treating AIDS is to insert, via white blood cells, genes that have been genetically engineered to produce a receptor that would attract HIV and reduce its chances of replicating.

Several cancers also have the potential to be treated with gene therapy. A therapy tested for melanoma, or skin cancer, involves introducing a gene with an anticancer protein called tumor necrosis factor (TNF) into test tube samples of the patient's own cancer cells, which are then reintroduced into the patient. In brain cancer, the approach is to insert a specific gene that increases the cancer cells' susceptibility to a common drug used in fighting the disease.

Gaucher disease is an inherited disease caused by a mutant gene that inhibits the production of an enzyme called glucocerebrosidase. Patients with Gaucher disease have enlarged livers and spleens and eventually their bones deteriorate. Clinical gene therapy trials focus on inserting the gene for producing this enzyme.

Gene therapy is also being considered as an approach to solving a problem associated with a surgical procedure known as balloon angioplasty. In this procedure, a stent (in this case, a type of tubular scaffolding) is used to open the clogged artery. However, in response to the trauma of the stent insertion, the body initiates a natural healing process that produces too many cells in the artery and results in restenosis, or reclosing of the artery. The gene therapy approach to preventing this unwanted side effect is to cover the outside of the stents with a soluble gel. This gel contains vectors for genes that reduce this overactive healing response.

The Human Genome Project

Although great strides have been made in gene therapy in a relatively short time, its potential usefulness has been limited by lack of scientific data concerning the multitude of functions that genes control in the human body. For instance, it is now known that the vast majority of genetic material does not store information for the creation of proteins, but rather is involved in the control and regulation of gene expression, and is therefore much more difficult to interpret. Even so, each individual cell in the body carries thousands of genes coding for proteins, with some estimates as high as 150,000 genes. For gene therapy to advance to its full potential, scientists must discover the biological role of each of these individual genes and where the base pairs that make them up are located on DNA.

To address this issue, the National Institutes of Health initiated the Human Genome Project in 1990. Led by James D. Watson (one of the co-discoverers of the chemical makeup of DNA), the project's 15-year goal is to map the entire human genome (a combination of the words gene and chromosomes). A genome map would clearly identify the location of all genes as well as the more than three billion base pairs that make them up. With a precise knowledge of gene locations and functions, scientists may one day be able to conquer or control diseases that have plagued humanity for centuries.

Scientists participating in the Human Genome Project have identified an average of one new gene a day, but many expect this rate of discovery to increase. By the year 2005, their goal is to determine the exact location of all the genes on human DNA and the exact sequence of the base pairs that make them up. Some of the genes identified through this project include a gene that predisposes people to obesity, one associated with programmed cell death (apoptosis), a gene that guides HIV viral reproduction, and the genes of inherited disorders like Huntington's disease, Lou Gehrig's disease, and some colon and breast cancers. In February of 2001, scientists published a rough draft of the complete human genome. With fewer than the anticipated number of genes found, between 30,000–40,000, the consequences of this announcement are enormous. Scientists caution however, that the initial publication is only a draft of the human genome and much more work is still ahead for the completion of the project. As the human genome is completed, there will be more information available for gene therapy research and implementation.

The future of gene therapy

Gene therapy seems elegantly simple in its concept: supply the human body with a gene that can correct a biological malfunction that causes a disease. However, there are many obstacles and some distinct questions concerning the viability of gene therapy. For example, viral vectors must be carefully controlled lest they infect the patient with a viral disease. Some vectors, like retroviruses, can also enter cells functioning properly and interfere with the natural biological processes, possibly leading to other diseases. Other viral vectors, like the adenoviruses, are often recognized and destroyed by the immune system so their therapeutic effects are short-lived. Maintaining gene expression so it performs its role properly after vector delivery is difficult. As a result, some therapies need to be repeated often to provide long-lasting benefits.

One of the most pressing issues, however, is gene regulation. Genes work in concert to regulate their functioning. In other words, several genes may play a part in turning other genes on and off. For example, certain genes work together to stimulate cell division and growth, but if these are not regulated, the inserted genes could cause tumor formation and cancer. Another difficulty is learning how to make the gene go into action only when needed. For the best and safest therapeutic effort, a specific gene should turn on, for example, when certain levels of a protein or enzyme are low and must be replaced. But the gene should also remain dormant when not needed to ensure it doesn't oversupply a substance and disturb the body's delicate chemical makeup.

