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November 1, 2021

Four Success Stories in Gene Therapy

The field is beginning to fulfill its potential. These therapies offer a glimpse of what’s to come

By Jim Daley

Design Cells Getty Images

After numerous setbacks at the turn of the century, gene therapy is treating diseases ranging from neuromuscular disorders to cancer to blindness. The success is often qualified, however. Some of these therapies have proved effective at alleviating disease but come with a high price tag and other accessibility issues: Even when people know that a protocol exists for their disease and even if they can afford it or have an insurance company that will cover the cost—which can range from $400,000 to $2 million—they may not be able to travel to the few academic centers that offer it. Other therapies alleviate symptoms but don’t eliminate the underlying cause.

“Completely curing patients is obviously going to be a huge success, but it’s not [yet] an achievable aim in a lot of situations,” says Julie Crudele, a neurologist and gene therapy researcher at the University of Washington. Still, even limited advances pave the way for ongoing progress, she adds, pointing to research in her patients who have Duchenne muscular dystrophy: “In most of these clinical trials, we learn important things.”

Thanks to that new knowledge and steadfast investigations, gene therapy researchers can now point to a growing list of successful gene therapies. Here are four of the most promising.

Gene Swaps to Prevent Vision Loss

Some babies are born with severe vision loss caused by retinal diseases that once led inevitably to total blindness. Today some of them can benefit from a gene therapy created by wife-and-husband team Jean Bennett and Albert Maguire, who are now ophthalmologists at the University of Pennsylvania.

When the pair first began researching retinal disease in 1991, none of the genes now known to cause vision loss and blindness had been identified. In 1993 researchers identified one potential target gene, RPE65 . Seven years later Bennett and Maguire tested a therapy targeting that gene in three dogs with severe vision loss—it restored vision for all three.

In humans, the inherited condition that best corresponds with the dogs’ vision loss is Leber congenital amaurosis (LCA). LCA prevents the retina, a layer of light-sensitive cells at the back of the eye, from properly reacting or sending signals to the brain when a photon strikes it. The condition can cause uncontrolled shaking of the eye (nystagmus), prevents pupils from responding to light and typically results in total blindness by age 40. Researchers have linked the disease to mutations or deletions in any one of 27 genes associated with retinal development and function. Until gene therapy, there was no cure.

Mutations in RPE65 are just one cause of inherited retinal dystrophy, but it was a cause that Bennett and Maguire could act on. The researchers used a harmless adeno-associated virus (AAV), which they programmed to find retinal cells and insert a healthy version of the gene, and injected it into a patient’s eye directly underneath the retina. In 2017, after a series of clinical trials, the Food and Drug Administration approved voretigene neparvovecrzyl (marketed as Luxturna) for the treatment of any heritable retinal dystrophy caused by the mutated RPE65 gene, including LCA type 2 and retinitis pigmentosa, another congenital eye disease that affects photoreceptors in the retina. Luxturna was the first FDA-approved in vivo gene therapy, which is delivered to target cells inside the body (previously approved ex vivo therapies deliver the genetic material to target cells in samples collected from the body, which are then reinjected).

Spark Therapeutics, the company that makes Luxturna, estimates that about 6,000 people worldwide and between 1,000 and 2,000 in the U.S. may be eligible for its treatment—few enough that Luxturna was granted “orphan drug” status, a designation that the FDA uses to incentivize development of treatments for rare diseases. That wasn’t enough to bring the cost down. The therapy is priced at about $425,000 per injection, or nearly $1 million for both eyes. Despite the cost, Maguire says, “I have not yet seen anybody in the U.S. who hasn’t gotten access based on inability to pay.”

Those treated show significant improvement: Patients who were once unable to see clearly had their vision restored, often very quickly. Some reported that, after the injections, they could see stars for the first time.

While it is unclear how long the effects will last, follow-up data published in 2017 showed that all 20 patients treated with Luxturna in a phase 3 trial had retained their improved vision three years later. Bennett says five-year follow-up with 29 patients, which is currently undergoing peer review, showed similarly successful results. “These people can now do things they never could have dreamed of doing, and they’re more independent and enjoying life.”

