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.
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
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 such inherited diseases as cystic fibrosis, sickle-cell disease, and hemophilia. Similarly, errors in entire chromosomes may cause such conditions 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 such diseases as atherosclerosis, cancer, and schizophrenia. These diseases have a genetic component, but are also influenced by such environmental factors 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, antiinflammatory 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.
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 likli-hood 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 herpesvirus 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.
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.
As of 2001, 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 (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
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. With fewer than the anticipated number of genes found, between 30,000 and 40,000, the consequences of this announcement are potentially profound. 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. 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, amyotrophic lateral aclerosis (Lou Gehrig's disease), and some colon and breast cancers. As the human genome is completed, more information will be available for gene therapy research and implementation.
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
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 such diseases 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. As of 2001, these approaches are experimental and have not been approved for treatment.
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 con trol 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 harm less goal. However, what if genetic manipulation were used to alter skin color, prevent homosexuality, or ensure good looks? If a gene is found that can enhance intelli gence 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,
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.
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—A type of molecule made by cells 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-related disease in an individual.
Vector—Something used to transport genetic information to a cell.
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 Lee Goldman, and J. Claude Bennett. 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, MA: 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.
Gottlieb S. "Gene therapy shows promise for hemophilia." British Medical Journal, 322 no.7300 (2001): 1442A-1443.
Gray SG. "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 CM, Lawrie A, Brisken AF, Cumberland DC. "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 IM. "Ombudsman or Hotline for Gene Therapy Clinical Trials?" Molecular Therapeutics, 3 no.6 (2001): 817-818.
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>.
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>.
US Food and Drug Administration. <http://www.fda.gov/fdac/features/2000/gene.html>.
Vanderbilt University. <http://www.mc.vanderbilt.edu/gcrc/gene>.
L. Fleming Fallon, Jr., MD, DrPH