One approach to gene regulation is to attach other genes that detect certain biological activities and then react as a type of automatic off-and-on switch that regulates the activity of the other genes according to biological cues. Although still in the rudimentary stages, researchers are making headway in inhibiting some gene functioning by using a synthetic DNA to block gene transcriptions (the copying of genetic information). This approach may have implications for gene therapy.

The ethics of gene therapy

While gene therapy holds promise as a revolutionary approach to treating disease, ethical concerns over its use and ramifications have been expressed by scientists and lay people alike. For example, since much needs to be learned about how these genes actually work and their long-term effect, is it ethical to test these therapies on humans, where they could have a disastrous result? As with most clinical trials concerning new therapies, including many drugs, the patients participating in these studies have usually not responded to more established therapies and are often so ill the novel therapy is their only hope for long-term survival.

Another questionable outgrowth of gene therapy is that scientists could possibly manipulate genes to genetically control traits in human offspring that are not health related. For example, perhaps a gene could be inserted to ensure that a child would not be bald, a seemingly harmless goal. However, what if genetic manipulation was used to alter skin color, prevent homosexuality, or ensure good looks? If a gene is found that can enhance intelligence of children who are not yet born, will everyone in society, the rich and the poor, have access to the technology or will it be so expensive only the elite can afford it?

The Human Genome Project, which plays such an integral role for the future of gene therapy, also has social repercussions. If individual genetic codes can be determined, will such information be used against people? For example, will someone more susceptible to a disease have to pay higher insurance premiums or be denied health insurance altogether? Will employers discriminate between two potential employees, one with a "healthy" genome and the other with genetic abnormalities?

Some of these concerns can be traced back to the eugenics movement popular in the first half of the twentieth century. This genetic "philosophy" was a societal movement that encouraged people with "positive" traits to reproduce while those with less desirable traits were sanctioned from having children. Eugenics was used to pass strict immigration laws in the United States, barring less suitable people from entering the country lest they reduce the quality of the country's collective gene pool . Probably the most notorious example of eugenics in action was the rise of Nazism in Germany, which resulted in the Eugenic Sterilization Law of 1933. The law required sterilization for those suffering from certain disabilities and even for some who were simply deemed "ugly." To ensure that this novel science is not abused, many governments have established organizations specifically for overseeing the development of gene therapy. In the United States, the Food and Drug Administration and the National Institutes of Health requires scientists to take a precise series of steps and meet stringent requirements before approving clinical trials.

In fact, gene therapy has been immersed in more controversy and surrounded by more scrutiny in both the health and ethical arena than most other technologies (except, perhaps, for cloning) that promise to substantially change society. Despite the health and ethical questions surrounding gene therapy, the field will continue to grow and is likely to change medicine faster than any previous medical advancement.

Resources

BOOKS

Hyde, Margaret O., and Lawrence E. Hyde. Cloning and theNew Genetics. Springfield, NJ: Enslow Publishers, Inc., 1984.

Stwertka, Eve, and Albert Stwertka. Genetic Engineering. New York: Franklin Watts, 1989.

Thompson, Larry. Correcting the Code: Inventing the GeneticCure for the Human Body. New York: Simon & Schuster, 1994.

PERIODICALS

Christensen R. "Cutaneous Gene Therapy—An Update." Histochemical Cell Biology (January 2001): 73-82.

"Initial Sequencing and Analysis of the Human Genome." Nature (February 15, 2001): 860-921.

Nevin, Norman. "What Has Happened to Gene Therapy?" European Journal of Pediatrics (2000): S240-S242.

Pekkanen, John. "Genetics: Medicine's Amazing Leap." Readers Digest (September 1991): 23-32.

Schemck, Harold M., Jr. "A New Era of Gene Therapy." FDAConsumer (December 1991): 14-19.

Weiss, Rick. "Gene Therapy at a Crossroads." Washington Post (October 19, 1994): 12-15.

ORGANIZATIONS

The National Human Genome Research Institute. The National Institutes of Health. 9000 Rockville Pike, Bethesda, MD 20892. (301) 496-2433. <http://www.nhgri.nih.gov>.

