How Gene Therapy Works

Understanding this medical treatment requires a working knowledge of genes. The good news is you probably covered this in your high-school biology class, but just in case you've forgotten, here's a quick recap. A gene refers to a single unit of hereditary information -- a factor that controls some specific activity or trait. Genes exist on chromosomes, which themselves reside in the nuclei of our cells.

Chromosomes, of course, contain long chains of DNA built with repeating subunits known as nucleotides. That means a single gene is a finite stretch of DNA with a specific sequence of nucleotides. Those nucleotides act as a blueprint for a specific protein, which gets assembled in a cell using a multistep process.

Proteins are the workhorses of cells. They help build the physical infrastructure, but they also control and regulate important metabolic pathways. If a gene malfunctions -- if, say, its sequence of nucleotides gets scrambled -- then its corresponding protein won't be made or won't be made correctly. Biologists call this a mutation, and mutations can lead to all sorts of problems, such as cancer and phenylketonuria.

Gene therapy tries to restore or replace a defective gene, bringing back a cell's ability to make a missing protein. On paper, it's straightforward: You simply insert the correct version of a gene into a strand of DNA. In reality, it's a little more complicated because cells require some outside assistance in the form of a virus. You probably think of viruses as agents that cause infections -- smallpox, influenza, rabies or AIDS. In gene therapy, scientists use these tiny living-but-not-living particles to give a cell a genetic makeover. In the next section, we'll explore which viruses are used and why.

Viruses perplexed biologists for years. They could see the effects of viruses -- illness -- but they couldn't isolate the infecting agent. At first, they thought they were dealing with extremely small bacteria cells. Then, amid a flurry of interest in viruses, American scientist Wendell Stanley crystallized the particles responsible for tobacco mosaic disease and described viruses for the world in 1935.

These strange entities don't have nuclei or other cellular structures, but they do have nucleic acid, either DNA or RNA. This small packet of genetic information is packed inside a protein coat, which, in some cases, is wrapped in a membranous envelope.

Unlike other living things, viruses can't reproduce on their own because they don't have the necessary cellular machinery. They can, however, reproduce if they invade a cell and borrow the cell's equipment and enzymes. The basic process works like this:

That ability to carry genetic information into cells makes viruses useful in gene therapy. What if you could replace a snippet of viral DNA with the DNA of a human gene and then let that virus infect a cell? Wouldn't the host cell make copies of the introduced gene and then follow the blueprint of the gene to churn out the associated protein? As it turns out, this is completely possible -- as long as scientists modify the virus to prevent it from causing disease or inducing an immune reaction by the host. When so modified, such a virus can become a vehicle, or vector, to deliver a specific gene therapy.

Today, researchers use several types of viruses as vectors. One favorite is adenovirus, the agent responsible for the common cold in humans. Adenoviruses introduce their DNA into the nucleus of the cell, but the DNA isn't integrated into a chromosome. This makes them good vectors, but they often stimulate an immune response, even when weakened. As an alternative, researchers may rely on adeno-associated viruses, which cause no known human diseases. Not only that, they integrate their genes into host chromosomes, making it possible for the cells to replicate the inserted gene and pass it on to future generations of the altered cells. Retroviruses, like the ones that cause AIDS and some types of hepatitis, also splice their genetic material into the chromosomes of the cells they invade. As a result, researchers have studied retroviruses extensively as vectors for gene therapy.

The idea of gene therapy has been bouncing around scientists' brains for decades. In fact, it was Edward Tatum, an American geneticist, who first suggested that genetic diseases might be cured with "genetic engineering" in 1966. That same year, another American, Joshua Lederberg, actually outlined the details of "virogenic therapy" in an article published in The American Naturalist. Numerous researchers then worked diligently to move gene therapy from concept to reality. In 1972, biochemist Paul Berg figured out how to snip out a section of human DNA and insert it into the genome of a virus, which he then used to infect bacteria cells. Eventually, he was able to get bacteria to produce human insulin. Ten years later, Ronald M. Evans inserted the gene for rat growth hormone into a retrovirus and then transferred that gene into mouse cells.

All of these efforts set the stage for a gene-therapy revolution. The first gene therapy trial approved by the U.S. Food and Drug Administration took place in 1990. The trial focused on patients with severe combined immunodeficiency (SCID), also known as "bubble boy" disease after David Vetter, who lived in the sterile environment of a plastic bubble until he died in 1984 at age 12.

Researchers in this trial used what is known as an out-of-body gene therapy. First, they harvested marrow from a patient by inserting a special needle through the skin and into the hip bone. Then, in the laboratory, they exposed the stem cells from the marrow to retroviruses whose RNA had been modified to contain the gene associated with SCID. The retroviruses infected the stem cells and inserted the functional gene into the host chromosome. Next, scientists took the engineered stem cells and injected them back into the patient's bloodstream. The cells made a beeline for the bone marrow and, like all good stem cells, matured into different cell types, including healthy T cells with functioning copies of the necessary gene. Using this technique, dozens of children with SCID have been completely cured. But it's not the only disease -- or approach -- on the playlist of geneticists [source: Nienhuis].

The second common way to administer gene therapy is to inject the gene-carrying virus directly into the region that has defective cells. James Wilson, professor of pathology and laboratory medicine at the University of Pennsylvania, pioneered this so-called "in-the-body" gene therapy in the 1990s. He used adenovirus as his vector, but he weakened it to limit the immune response in the recipient. In early tests, his modified virus seemed to cause no harm at all -- not even sniffles -- in test subjects. That meant it could deliver genes reliably with few side effects.

