Gene therapy

Definition

Gene therapy is a type of treatment designed to modify the expression of an individual’s genes or to correct abnormal genes to treat a disease.

R. Michael Blaese, W. French Anderson and Kenneth Culver at a press conference announcing the start of the first gene therapy trial for treating children with severe combined immunodeficiency, 13 September 1990. Source: National Cancer Institute

Importance

Gene therapy gained a lot of commercial interest in the 1980s. In part this was because many assumed such treatment would move swiftly and easily from proof of concept into clinical trials. Such hopes, however, were dashed following the death of the first patient in a gene therapy trial in 1999. It would take another decade before optimism about the therapy resurfaced. From 2008 onwards dozens of new start-ups began to be created around gene therapy. These were founded on the back of sponsorship from pharmaceutical companies and the stock market. Just how much weight began to be attached to gene therapy can be seen by the stock market’s valuation of Juno Therapeutics. In 2014, just one year after Juno was set up, the company was valued at US$4 billion. When the first gene therapy was approved in the United States there were 854 companies developing such therapies. According to the Alliance for Regenerative Medicine there were 1085 companies in that space by the end of 2020 and more than 400 gene therapy trials under way.

Discovery

Scientists first demonstrated the feasibility of incorporating new genetic functions in mammalian cells in the late 1960s. Several methods were used. One involved injecting genes with a micropipette directly into a living mammalian cell. Another exposed cells to a precipitate of DNA containing the desired genes. A virus could also be used as a vehicle, or vector, to deliver the genes into cells.

One of the first people to report the direct incorporation of functional DNA into a mammalian cell was Lorraine Kraus at the University of Tennessee. In 1961 she managed to genetically alter the haemoglobin of cells from bone marrow taken from a patient with sickle-cell anaemia. She did this by incubating the patient’s cells in tissue culture with DNA extracted from a donor with normal haemoglobin. Seven years later, Theodore Friedmann, Jay Seegmiller and John Subak-Sharpe at the National Institutes of Health (NIH), Bethesda, successfully corrected genetic defects associated with Lesch-Nyhan syndrome, a debilitating neurological disease. They did this by adding foreign DNA to cultured cells collected from patients suffering from the disease.

The first humans to receive gene therapy took place in 1970. It was administered to two very young West German sisters suffering from hyperargininemia, an extremely rare genetic disorder that prevents the production of arginase. This is an enzyme that helps prevent the build up of arginine in bodily fluids. Any accumulation can cause brain damage, epilepsy and other neurological and muscular problems. Each sister received an injection of a rabbit virus (Shope papilloma) known to induce the production of arginase. The injection was given as a last desperate measure to rescue the children. The treatment was carried out by Stanfield Rogers, an American physician, together with H. G. Terheggen, a German paediatrician. They took the risk based on observations Rogers had previously made with some laboratory technicians at Oak Ridge National Laboratory who became infected with the rabbit virus when working with it. None of the technicians experienced ill-effects from the virus but had abnormally low levels of arginine in their blood. This was apparent even in a technician whose last exposure to the virus had been 20 years before. Rogers connected the technicians’ abnormal arginine levels with a gene in the rabbit virus which was known to encourage the production of arginase in rabbits. By giving the rabbit virus to the girls, Rogers hoped to transfer genetic instructions to their cells to produce arginase. After the two sisters were treated a third sister was born afflicted with hyperargininemia. She was also injected with the virus. Disappointingly none of the sisters responded to the treatment.

A new pathway for gene therapy opened up with the development of genetic engineering in the early 1970s. The technique provided two key tools. Firstly, a means to clone specific disease genes. Secondly, an efficient method for gene transfer. The potential of the technology for gene therapy was first highlighted by the US scientists Theodore Friedmann and Richard Roblin. In 1972 they published an article in Science suggesting genetically modified tumour viruses might be used to transfer the necessary genetic information to treat genetic disorders in patients.

The technique was first tried out in the case of beta-thalassemia. Linked to an inherited defect in a gene for beta-globin, this blood disorder usually causes premature death. The beta-globin gene was first cloned by scientists at Cold Spring Harbor Laboratory and Harvard University in 1976. It was the first disease gene ever cloned. Three years later, a team led by Martin Cline at the University of California, Los Angeles, reported the successful introduction of the gene into the bone marrow of irradiated mice. Following this, Cline and his team unsuccessfully tried to treat two beta-thalassemia patients, one in Italy and another in Israel by inserting the gene into bone marrow extracted from them and then reinfusing the cells. Cline was immediately reprimanded for failing to secure the necessary permission from his home institution’s Institutional Review Board to carry out the work and having insufficient animal data to demonstrate the effectiveness of his procedure. The incident cost Cline his university chair and most of his funding from the NIH. It also ignited a furious public debate about the social and ethical implications of gene therapy. This led to the tightening up of regulations for the future testing of gene therapy in humans, which were to be overseen by the NIH’s Recombinant DNA Advisory Committee (RAC).