WEBSITES

Online Mendelian Inheritance in Man. Online genetic testing information sponsored by National Center for Biotechnology Information. <http://www.ncbi.nlm.nih.gov/Omim/>.

Katherine Hunt, MS

Gene therapy

Gene therapy is a rapidly growing field of medicine in which genes are introduced into the body to treat diseases. Genes control heredity and provide the basic biological code for determining a cell's specific functions. Gene therapy seeks to provide genes that correct or supplant the disease-controlling functions of cells that are not, in essence, doing their job. Somatic gene therapy introduces therapeutic genes at the tissue or cellular level to treat a specific individual. Germ-line gene therapy inserts genes into reproductive cells or possibly into embryos to correct genetic defects that could be passed on to future generations. Initially conceived as an approach for treating inherited diseases, like cystic fibrosis and Huntington's disease, the scope of potential gene therapies has grown to include treatments for cancers, arthritis, and infectious diseases. Although gene therapy testing in humans has advanced rapidly, many questions surround its use. For example, some scientists are concerned that the therapeutic genes themselves may cause disease. Others fear that germ-line gene therapy may be used to control human development in ways not connected with disease, like intelligence or appearance.

The biological basis of gene therapy

Gene therapy has grown out of the science of genetics or how heredity works. Scientists know that life begins in a cell, the basic building block of all multicellular organisms. Humans, for instance, are made up of trillions of cells, each performing a specific function. Within the cell's nucleus (the center part of a cell that regulates its chemical functions) are pairs of chromosomes. These threadlike structures are made up of a single molecule of DNA (deoxyribonucleic acid) , which carries the blueprint of life in the form of codes, or genes, that determine inherited characteristics.

A DNA molecule looks like two ladders with one of the sides taken off both and then twisted around each other. The rungs of these ladders meet (resulting in a spiral staircase-like structure) and are called base pairs. Base pairs are made up of nitrogen molecules and arranged in specific sequences. Millions of these base pairs, or sequences, can make up a single gene, specifically defined as a segment of the chromosome and DNA that contains certain hereditary information. The gene, or combination of genes formed by these base pairs ultimately direct an organism's growth and characteristics through the production of certain chemicals, primarily proteins, which carry out most of the body's chemical functions and biological reactions.

Scientists have long known that alterations in genes present within cells can cause inherited diseases like cystic fibrosis, sickle-cell anemia, and hemophilia . Similarly, errors in the total number of chromosomes can cause conditions such as Down syndrome or Turner syndrome . As the study of genetics advanced, however, scientists learned that an altered genetic sequence can also make people more susceptible to diseases, like atherosclerosis, cancer , and even schizophrenia . These diseases have a genetic component, but are also influenced by environmental factors (such as diet and lifestyle). The objective of gene therapy is to treat diseases by introducing functional genes into the body to alter the cells involved in the disease process by either replacing missing genes or providing copies of functioning genes to replace nonfunctioning ones. The inserted genes can be naturally occurring genes that produce the desired effect or may be genetically engineered (or altered) genes.

Scientists have known how to manipulate a gene's structure in the laboratory since the early 1970s through a process called gene splicing. The process involves removing a fragment of DNA containing the specific genetic sequence desired then inserting it into the DNA of another gene. The resultant product is called recombinant DNA and the process is genetic engineering.

There are basically two types of gene therapy. Germ-line gene therapy introduces genes into reproductive cells (sperm and eggs) or someday possibly into embryos in hopes of correcting genetic abnormalities that could be passed on to future generations. Most of the current work in applying gene therapy, however, has been in the realm of somatic gene therapy. In this type of gene therapy, therapeutic genes are inserted into tissue or cells to produce a naturally occurring protein or substance that is lacking or not functioning correctly in an individual patient.

Viral vectors

In both types of therapy, scientists need something to transport either the entire gene or a recombinant DNA to the cell's nucleus, where the chromosomes and DNA reside. In essence, vectors are molecular delivery trucks. One of the first and most popular vectors developed were viruses because they invade cells as part of the natural infection process. Viruses have the potential to be excellent vectors because they have a specific relationship with the host in that they colonize certain cell types and tissues in specific organs. As a result, vectors are chosen according to their attraction to certain cells and areas of the body.