In 1999, he led a phase I clinical trial to test adenovirus-based therapy for the treatment of a rare genetic disorder called ornithine transcarbamylase (OTC) deficiency. OTC is an enzyme that helps the body break down excess nitrogen. Without it, ammonia levels increase until the brain becomes poisoned. A single gene on the X chromosome codes for the enzyme, making it an ideal candidate for the experimental therapy. Wilson inserted the OTC gene into weakened adenovirus particles and then injected those into the livers of 18 patients [source: Neimark].

The idea was simple: The virus would infect the liver cells, which would then proceed to replicate the OTC gene and begin manufacturing the enzyme. Unfortunately, one of the patients, 18-year-old Jesse Gelsinger, died just three days after receiving his injection of the engineered virus. Scientists now think that Gelsinger's body mounted a massive immune response, leading to widespread organ failure. That's just one of the risks of gene therapy, as we'll see on the next page.

Jesse Gelsinger's death stunned the public, and it sent shock waves through the scientific community, too. Geneticists came to the painful realization that gene therapy, while easy to diagram on paper, came loaded with challenges and pitfalls. And it didn't matter how they tackled the problem -- both in- and out-of-the body approaches came with inherent risks.

For in-the-body gene therapy, the biggest issue is the immune system of the patient. The body views adenovirus particles, even those carrying a human gene, as foreign objects. When they enter host cells, the host responds by mounting a counterattack to get rid of the invaders. This is what happened with Jesse Gelsinger. His immune system didn't realize the viruses were trying to be helpful, and it launched a vigorous attack, shutting down his organs in the process. Today, researchers might give Gelsinger lower therapy doses or pretreat him with immunosuppressive drugs. Another option being explored involves "naked" DNA, which refers to a nucleic acid molecule stripped of its viral carrier.

Out-of-the-body therapies relying on retroviruses have their own problems. Remember, retroviruses stitch their DNA into the host chromosome, which is a bit like picking up a short phrase from one sentence and plugging it into a longer sentence. If the insertion doesn't occur in just the right place, the resulting "language" might not make any sense. In some gene therapy trials using retroviruses, patients have developed leukemia and other forms of cancer because inserting one gene disrupts the function of other surrounding genes. This complication has affected several children in the SCID trials, although many of them have beaten the cancer with other therapies.

Because of these issues, the U.S. Food and Drug Administration (FDA) regulates all gene therapy products in the United States through its Center for Biologics Evaluation and Research, or CBER. The center also provides proactive scientific and regulatory advice to medical researchers and manufacturers interested in developing human gene therapy products. Investigators can also turn to the National Institutes of Health for guidance and guidelines when conducting clinical trials with gene therapy.

To date, the center hasn't yet approved any human gene therapy product for sale, although several ongoing trials are producing promising results. Up next, we'll look at a few recent successes in what many believe is the second revolution of gene therapy.

In the wake of Jesse Gelsinger's death, the FDA banned James Wilson from conducting gene-therapy experiments using human subjects. Other researchers, however, didn't operate under those same restrictions.

In 2007, Jean Bennett, a molecular geneticist and physician at the University of Pennsylvania School of Medicine, and her husband, Albert Maguire, a retinal surgeon at Children's Hospital of Philadelphia, began a clinical trial to study a gene-therapy treatment for a rare form of blindness known as leber congenital amaurosis (LCA). A mutation in a gene known as RPE65 leads to a deficiency in a protein that's vital to the normal function of the retina. People who lack this protein suffer progressive loss of vision until they lose all sight, usually by the age of 40.

Bennett and Maguire inserted the RPE65 gene into an adeno-associated virus, the kinder, gentler version of adenovirus. They then injected the engineered virus in low doses into the retinas of three patients. The viruses infected the retinal cells, which started churning out the RPE65 protein. Lo and behold, the vision of all three subjects improved, and no nasty side effects -- including dangerous immune responses -- were reported. The team decided to test a larger test population with a stronger dose of the virus. Six more LCA patients received the gene therapy and enjoyed even better results [source: Kaiser].

That puts SCID and LCA into a rarefied category -- diseases successfully treated by gene therapy. And yet geneticists and molecular biologists feel confident there will be more. James Wilson, who continues to contribute to the field, has isolated 120 types of adeno-associated viruses, many of which perform more effectively in some tissues than others. For example, some of these vectors have an affinity for heart tissue, while others have an affinity for cells in the spinal cord and brain. Future research may yield viable treatments for spinal injuries and for diseases such as Parkinson's [source: Neimark].

Researchers are also making great progress with out-of-the-body therapies. In July 2013, the journal Science published the results of two studies investigating the use of lentiviruses as gene therapy vectors. Lentiviruses are retroviruses, but they are unique in their ability to transfer genes efficiently and permanently in both dividing and nondividing cells. Other retroviruses must do their genetic voodoo on dividing cells. Perhaps more important, lentiviruses seem less prone to activating other cancer-related genes when they insert their payload into the host's DNA. When researchers tested lentiviral-based therapy on patients with adrenoleukodystrophy, an X-linked neurodegenerative disease that affects young males, and metachromatic leukodystrophy, a rare neurodegenerative disease caused by mutations in a single gene, they were able to arrest the progression of both diseases with no harmful side effects [source: Cossins].

In the future, other promising gene therapies are sure to emerge, mostly for hereditary diseases, such as cystic fibrosis, muscular dystrophy, sickle cell anemia and hemophilia. Even phenylketonuria may become a thing of the past, something that probably would make Charlie Gordon pretty happy.