Gene therapy entered a new era in the 1980s following the discovery of retroviruses which proved a much more efficient tool for gene transfer. The first suitable retroviral vector for gene therapy was developed by Richard Mulligan, a researcher at Massachusetts Institute of Technology and former doctoral student of Paul Berg, a key pioneer in genetic engineering at Stanford University. By 1983 Mulligan had managed to genetically modify a mouse leukemia retrovirus with his colleagues so that it could deliver any desired DNA without reproducing in humans. The new vector also contained a selective marker, a piece of DNA from Escherichia coli bacteria, which made it possible to identify how many genes a cell picked up during gene transfer.

One of the first people to use Mulligan’s new vector was French Anderson, a geneticist at the NIH’s National Heart, Lung and Blood Institute. By 1989 he had secured permission from the RAC to begin the first approved clinical trial with gene therapy. This was to be done with the help of Michael Blease, a paediatrician and immunologist. The team’s aim was to test gene therapy in children with severe combined immunodeficiency, an inherited immune disorder caused by a defective adenosine deaminase (ADA) gene. Most children born with the disorder did not live long and only survived by being confined in sterile plastic enclosures, giving rise to the term ‘bubble disease’. Those with the condition had only two treatment options. The first was to have a bone marrow transplant, but this was hampered by the need to find a matching donor and the risks of an immune reaction. The second was to have frequent injections of PEG-ADA, a synthetic enzyme. Children who had such treatment usually showed a marked improvement after the first injection but this was usually of short duration and subsequent doses were largely ineffective.

Prior to treating the children the team partnered with Steven Rosenberg at the National Cancer Institute (NCI) conducted a test of their proposed procedure in a 52 year old man dying from malignant melanoma in May 1989. This was designed to assess three things: assess the safety of Mulligan’s retroviral vector, determine how much of the marked gene it could transfer and how long the gene lasted. The experiment involved a number of stages. In the first instance, the scientists needed to cultivate tumour infiltrating lymphocytes (TIL cells), a type of tumour-killing cell. This involved incubating white blood cells removed from the man’s tumour with interleukin-2, a molecule found to activate T in the destruction of cancer cells in the 1960s. A DNA marker was then inserted into the TIL cells before they were reinfused into the patient. The same procedure was repeated in seven more patients at the NCI with terminal malignant melanoma. Encouragingly all of the patients absorbed the marker genes with no ill-effects and a third of them responded positively to the treatment. One experienced a near-complete remission. The study marked a major turning point. Firstly, it established the feasibility and safety of gene therapy. Secondly, it opened the door to the development of gene therapy for cancer.

Anderson’s team started trying out the gene therapy in children with ADA-SCID in early 1990. The first patient to receive the therapy was Ashanti DeSilva, a four year old girl. Her treatment lasted twelve days. It necessitating extracting Ashanti’s blood cells, inserting a new working copy of the ADA gene into them and then reinfusing the cells into her. Overall, the procedure was similar to a bone marrow transplant. The goal was to replenish Ashanti’s blood cells with ones that could produce ADA. Gene therapy had the advantage that the cells originated from Ashanti so there was no risk of rejection. To everyone’s delight Ashanti improved so much she no longer needed to be kept in isolation and was able to start school. She remains alive to this day.

Numerous gene therapy trials were launched in the 1990s in the light of the success with Ashanti. A significant shift took place during this decade. Critically the field moved away from just looking to treat rare diseases caused by a single gene, as had been the case with Ashanti. By 2000 gene therapy had been tried out in nearly 3,000 patients in almost 400 trials. Most of the trials targeted cancer, but cardiovascular disease, AIDS, cystic fibrosis and Gaucher disease were also investigated.

Some of the early enthusiasm for gene therapy witnessed at the beginning of the decade, however, had begun to disappear by the end of the 1990s. This was because researchers struggled to get the therapy to work because of the inefficiency of the retroviral vectors they had to hand. Negative attitudes to gene therapy increased following the first death in a trial. In September 1999, Jesse Gelsinger, an 18 year old American died while taking part as a volunteer in a dosing escalation trial. Led by James M Wilson, the trial was designed to treat newborn infants with a fatal inherited a metabolic disorder, known as ornithine transcarbamylase deficiency, which leads to the buildup of excessive ammonia in the body. Gelsinger had himself been born with the condition, but had managed to keep it in check by restricting his diet and taking special medications. He was allocated to the last group in the trial who received the highest dose. Four days after treatment Gelsinger died from major organ failure because of his violent immune reaction to the vector used in the treatment. The vector was derived from adenovirus, a group of viruses first isolated from the tonsils and adenoid tissue of children in the early 1950s. One of the reasons such a virus was used was because such viruses were well characterised and had a small genome so were easy to manipulate. Moreover, most people carry adenoviruses without experiencing any significant clinical symptoms. Investigations into Gelsinger’s death revealed insufficient care had been taken during the trial and poor clarity in terms of its safety guidelines.

