How Does Gene Therapy Work?
Gene replacement therapy targets disease at the root cause—your DNA—by introducing healthy nucleotide sequences into cells. So how does gene therapy work?
What is Gene Therapy?
Gene therapy seeks to replace broken sections of DNA, or add new genes to faulty cells, to address the underlying cause of disease. It has applications in treating cancer, heart disease, diabetes, cystic fibrosis, haemophilia, and AIDS.
Let's look at the rationale. Your DNA lives inside every cell of your body, comprising 46 chromosomes, which in turn are made up of around 20,000 genes. They provide the blueprint your cells need to make life-sustaining proteins.
Your various cells makes around 20,000 proteins to make your body work. They run the show, keeping you alive on a moment-to-moment basis. Examples of proteins are insulin (turning glucose into energy), haemoglobin (transporting oxygen in the blood), and neurexin (connecting neurons at the synapse).
But when genes mutate, they can become faulty, affecting the ability of cells to produce healthy proteins (if at all). Disease is born. For instance, cystic fibrosis is a single-gene mutation that affects the production of the CFTR protein. A chain reaction ensues: chloride isn't moved around cells, water isn't attracted to the cell surface, and mucus in the organs becomes thick and sticky. In CF patients, this has disastrous effects on the lungs.
Gene therapy aims to replace such mutant genes with healthy genes. There are two main types:
Somatic Gene Therapy treats disease in an individual, by targeting malfunctioning body cells.
Germline Gene Therapy prevents disease in future offspring, by targeting cells that produce eggs and sperm.
It's an elegant solution, correcting myriad diseases at the most fundamental level. Yet gene therapy is still in its infancy. Certain aspects of the procedure have been described as crude and, as we'll see in a moment, vulnerable to catastrophic failure. Even so, scientists believe gene therapy will become a staple of 21st century medicine.
The First Gene Therapy Trial
In the 1980s, Ashanthi DeSilva was born without the ability to make the protein adenosine deaminase (ADA). She'd inherited broken ADA genes from both her mother and father. If she'd inherited just one mutant gene, she would have been ok. But two faulty genes left her with a critical shortage.
No-one knew this at the time. At birth, Ashanthi seemed like a healthy baby. But as she became exposed to common bacteria and viruses in the environment, her deficiency became manifest.
"Ashi had her first infection at just two days old. By the time she was walking, she was constantly hacking and dripping with coughs and colds." - Ricki Lewis, The Forever Fix: Gene Therapy and The Boy Who Saved It
The protein ADA plays a crucial role in our resistance to infections. Without it, Ashanthi was unable to produce healthy white blood cells, and her body quickly became overwhelmed by viruses and bacteria.
Most infants with ADA deficiency don't survive past their second birthday. Ashanthi was lucky. After she turned two, she was diagnosed with Severe Combined Immunodeficiency (SCID) and put on ADA replacement therapy. The missing protein was sourced from cows and infused into her bloodstream on a regular basis.
But for Ashanthi, the replacement protein was only partially effective in treating her SCID, and was also expensive as a long term treatment. Still, it kept her alive until she was four years old and recruited for a new trial that sought to replace the very genes that make that protein. It was the world's first gene therapy trial...
How Does Gene Therapy Work?
Let's examine how gene therapy works in the context of the groundbreaking gene therapy trial performed on four-year-old Ashanthi DeSilva in 1990.
Step 1. Harvest Target Cells from The Patient
Gene therapy starts by taking a sample of the patient's target cells. For Ashanthi, this meant collecting some of her malfunctioning white blood cells, which were then replicated in culture to scale the operation.
Step 2. Insert Healthy Genes into The Vector
Next you need a sample of healthy DNA. Ashanthi's doctors collected ADA-coding genes found on chromosome 20 and inserted them into a vector—a carrier vessel to transport the genes into the target cells.
In gene therapy, viruses devoid of their own genetic material are widely used as gene-delivery vehicles. Viruses evolved to transport viral DNA from host to host and inject it into cells for replication, making them a natural transport vessel for gene therapy.