Training the Immune System to Fight Cancer

Gene therapy has made inroads against cancer, too. An approach known as chimeric antigen receptor (CAR) T cell therapy works by programming a patient’s immune cells to recognize and target cells with cancerous mutations. Steven Rosenberg, chief of surgery at the National Cancer Institute, helped to develop the therapy and published the first successful results in a 2010 study for the treatment of lymphoma.

“That patient had massive amounts of disease in his chest and his belly, and he underwent a complete regression,” Rosenberg says—a regression that has now lasted 11 years and counting.

CAR T cell therapy takes advantage of white blood cells, called T cells, that serve as the first line of defense against pathogens. The approach uses a patient’s own T cells, which are removed and genetically altered so they can build receptors specific to cancer cells. Once infused back into the patient, the modified T cells, which now have the ability to recognize and attack cancerous cells, reproduce and remain on alert for future encounters.

In 2016 researchers at the University of Pennsylvania reported results from a CAR T cell treatment, called tisagenlecleucel, for acute lymphoblastic leukemia (ALL), one of the most common childhood cancers. In patients with ALL, mutations in the DNA of bone marrow cells cause them to produce massive quantities of lymphoblasts, or undeveloped white blood cells, which accumulate in the bloodstream. The disease progresses rapidly: adults face a low likelihood of cure, and fewer than half survive more than five years after diagnosis.

When directed against ALL, CAR T cells are ruthlessly efficient—a single modified T cell can kill as many as 100,000 lymphoblasts. In the University of Pennsylvania study, 29 out of 52 ALL patients treated with tisagenlecleucel went into sustained remission. Based on that study’s results, the FDA approved the therapy (produced by Novartis as Kymriah) for treating ALL, and the following year the agency approved it for use against diffuse large B cell lymphoma. The one-time procedure costs upward of $475,000.

CAR T cell therapy is not without risk. It can cause severe side effects, including cytokine release syndrome (CRS), a dangerous inflammatory response that ranges from mild flulike symptoms in less severe cases to multiorgan failure and even death. CRS isn’t specific to CAR T therapy: Researchers first observed it in the 1990s as a side effect of antibody therapies used in organ transplants. Today, with a combination of newer drugs and vigilance, doctors better understand how far they can push treatment without triggering CRS. Rosenberg says that “we know how to deal with side effects as soon as they occur, and serious illness and death from cytokine release syndrome have dropped drastically from the earliest days.”

Through 2020, the remission rate among ALL patients treated with Kymriah was about 85 percent. More than half had no relapses after a year. Novartis plans to track outcomes of all patients who received the therapy for 15 years to better understand how long it remains effective.

Precision Editing for Blood Disorders

One new arrival to the gene therapy scene is being watched particularly closely: in vivo gene editing using a system called CRISPR, which has become one of the most promising gene therapies since Jennifer Doudna and Emmanuelle Charpentier discovered it in 2012—a feat for which they shared the 2020 Nobel Prize in Chemistry. The first results from a small clinical trial aimed at treating sickle cell disease and a closely related disorder, called beta thalassemia, were published this past June.

Sickle cell disease affects millions of people worldwide and causes the production of crescent-shaped red blood cells that are stickier and more rigid than healthy cells, which can lead to anemia and life-threatening health crises. Beta thalassemia, which affects millions more, occurs when a different mutation causes someone’s body to produce less hemoglobin, the iron-rich protein that allows red blood cells to carry oxygen. Bone marrow transplants may offer a cure for those who can find matching donors, but otherwise treatments for both consist primarily of blood transfusions and medications to treat associated complications.

