Gene therapy

Definition

Classic gene therapy is the direct use of genetic material in the treatment of disease. This usually involves inserting a functional gene or DNA fragment into key cells to mitigate, or cure, a disease. A broader definition of gene therapy includes all applications of DNA technology to treat disease. For people with certain neurological conditions such as Parkinson's disease and Canavan disease, initial gene therapy trials have shown promise. Developing gene therapies for treating disorders of the nervous system poses unique challenges, such as how to introduce the therapeutic gene across the blood-brain barrier or how to target the therapeutic gene to one specific area of the brain.

Purpose

Genes play a role in every function of the human body. Defects or mutations within a gene can lead to malfunction or disease of cells, tissues, and/or organs. Although standard drug therapy is usually effective in treating the symptoms of a disorder, a patient may be required to take the drugs for an extended time and there may be serious or unpleasant side effects. However, a patient may be cured with few negative consequences if treatment can be targeted directly at the specific cause of the disease (the gene defect), or if that cause can be neutralized or reversed. Therefore, gene therapy provides an attractive alternative to drug therapy as it seeks to provide treatment strategies that will be more complete and less toxic to the patient. Furthermore, gene therapy may provide a way of treating diseases that cannot be managed by standard therapies.

Description

There are many diverse approaches to gene therapy since the biological basis of each disease is unique, presenting a different set of parameters and challenges. However, in each case, a basic set of criteria must be met. First, it is essential to fully understand the disease to be treated. The cells or tissues associated with the disease must be well defined and accessible. The gene and the specific mutation or mutations causing the disease must be known, and it must be possible to isolate or synthesize a normal, functional copy of that gene and to incorporate it into a vector. The vector then transfers the new gene to the target cells where, hopefully, the gene will become fully active. The most common roles for the expressed gene include replacing a defective gene, inhibiting or degrading a deleterious DNA, RNA, or protein, or directly or indirectly killing the cell.

Single gene disorders resulting in a loss of gene function in one specific target tissue provide the easiest options for gene therapy, though strategies for many types of mutations have been investigated. A broad spectrum of diseases has been considered for gene therapy, including:

neurological disorders, e.g., Parkinson disease, Huntington disease
muscular dystrophies
immunological disorders, e.g., severe combined immunodeficiency syndrome (SCIDS)
blood abnormalities, e.g., thalassemias, hemophilia
cancer

Unfortunately, many of the more commonly occurring disorders, including heart disease, diabetes, and high blood pressure, result from defects in multiple genes making them unlikely candidates for gene therapy using existing technologies.

For each disease, it must be determined if ex vivo or in vitro technology is the best approach. In ex vivo technology, patient cell samples are collected and cultured in the laboratory. The new gene is incorporated into the growing cells, and these are subsequently transferred back into the patient. Not all of the cultured cells will include the new gene, and not all will survive the transfer. The hope is that a sufficient number of the modified cells will be functional in the patient such that the therapy will reverse the disease. In vitro therapy involves injecting the new gene directly into the target tissue where the individual cells must pick it up. Of the two, this method is technically easier and cheaper, but it is harder to determine how many of the target cells actually acquire the new gene. Ex vivo therapy is more expensive and time consuming, but allows greater control of the conditions.

Both processes require the use of a vector to get the new gene across the cell membrane and into a cell. Viruses have proven to be highly effective as vectors since these are biological entities with a natural function of infecting host cells. DNA technology allows viruses to be manipulated to replace the normal payload of disease-causing genetic material with therapeutic genes. The virus will retain its ability to infect a host cell but, instead of causing a disease, it will deposit the new gene into the cell.

Other mechanisms of gene transfer have also been investigated. Artificial chromosomes have been developed, but these are often too large to move across cell membranes. Liposomes, structures with lipid membranes, that encompass genetic material can be successfully used as vectors if the liposome is absorbed by the cell or if its membrane fuses with the cell membrane releasing the new gene inside the cell.