While the tragedy led to the enforcement of more stringent regulations for gene therapy trials, Gelsinger was not the last to suffer the consequences of an adenoviral vector. Three years later, in 2002, a number of British and French children were discovered to have developed T cell leukaemia three years after receiving gene therapy for a form of SCID linked to a defect on the X chromosome. Their cancer turned out to have been caused by an adenoviral vector that integrated into a part of their genome that activated a gene for leukaemia. This too the scientists by total surprise because most adenoviruses are unable to integrate into the host genome.

Despite the difficulties, gene therapy began to turn a corner the following decade, aided by the arrival of safer and more effective vectors. Positive results began to be reported for a number of gene therapy trials. Most were small-scale academic studies. In 2007 Jean Bennett, an ophthalmologist at the University of Pennsylvania, demonstrated in a small trial that gene therapy could provide a promising treatment for inherited retinal disease. Subsequent trials in more patients carried out in 2015 backed this up. In addition to eye disease, gene therapy was found to help haemophilic patients, a number of whom no longer needed to take blood clotting factor drugs. Good news also emerged in 2015 from trials of gene therapy for rare single-mutation blood diseases like thalassemia and sickle-cell anaemia, with some patients able to stay healthy without blood transfusions. A year later, two small trials showed gene therapy could help in the treatment of patients with cerebral adrenoleukodystrophy, an inherited disorder that affects the central nervous system, and with spinal muscular atrophy, a neuromuscular disease that is one of the leading causes of genetic death in infants.

The first gene therapy was licensed in China in 2003. Designed for the treatment of neck and head cancer, this treatment did not make it across to other countries. The first gene therapy was approved in Europe nine years later. It was developed by UniQure, a Dutch company for treating lipoprotein lipase deficiency, a rare metabolic disease that causes acute and recurrent abdominal pain and inflammation of the pancreas. The drug, however, failed to be a commercial success because too few patients needed the drug. This led to UniQure withdrawing marketing authorisation for the drug by 2017.

In 2016 Europe licensed a second gene therapy, developed by GlaxoSmithKline for children suffering from ADA-SCID. A year later Novartis secured approval for the first gene therapy in the United States. Designed to treat acute lymphoblastic leukaemia, the therapy had grown out of the preliminary work Anderson and Rosenberg had originally undertaken to establish the safety of gene therapy for treating children with ADA-SCID in 1989.

Application

Gene therapy takes different forms. It can involve the insertion of a copy of a new gene, modifying or inactivating a gene, or correcting a gene mutation. This is done with the help of a vector derived from a genetically modified virus. Several different viral vectors are now used for this purpose.

Adenoviral vectors are some of the most common ones. These vectors work best in nondividing cells such as found in the brain or retina. Lentiviral vectors are also popular. These are derived from lentiviruses, a group of retroviruses. Two of the most commonly used, which emerged in the late 1990s, are the human immunodeficiency virus and the herpes simplex virus. Such vectors have the advantage that they can carry large quantities of genes and work in non-dividing cells. Nonetheless, they, present some safety issues because it is difficult to predict where they will integrate into the host genome. For this reason, lentiviral vectors are generally deployed in the genetic alteration of cells extracted from patients. Lentiviral vectors are particularly helpful in the introduction of genes into the genome of cells that are generally difficult to modify. Lentiviral vectors made from the herpes simplex virus are currently being used in gene therapies being explored for pain and brain diseases.

New horizons have opened up for gene therapy with the recent development of CRISPR-Cas9, a much more precise technique for altering genes. At the end of 2016 a group of Chinese scientists, led by the oncologist Lu You at Sichuan University, launched a safety trial to see if it was possible to treat cancer patients by using CRISPR-Cas to disable a particular gene in their cells that codes for the PD1 protein which often impedes a cell’s immune response to cancer. A few months later, in 2017, a similar trial was initiated by an American team headed by Carl June at the University of Pennsylvania.

Issues

While gene therapy has made remarkable progress in the last few years, its development still raises significant questions in terms of safety. One of the major differences between gene therapy and conventional small molecule drugs or other biological products, like protein therapeutics, is that once gene therapy has been administered it is difficult to stop treatment. It is also too early to know how long the effects of a gene therapy last. Moreover, too few patients have been given gene therapy for any length of time to know whether it poses any safety risks long term.

Another major stumbling block is that so far the price of gene therapy has been incredibly high. Gene therapies are currently some of the most expensive treatments on the market. In part this reflects the fact that most of them are custom-made for individual patients.

This piece was written by Lara Marks in January 2018. It draws on the work of Courtney Addison and her chapter ‘Gene therapy: An evolving story’, in Lara V Marks, ed, Engineering Health: How biotechnology changed medicine, (Royal Society of Chemistry, October 2017).

Gene therapy: timeline of key events

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