One of the first vectors used was the retrovirus. Because these viruses are easily cloned (artificially reproduced) in the laboratory, scientists have studied them extensively and learned a great deal about their biological action. They have also learned how to remove the genetic information which governs viral replication, thus reducing the chances of infection.

Retroviruses work best in actively dividing cells, but cells in the body are relatively stable and do not divide often. As a result, these cells are used primarily for ex vivo (outside the body) manipulation. First, the cells are removed from the patient's body, and the virus, or vector, carrying the gene is inserted into them. Next, the cells are placed into a nutrient culture where they grow and replicate. Once enough cells are gathered, they are returned to the body, usually by injection into the blood stream. Theoretically, as long as these cells survive, they will provide the desired therapy.

Another class of viruses, called the adenoviruses, may also prove to be good gene vectors. These viruses can effectively infect nondividing cells in the body, where the desired gene product is then expressed naturally. In addition to being a more efficient approach to gene transportation, these viruses, which cause respiratory infections, are more easily purified and made stable than retroviruses, resulting in less chance of an unwanted viral infection. However, these viruses live for several days in the body, and some concern surrounds the possibility of infecting others with the viruses through sneezing or coughing. Other viral vectors include influenza viruses, Sindbis virus, and a herpes virus that infects nerve cells.

Scientists have also delved into nonviral vectors. These vectors rely on the natural biological process in which cells uptake (or gather) macromolecules. One approach is to use liposomes, globules of fat produced by the body and taken up by cells. Scientists are also investigating the introduction of raw recombinant DNA by injecting it into the bloodstream or placing it on microscopic beads of gold shot into the skin with a "gene-gun." Another possible vector under development is based on dendrimer molecules. A class of polymers (naturally occurring or artificial substances that have a high molecular weight and formed by smaller molecules of the same or similar substances), is "constructed" in the laboratory by combining these smaller molecules. They have been used in manufacturing Styrofoam, poly-ethylene cartons, and Plexiglass. In the laboratory, dendrimers have shown the ability to transport genetic material into human cells. They can also be designed to form an affinity for particular cell membranes by attaching to certain sugars and protein groups.

The history of gene therapy

In the early 1970s, scientists proposed "gene surgery" for treating inherited diseases caused by faulty genes. The idea was to take out the disease-causing gene and surgically implant a gene that functioned properly. Although sound in theory, scientists, then and now, lack the biological knowledge or technical expertise needed to perform such a precise surgery in the human body.

However, in 1983, a group of scientists from Baylor College of Medicine in Houston, Texas, proposed that gene therapy could one day be a viable approach for treating Lesch-Nyhan disease, a rare neurological disorder. The scientists conducted experiments in which an enzyme-producing gene (a specific type of protein) for correcting the disease was injected into a group of cells for replication. The scientists theorized the cells could then be injected into people with Lesch-Nyhan disease, thus correcting the genetic defect that caused the disease.

As the science of genetics advanced throughout the 1980s, gene therapy gained an established foothold in the minds of medical scientists as a promising approach to treatments for specific diseases. One of the major reasons for the growth of gene therapy was scientists' increasing ability to identify the specific genetic malfunctions that caused inherited diseases. Interest grew as further studies of DNA and chromosomes (where genes reside) showed that specific genetic abnormalities in one or more genes occurred in successive generations of certain family members who suffered from diseases like intestinal cancer, manic-depression, Alzheimer's disease, heart disease, diabetes , and many more. Although the genes may not be the only cause of the disease in all cases, they may make certain individuals more susceptible to developing the disease because of environmental influences, like smoking, pollution, and stress. In fact, some scientists theorize that all diseases may have a genetic component.

On September 14, 1990, a four-year-old girl with a genetic disorder that prevented her body from producing a crucial enzyme became the first person to undergo gene therapy in the United States. Because her body could not produce adenosine deaminase (ADA), she had a weakened immune system, making her extremely susceptible to severe, life-threatening infections. W. French Anderson and colleagues at the National Institutes of Health's Clinical Center in Bethesda, Maryland, took white blood cells (which are crucial to proper immune system functioning) from the girl, inserted ADA producing genes into them, and then transfused the cells back into the patient. Although the young girl continued to show an increased ability to produce ADA, debate arose as to whether the improvement resulted from the gene therapy or from an additional drug treatment she received.