Once the gene enters the cell, one of two things occurs. It may be degraded and lost, which is an unfavorable outcome. Preferably, the gene will stably incorporate into the DNA of the target cell so that it can be processed as a normal part of that genome. If the gene therapy is designed to replace a defective gene, the best-case scenario is for the new gene to integrate into a completely renewable cell such as a stem cell. Theoretically, in this situation, the gene will be permanently incorporated into the patient's body and no further therapy will be required. Alternatively, if the gene integrates into a genome of a cell with a finite lifespan, the beneficial effects of the gene will only exist while that cell lives, requiring the gene therapy to be repeated at a later time.

One of the early successes of gene therapy was for a four-year-old girl with adenine deaminase (ADA) deficiency. This is a form of SCIDS that results in malfunction of the immune system and can lead to death as a result of severe infection. Conventional treatment had failed for this patient, making her a candidate for gene therapy. A normal ADA gene was incorporated into a retroviral vector that transferred the gene into the patient's lymphocytes in vitro. The modified cells were returned to her circulation by transfusion. After five months, her levels of ADA activity had risen from less than 1% to 50%. With additional therapies over the next two years, her health improved as the enzyme activity stabilized, and she was able to begin a normal life. Twelve years later, she still demonstrates reasonable levels of ADA activity, but the gene therapy was not a cure as she must continue to receive the standard enzyme replacement therapy to maintain her health.

Acquired diseases can also be treated with gene therapy as demonstrated by a novel strategy for treating brain cancer. The thymidine kinase (TK) gene from the herpes simplex virus (HSV) has an enzymatic property that converts the drug ganciclovir into a toxic substance that can kill human cells. It was postulated that this could be used as a targeted killing tool. To investigate, cloned HSV TK genes were injected into brain tumors. In the brain, only the tumor cells are dividing, so these are the only cells that will be infected by the viral vector, and are thus the only cells that will receive the HSV TK gene. When the patient is subsequently treated with ganciclovir, the tumor cells that have incorporated the HSV TK gene will be selectively killed. Clinical trials proved that tumor cells could be selectively eliminated by demonstrating a reduction in the size of the brain tumors in seven of nine patients.

A completely different set of therapies is possible if the idea of gene therapy includes the use of DNA for patient treatment in ways other than inserting new genes into cells. One example is the drug Gleevec that was approved in 2001 for use in patients with chronic myelogenous leukemia (CML). Gleevec is a substance that binds to the defective protein produced in CML, blocking that protein's activity and alleviating the symptoms of the disease. This is a targeted therapy that affects only the cells with the CML mutation, so there are very few side effects. Recombinant DNA technology has also been utilized to generate genetically engineered copies of vaccines (Recombivax HB), antibodies, and normal gene products (insulin).

Aftercare

If the new DNA can be stably incorporated into the proper regenerative target cells, the patient may be cured of disease. No additional care should be required, although periodic monitoring of the patient is appropriate.

For gene therapies in which the new DNA is inserted into cells with a finite lifespan, the therapeutic effect will be lost when those cells die. In these situations, the patient will require continuing treatments. Monitoring of patients who receive drugs and substances arising from recombinant DNA technology is the same as standard drug therapy.

Precautions

Currently classic gene therapy is still experimental. Although many patients have shown significant improvement following their treatment, at least two individuals have died as a result of this type of therapy. Therefore, experts carefully review all protocols before any studies are undertaken. Initial research is done in an animal model system, and any problems detected are carefully evaluated before the same treatments are attempted in humans.

Risks

A patient who is receiving gene therapy may face a number of potential problems. The viral vectors used may cause infection and/or inflammation of tissues, and artificial introduction of viruses into the body may initiate other disease processes. Functional gene therapy relies on stable incorporation of a new gene into an individual's own DNA. As the integration is random, occasionally the new gene may insert within another normally functioning gene, causing its damage or inactivation. This, in turn, could lead to cancer or other disease. It is also critical that the new gene have the proper regulatory controls so that the gene product is produced in the proper amount. Over-expression of certain genes can have deleterious results. Any of these problems could render the gene therapy ineffective, or, at worst, cause the death of the subject.

Normal results

Classic gene therapy seeks to treat or cure a defined disease by incorporating a functional gene or gene product into target cells of an affected individual.