Step 3. Combine The Vector with Target Cells
When introduced to the patient's target cells, the virus does what viruses do best: hijack cells. Using its biological entry codes, the virus gains access at the cell membrane and infiltrates the factory floor. Then it sneaks in its genetic payload for replication.
Besides curing horrible diseases, this is the coolest part of gene therapy. It actually hijacks the hijacker.
Ashanthi's gene edits were performed ex vivo (outside the body) before the cells were returned to her. Later therapies would be performed in vivo (inside the body) with fatal consequences, as we'll see below.
Step 4. Return Modified Cells to The Patient
Once the target cells have taken up the healthy genes, they're returned to the patient. Ashanthi received an infusion of a pint or so of murky fluid containing 10 million edited white blood cells. The infusion took just 28 minutes, marking the beginning of a new era in human gene therapy.
The trial was a success. The modified white blood cells produced ADA and were then able to make functional antibodies. They also went on to replicate with the healthy genes intact, providing long-term gene correction. Although she needed 10 more infusions, Ashanthi suffered no side effects, and was effectively cured of SCID over the next two years. Today, Ashanthi is alive and well, married and with a Masters in Public Policy.
That first gene therapy trial prompted researchers to treat newborn babies with ADA deficiency in the same way. They even took white blood cells straight from the umbilical cord, enabling the infants to produce healthy immune cells from the start of their lives.
That was more than 30 years ago. At the time, you'd be forgiven for thinking that gene therapy was on a fast-track to success; that we'd soon see it successfully treating other inherited diseases like cystic fibrosis, diabetes, haemophilia, and cancer. But you'd be wrong.
The Scientific Breakthrough Myth
When it comes to invention, Hollywood has taught us to imagine the lone scientist working late at night, hunched over a microscope. Suddenly, a test tube starts fizzing and he's got his winning formula.
The unsexy reality is it often takes many teams of researchers decades to develop new treatments and introduce them safely into widespread use.
When the first bone marrow transplant was performed in 1957, the survival rate was less than 1%. It was 30 years before immune suppressing drugs emerged, bringing the survival rate to 35-90% depending on the donor.
Chemotherapy was another slow burner. Chemotherapy was invented when, during post-World War I autopsies, doctors found that soldiers poisoned with mustard gas had too few white blood cells. Even though the gas was a harmful toxin, it prompted researchers to investigate if it could reduce the excess white blood cells in leukaemia patients.
The first chemotherapies using mustard gas were introduced in the 1940s. However, toxicity issues persisted, and it was several decades before mustard gas became a viable treatment for leukaemia.
Gene therapy is turning out to be no different. Although some 4,000 patients would receive gene therapy in the decade after Ashanthi's successful treatment, the introduction of a different method would reveal the fatal risks of gene therapy.
When Gene Therapy Goes Wrong
In 1999, Jesse Gelsinger was an 18-year-old with a partial ornithine transcarbamylase (OTC) deficiency. It meant he was unable to digest protein in his diet. Most babies born with this disease suffer excessive build-up of ammonia and half of them die within a month.
With one healthy gene and one faulty gene, Jesse had a partial OTC deficiency. It enabled him to manage his condition with a low-protein diet and nearly 50 pills a day. It also made him eligible to take part in a gene therapy trial. He signed up not only in the hope of curing his own disease, but to help babies born with the fatal form.
The trial was designed to test the adenovirus as a gene vector, and whether it could be used safely in vivo targeting liver cells. The researchers knew that the adenovirus could stimulate an immune response even when inactive, and that adenoviruses are more dangerous in OTC-deficient patients. Yet the trial went ahead.
As the eighteenth and final patient in the trial, Jesse received the highest dose of the virus: 3.8x1013 (380,000,000,000,000) viral particles. This was orders of magnitude greater than the dose required for treatment.