Nevertheless, a new era of gene therapy began as more and more scientists sought to conduct clinical trial (testing in humans) research in this area. In that same year, gene therapy was tested on patients with melanoma (skin cancer). The goal was to help them produce anti-bodies (disease fighting substances in the immune system) to battle the cancer.

These experiments have spawned an ever-growing number of attempts at gene therapies designed to perform a variety of functions in the body. For example, a gene therapy for cystic fibrosis aims to supply a gene that alters cells, enabling them to produce a specific protein to battle the disease. Another approach was used for brain cancer patients, in which the inserted gene was designed to make the cancer cells more likely to respond to drug treatment. Gene therapy for patients who have artery blockage, which can lead to strokes, induces the growth of new blood vessels near clogged arteries, thus ensuring normal blood circulation.

Currently, there are a host of new gene therapy agents in clinical trials. In the United States, both nucleic acid-based (in vivo) treatments and cell-based (ex vivo) treatments are being investigated. Nucleic acid-based gene therapy uses vectors (like viruses) to deliver modified genes to target cells. Cell-based gene therapy techniques remove cells from the patient in order to genetically alter them then reintroduce them to the patient's body. Presently, gene therapies for the following diseases are being developed: cystic fibrosis (using adenoviral vector), HIV infection (cell-based), malignant melanoma (cell-based), Duchenne muscular dystrophy (cellbased), hemophilia B (cell-based), kidney cancer (cellbased), Gaucher disease (retroviral vector), breast cancer (retroviral vector), and lung cancer (retroviral vector). When a cell or individual is treated using gene therapy and successful incorporation of engineered genes has occurred, the cell or individual is said to be transgenic.

The medical establishment's contribution to transgenic research has been supported by increased government funding. In 1991, the U.S. government provided $58 million for gene therapy research, with increases in funding of $15–40 million dollars a year over the following four years. With fierce competition over the promise of societal benefit in addition to huge profits, large pharmaceutical corporations have moved to the forefront of transgenic research. In an effort to be first in developing new therapies, and armed with billions of dollars of research funds, such corporations are making impressive strides toward making gene therapy a viable reality in the treatment of once elusive diseases.

Diseases targeted for treatment by gene therapy

The potential scope of gene therapy is enormous. More than 4,200 diseases have been identified as resulting directly from abnormal genes, and countless others that may be partially influenced by a person's genetic makeup. Initial research has concentrated on developing gene therapies for diseases whose genetic origins have been established and for other diseases that can be cured or ameliorated by substances genes produce.

The following are examples of potential gene therapies. People suffering from cystic fibrosis lack a gene needed to produce a salt-regulating protein. This protein regulates the flow of chloride into epithelial cells, (the cells that line the inner and outer skin layers) which cover the air passages of the nose and lungs. Without this regulation, patients with cystic fibrosis build up a thick mucus that makes them prone to lung infections. A gene therapy technique to correct this abnormality might employ an adenovirus to transfer a normal copy of what scientists call the cystic fibrosis transmembrane conductance regulator, or CTRF, gene. The gene is introduced into the patient by spraying it into the nose or lungs.

Familial hypercholesterolemia (FH) is also an inherited disease, resulting in the inability to process cholesterol properly, which leads to high levels of artery-clogging fat in the blood stream. Patients with FH often suffer heart attacks and strokes because of blocked arteries. A gene therapy approach used to battle FH is much more intricate than most gene therapies because it involves partial surgical removal of patients' livers (ex vivo transgene therapy). Corrected copies of a gene that serve to reduce cholesterol build-up are inserted into the liver sections, which are then transplanted back into the patients.

Gene therapy has also been tested on patients with AIDS. AIDS is caused by the human immunodeficiency virus (HIV), which weakens the body's immune system to the point that sufferers are unable to fight off diseases like pneumonias and cancer. In one approach, genes that produce specific HIV proteins have been altered to stimulate immune system functioning without causing the negative effects that a complete HIV molecule has on the immune system. These genes are then injected in the patient's blood stream. Another approach to treating AIDS is to insert, via white blood cells, genes that have been genetically engineered to produce a receptor that would attract HIV and reduce its chances of replicating.