Both sickle cell disease and beta thalassemia are caused by heritable, single-gene mutations, making them good candidates for gene-editing therapy. The method, CRISPR-Cas9, uses DNA sequences from bacteria (clustered regularly interspaced short palindromic repeats, or CRISPR) and a CRISPR-associated enzyme (Cas for short) to edit the patient’s genome. The CRISPR sequences are transcribed onto RNA that locates and identifies DNA sequences to blame for a particular condition. When packaged together with Cas9, transcribed RNA locates the target sequence, and Cas9 snips it out of the DNA, thereby repairing or deactivating the problematic gene.

At a conference this past June, Vertex Pharmaceuticals and CRISPR Therapeutics announced unpublished results from a clinical trial of beta thalassemia and sickle cell patients treated with CTX001, a CRISPR-Cas9-based therapy. In both cases, the therapy does not shut off a target gene but instead delivers a gene that boosts production of healthy fetal hemoglobin—a gene normally turned off shortly after birth. Fifteen people with beta thalassemia were treated with CTX001; after three months or more, all 15 showed rapidly improved hemoglobin levels and no longer required blood transfusions. Seven people with severe sickle cell disease received the same treatment, all of whom showed increased levels of hemoglobin and reported at least three months without severe pain. More than a year later those improvements persisted in five subjects with beta thalassemia and two with sickle cell. The trial is ongoing, and patients are still being enrolled. A Vertex spokesperson says it hopes to enroll 45 patients in all and file for U.S. approval as early as 2022.

Derailing a Potentially Lethal Illness

Spinal muscular atrophy (SMA) is a neurodegenerative disease in which motor neurons—the nerves that control muscle movement and that connect the spinal cord to muscles and organs—degrade, malfunction and die. It is typically diagnosed in infants and toddlers. The underlying cause is a genetic mutation that inhibits production of a protein involved in building and maintaining those motor neurons.

The four types of SMA are ranked by severity and related to how much motor neuron protein a person’s cells can still produce. In the most severe or type I cases, even the most basic functions, such as breathing, sitting and swallowing, prove extremely challenging. Infants diagnosed with type I SMA have historically had a 90 percent mortality rate by one year.

Adrian Krainer, a biochemist at Cold Spring Harbor Laboratory, first grew interested in SMA when he attended a National Institutes of Health workshop in 1999. At the time, Krainer was investigating how RNA mutations cause cancer and genetic diseases when they disrupt a process called splicing, and researchers suspected that a defect in the process might be at the root of SMA. When RNA is transcribed from the DNA template, it needs to be edited or “spliced” into messenger RNA (mRNA) before it can guide protein production. During that editing process, some sequences are cut out (introns), and those that remain (exons) are strung together.

Krainer realized that there were similarities between the defects associated with SMA and one of the mechanisms he had been studying—namely, a mistake that occurs when an important exon is inadvertently lost during RNA splicing. People with SMA were missing one of these crucial gene sequences, called SMN1 .

“If we could figure out why this exon was being skipped and if we could find a solution for that, then presumably this could help all the [SMA] patients,” Krainer says. The solution he and his colleagues hit on, antisense therapy, employs single strands of synthetic nucleotides to deliver genetic instructions directly to cells in the body [see “ The Gene Fix ”]. In SMA’s case, the instructions induce a different motor neuron gene, SMN2 , which normally produces small amounts of the missing motor neuron protein, to produce much more of it and effectively fill in for SMN1 . The first clinical trial to test the approach began in 2010, and by 2016 the FDA approved nusinersen (marketed as Spinraza). Because the therapy does not incorporate itself into the genome, it must be administered every four months to maintain protein production. And it is staggeringly expensive: a single Spinraza treatment costs as much as $750,000 in the first year and $375,000 annually thereafter.

Since 2016, more than 10,000 people have been treated with it worldwide. Although Spinraza can’t restore completely normal motor function (a single motor neuron gene just can’t produce enough protein for that), it can help children with any of the four types of SMA live longer and more active lives. In many cases, Spinraza has improved patients’ motor function, allowing even those with more severe cases to breathe, swallow and sit upright on their own. “The most striking results are in patients who are being treated very shortly after birth, when they have a genetic diagnosis through newborn screening,” Krainer says. “Then, you can actually prevent the onset of the disease—for several years and hopefully forever.”