From this massive viral load, Jesse had a violent immune response to the adenovirus. His body produced fever, blood clots, and widescale inflammation. Multiple organ failure ensued, and four days later, he died.
An FDA investigation and concluded multiple factors in the death of Jesse Gelsinger. As a result of his partial OTC deficiency, Jesse had high ammonia levels at the time of his treatment, which should have triggered exclusion protocols. Likewise, two previous gene therapy participants had suffered serious side effects from the adenovirus. And in earlier animal studies, three monkeys had died from blood clots and liver inflammation, all of which went unreported.
Early clinical trials are inherently dangerous; that's why trials are necessary. Yet multiple warning signs were present in Jesse's case, making his death an even crueller tragedy. And while the event led to tighter regulations for gene therapy trials, Jesse was not the last to suffer from this experimental treatment.
In 2002, a gene therapy trial in France treated children suffering from X-SCID, a form of Ashanthi's disease. As with Jesse, they used an adenovirus as the vector, and performed the therapy in vivo.
Initially, the children responded well to the treatment. But in the years that followed, 4 out of 9 infants in the trial developed leukaemia, a cancer leading to the excessive production of abnormal white blood cells.
It was a baffling result. The FDA temporarily halted all gene therapy trials in the US until the cause could be identified. Soon, researchers found that the adenovirus had inserted the healthy genes alongside cancer-causing genes. This activated the cancer genes which, along with other existing mutations, caused leukaemia to develop. This insertional mutagenesis can happen in nature when mediated by viruses, or artificially as we've now seen in gene therapy.
It's pretty damning when your experimental therapy starts giving children cancer. The episode was widely publicised, and the leukaemia ended up claiming the life of one of the French children. Gene therapy was halted in its tracks for some time. But, eventually, the trial led to modifications in procedures. It was concluded that all gene delivery models present the risk of insertional mutagenesis, and insulators were subsequently built into the virus to prevent it from activating other genes.
This is still very much a technology in development, and as new methods are trialled on different diseases, further risks remain unknown. But proponents of gene therapy argue that the diseases they're targeting are often fatal. Gene therapy still offers the best chance of survival for children who are otherwise running out of time.
Gene Therapy Trials Today
The history of gene therapy trials is riddled with these setbacks—and we may still be in the thick of it. In 2021, several trials for sickle-cell disease were shut down when three patients developed cancer. Sometimes these adverse events take years to come to light.
"Most gene therapies are designed to achieve permanent or long-lasting effects in the human body, and this inherently increases the risk of delayed adverse events." - Gene Therapy Needs a Long Term Approach, Nature
Yet clinical studies are on the rise. The US performs two-thirds of gene therapy trials in the world today, and by 2025, the FDA is expected to approve 10-20 gene therapies per year. Will clinical frameworks keep up with this pace to ensure long term safety?
We are still delving into unknowns. Today, gene transfer technology uses two types of vectors: viral and non-viral. Among viral vectors, the retrovirus and the adenovirus are the most effective, with work ongoing to re-engineer them to improve safety and efficacy.
The newer, non-viral vectors come in various forms: polymers, lipids, and ultrasound-mediated microbubbles (gas-filled spheres) which can deliver healthy gene payloads into cells. These systems are currently less efficient, but they are thought to be safer, cheaper, and present no DNA size limit. Researchers are also exploring recently FDA-approved nanomedicines such as gold nanoparticles and quantum dots.
Ultimately, if scientists tread lightly, gene therapy could be used safely to treat a wide range of diseases. Many trials are now targeted at cancer because of its widespread incidence. Monogenic diseases (those caused by a single-gene mutation) are other major targets. Gene replacements for cardiovascular, infectious, and inflammatory diseases are also sought, with unique protocols needed for each new treatment.
But as with the development of any new therapy, there are always risks and challenges to overcome. That is what most people, and perhaps many trial participants, don't fully recognise. Gene therapy trials are a necessary gamble, producing inevitable victims along the way to cures that will one day save countless other lives.