Several cancers also have the potential to be treated with gene therapy. A therapy tested for melanoma, or skin cancer, involves introducing a gene with an anti-cancer protein called tumor necrosis factor (TNF) into test tube samples of the patient's own cancer cells, which are then reintroduced into the patient. In brain cancer, the approach is to insert a specific gene that increases the cancer cells' susceptibility to a common drug used in fighting the disease.

Gaucher disease is an inherited disease caused by a mutant gene that inhibits the production of an enzyme called glucocerebrosidase. Patients with Gaucher disease have enlarged livers and spleens and eventually their bones deteriorate. Clinical gene therapy trials focus on inserting the gene for producing this enzyme.

Gene therapy is also being considered as an approach to solving a problem associated with a surgical procedure known as balloon angioplasty. In this procedure, a stent (in this case, a type of tubular scaffolding) is used to open the clogged artery. However, in response to the trauma of the stent insertion, the body initiates a natural healing process that produces too many cells in the artery and results in restenosis, or reclosing of the artery. The gene therapy approach to preventing this unwanted side effect is to cover the outside of the stents with a soluble gel. This gel contains vectors for genes that reduce this overactive healing response.

The Human Genome Project

Although great strides have been made in gene therapy in a relatively short time, its potential usefulness has been limited by lack of scientific data concerning the multitude of functions that genes control in the human body. For instance, it is now known that the vast majority of genetic material does not store information for the creation of proteins, rather, it is involved in the control and regulation of gene expression, and is therefore much more difficult to interpret. Even so, each individual cell in the body carries thousands of genes coding for proteins, with some estimates as high as 150,000 genes. For gene therapy to advance to its full potential, scientists must discover the biological role of each of these individual genes and where the base pairs that make them up are located on DNA.

To address this issue, the National Institutes of Health initiated the Human Genome Project in 1990. Led by James D. Watson (one of the co-discoverers of the chemical makeup of DNA), the project's 15-year goal is to map the entire human genome (a combination of the words gene and chromosomes). A genome map would clearly identify the location of all genes as well as the more than three billion base pairs that make them up. With a precise knowledge of gene locations and functions, scientists may one day be able to conquer or control diseases that have plagued humanity for centuries.

Scientists participating in the Human Genome Project have identified an average of one new gene a day, but many expect this rate of discovery to increase. Their goal is to determine the exact location of all the genes on human DNA and the exact sequence of the base pairs that make them up (see the "Human Genome Project" entry for updated information). Some of the genes

identified through this project include a gene that predisposes people to obesity, one associated with programmed cell death (apoptosis), a gene that guides HIV viral reproduction, and the genes of inherited disorders like Huntington disease , Lou Gehrig's disease, and some colon and breast cancers. With fewer than the anticipated number of genes found, between 30,000 and 40,000, the consequences of this announcement are enormous. Scientists caution however, that the initial publication is only a draft of the human genome and much more work is still ahead for the completion of the project. As the human genome is completed, there will be more information available for gene therapy research and implementation.

The future of gene therapy

Gene therapy seems elegantly simple in its concept: supply the human body with a gene that can correct a biological malfunction that causes a disease. However, there are many obstacles and some distinct questions concerning the viability of gene therapy. For example, viral vectors must be carefully controlled lest they infect the patient with a viral disease. Some vectors, like retroviruses, can also enter cells functioning properly and interfere with the natural biological processes, possibly leading to other diseases. Other viral vectors, like the adenoviruses, are often recognized and destroyed by the immune system so their therapeutic effects are short-lived. Maintaining gene expression so it performs its role properly after vector delivery is difficult. As a result, some therapies need to be repeated often to provide long-lasting benefits.

One of the most pressing issues, however, is gene regulation. Genes work in concert to regulate their functioning. In other words, several genes may play a part in turning other genes on and off. For example, certain genes work together to stimulate cell division and growth, but if these are not regulated, the inserted genes could cause tumor formation and cancer. Another difficulty is learning how to make the gene go into action only when needed. For the best and safest therapeutic effort, a specific gene should turn on, for example, when certain levels of a protein or enzyme are low and must be replaced. But the gene should also remain dormant when not needed to ensure it does not oversupply a substance and disturb the body's delicate chemical makeup.