This article is part of “ Innovations In: Gene Therapy ,” an editorially independent special report that was produced with financial support from Pfizer .

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• Published: 05 August 2021

Record number of gene-therapy trials, despite setbacks

• Carrie Arnold 1  

Nature Medicine volume  27 ,  pages 1312–1315 ( 2021 ) Cite this article

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The recent failure of a gene therapy for Huntington’s disease was devastating for patients, but researchers remain optimistic.

For decades, Claudia Testa has longed to tell her patients with Huntington’s disease that she has a treatment for them. The neurologist at Virginia Commonwealth University could prescribe medications to help control the psychiatric symptoms of the deadly neurodegenerative condition, and to manage its characteristic jerking and writhing, but when it came to drugs that could slow the disease’s progression, Testa had nothing. Neither did anyone else.

This is why Testa and so many others in the Huntington’s community were avidly following the progress of a phase 3 trial for tominersen . The drug is an antisense oligonucleotide (ASO), a short, single-stranded piece of DNA designed to bind to a specific mRNA target. Tominersen, developed by Ionis Pharmaceuticals and licensed to Roche, binds to the mRNA encoding the mutant huntingtin protein and targets it for degradation by the cell. Tominersen sailed through phase 1/2 trials showing it was safe and that it lowered huntingtin levels. So when Roche announced in March 2021 that it was pulling the plug on the phase 3 trial of tominersen, Testa and others were devastated.

“This resets the timeline,” she says. The field was hopeful that 2021 would be the first year they would have a real disease-modifying therapeutic, Testa says. “And now we’re back to infinity.”

An interim analysis of the trial data showed no difference between the patients who received an intrathecal infusion of tominersen every 16 weeks and those who received the placebo. Worse, the patients who received tominersen every 8 weeks appeared to be getting sicker than the control participants.

“They made the only decision they could possibly make. They had to stop,” Testa says.

At the same time, a smaller phase 1/2 trial of an ASO for treating patients with Huntington’s disease by Wave Therapeutics (from whom Testa receives consulting fees) was also shuttered after preliminary results showed the compound failed to decrease levels of mutant huntingtin. The company is now throwing its support behind a newer, potentially more effective ASO. As the Huntington’s community reeled from back-to-back blows, two other biotechnology companies—Cambridge, Massachusetts–based Voyager Therapeutics , and uniQure in Amsterdam—moved forward with gene-therapy trials that use adeno-associated virus (AAV) vectors to deliver microRNAs that are designed to reduce toxic levels of mutant huntingtin protein.

This rollercoaster ride is emblematic of the gene-therapy field’s recent growing pains, although safety concerns and lack of efficacy have not stopped the explosive growth in trials. Clinicaltrials.gov lists nearly 5,000 gene-therapy trials, and more than 100 trials of ASOs from around the world, more than ever before. But the long-term safety of some of these therapies remains unclear.

Everything stopped

In the late 1990s, gene therapy seemed just around the corner. Scientists had nearly finished sequencing the human genome, and genetic engineering allowed them to transform viruses that caused diseases such as the common cold and even AIDS into delivery vehicles for life-saving gene therapies. The idea that researchers could cure even the deadliest of human diseases did not sound so far-fetched. Then everything went horribly wrong.

In 1999, 18-year-old Jesse Gelsinger received an injection of an adenovirus vector that carried a corrected copy of his mutated gene encoding ornithine transcarbamylase, which caused a rare X-linked liver disorder. The adenovirus vector sparked an overwhelming immune response that led to his death four days later . The US Food and Drug Administration (FDA) placed an immediate clinical hold on gene-therapy trials, and overnight, the field went from sizzling to frigid.

“Everything just seemed to stop,” says Roland Herzog , an immunologist at Indiana University, who was a postdoctoral fellow at Children’s Hospital of Philadelphia at the time.