One approach to gene regulation is to attach other genes that detect certain biological activities and then react as a type of automatic off-and-on switch that regulates the activity of the other genes according to biological cues. Although still in the rudimentary stages, researchers are making headway in inhibiting some gene functioning by using a synthetic DNA to block gene transcriptions (the copying of genetic information). This approach may have implications for gene therapy.

The ethics of gene therapy

While gene therapy holds promise as a revolutionary approach to treating disease, ethical concerns over its use and ramifications have been expressed by scientists and lay people alike. For example, since much needs to be learned about how these genes actually work and their long-term effect, is it ethical to test these therapies on humans, where they could have a disastrous result? As with most clinical trials concerning new therapies, including many drugs, the patients participating in these studies have usually not responded to more established therapies and are often so ill the novel therapy is their only hope for long-term survival.

Another questionable outgrowth of gene therapy is that scientists could possibly manipulate genes to genetically control traits in human offspring that are not health related. For example, perhaps a gene could be inserted to ensure that a child would not be bald, a seemingly harmless goal. However, what if genetic manipulation was used to alter skin color, prevent homosexuality, or ensure good looks? If a gene is found that can enhance intelligence of children who are not yet born, will everyone in society, the rich and the poor, have access to the technology or will it be so expensive only the elite can afford it?

The Human Genome Project, which plays such an integral role for the future of gene therapy, also has social repercussions. If individual genetic codes can be determined, will such information be used against people? For example, will someone more susceptible to a disease have to pay higher insurance premiums or be denied health insurance altogether? Will employers discriminate between two potential employees, one with a "healthy" genome and the other with genetic abnormalities?

Some of these concerns can be traced back to the eugenics movement popular in the first half of the twentieth century. This genetic "philosophy" was a societal movement that encouraged people with "positive" traits to reproduce while those with less desirable traits were sanctioned from having children. Eugenics was used to pass strict immigration laws in the United States, barring less suitable people from entering the country lest they reduce the quality of the country's collective gene pool . Probably the most notorious example of eugenics in action was the rise of Nazism in Germany, which resulted in the Eugenic Sterilization Law of 1933. The law required sterilization for those suffering from certain disabilities and even for some who were simply deemed "ugly." To ensure that this novel science is not abused, many governments have established organizations specifically for overseeing the development of gene therapy. In the United States, the Food and Drug Administration and the National Institutes of Health requires scientists to take a precise series of steps and meet stringent requirements before approving clinical trials.

In fact, gene therapy has been immersed in more controversy and surrounded by more scrutiny in both the health and ethical arena than most other technologies (except, perhaps, for cloning) that promise to substantially change society. Despite the health and ethical questions surrounding gene therapy, the field will continue to grow and is likely to change medicine faster than any previous medical advancement.

Resources

BOOKS

Hyde, Margaret O., and Lawrence E. Hyde. Cloning and the New Genetics. Springfield, NJ: Enslow Publishers, Inc., 1984.

Stwertka, Eve, and Albert Stwertka. Genetic Engineering. New York: Franklin Watts, 1989.

Thompson, Larry. Correcting the Code: Inventing the Genetic Cure for the Human Body. New York: Simon & Schuster, 1994.

PERIODICALS

Christensen R. "Cutaneous Gene Therapy—An Update." Histochemical Cell Biology (January 2001): 73-82.

"Initial Sequencing and Analysis of the Human Genome." Nature (February 15, 2001): 860-921.

Nevin, Norman. "What Has Happened to Gene Therapy?" European Journal of Pediatrics (2000): S240-S242.

Pekkanen, John. "Genetics: Medicine's Amazing Leap." Readers Digest (September 1991): 23-32.

Schemck, Harold M., Jr. "A New Era of Gene Therapy." FDA Consumer (December 1991): 14-19.

Weiss, Rick. "Gene Therapy at a Crossroads." Washington Post (October 19, 1994): 12-15.

ORGANIZATIONS

The National Human Genome Research Institute. The National Institutes of Health. 9000 Rockville Pike, Bethesda, MD 20892. (301) 496-2433. <http://www.nhgri.nih.gov>.

WEBSITES

Online Mendelian Inheritance in Man. Online genetic testing information sponsored by National Center for Biotechnology Information. <http://www.ncbi.nlm.nih.gov/Omim/>.

Katherine Hunt, MS