The problem was not the new gene to be delivered but instead the vector delivering the gene. Finding a way to safely and effectively introduce a novel gene (or gene-editing system) into the body’s cells continues to be the lynchpin of any gene-therapy endeavor, says Jennifer Hamilton, a postdoctoral fellow in the lab of Nobel Prize–winner Jennifer Doudna at the University of California, Berkeley.

“Delivery is the key thing that needs to be considered for these types of approaches,” she says.

The adenovirus vector’s ability to elicit a robust immune response is partly the reason vaccinologists have used adenovirus to build the ChAdOx1 vaccine against COVID-19. But this immunogenicity has led researchers to turn to other viruses to deliver gene therapy. The fact that scientists would continue to rely on viruses was no surprise, according to Hamilton. “Viruses are well evolved to get genes from outside the cell to inside,” she says.

Instead of adenovirus, they began experimenting with engineered lentiviruses such as HIV and with AAVs. “AAVs were a huge advance. It changed the whole landscape of gene therapy,” says Mark Sands , a gene-therapy expert at Washington University School of Medicine in St. Louis.

AAV continues to be the most popular vector partly due to its multiplicity of serotypes, which are able to infect nearly every type of tissue, even non-dividing cells. Crucially, AAV, unlike adenovirus, rarely elicits a strong immune response, so researchers could give patients the high doses of the virus that are often needed to get the correct gene into an adequate number of cells.

“It’s amazing that, for the most part, you can actually get away with these high doses. It tells you how benign this vector is,” Herzog says.

Over time, more safety information was gathered from preclinical trials, and regulators gained experience evaluating this information, says Peter Marks, Director of the Center for Biologics Evaluation and Research at the FDA. After more than a decade, gene therapy seemed poised to take its first hesitant steps forward. This, along with the advent of CRISPR-based gene-editing tools, meant that at first a few trials launched under nervous eyes, then hundreds. The field seemed unstoppable.

Then, yet again, everything went horribly wrong.

Too high a dose

On 6 May 2020, alerts from Nicole Paulk’s mobile phone woke the gene-therapy expert at the University of California, San Francisco, several hours before dawn. Colleagues had questions about the death of a participant in the ASPIRO gene-therapy trial by Audentes Therapeutics. The Bay Area biotechnology company (now part of the Japanese pharmaceutical company Astellas Pharma) had developed a gene therapy for the rare neuromuscular disease X-linked myotubular myopathy. They delivered a very high dose of AAV: 3 × 10 14 vector genomes per kilogram of body weight. The boy subsequently died as a result of sepsis .

Seven weeks later, a second boy who had received the same high dose of the same AAV gene therapy died. On 20 August, so did a third boy—all three died of sepsis. To Paulk, the deaths were a stark warning about the potential dangers of high-dose AAVs.

“I don’t think anyone expected such a profound response that couldn’t be controlled,” she says.

The question on her mind—on everyone’s mind—was whether the field was having another ‘Jesse Gelsinger moment’. As in the Gelsinger case, scientists had noted severe toxicity in preclinical tests when non-human primates were given high doses of the therapy. High-dose AAVs had also caused liver and immune system toxicities in animal models and in humans given gene therapy for Duchenne muscular dystrophy.

“Back when the original events occurred at the turn of the millennium, we did not have the same amount of experience at FDA,” Marks says. “The scientific knowledge and regulatory maturation have given us more confidence. It’s why these deaths didn’t stop the field like it did in 2000.”

In December 2020, the FDA lifted its clinical hold on the ASPIRO trials. “The exact biological mechanism that led to the patients’ deaths has not been conclusively determined,” says Edward Conner, Astellas Gene Therapies Site Lead. “Within the comprehensive review, Astellas has not identified clinical evidence, either direct or indirect, that immune responses contributed to the liver injury.”

A detailed analysis of the company’s internal findings was published . Still, scientists held their breath, and fears grew as serious adverse events appeared in other gene-therapy trials. In December 2020, Amsterdam-based uniQure reported that one person in its phase 3 trial of an AAV5-based therapy for hemophilia B had developed hepatocellular carcinoma (in March 2021, an independent investigation cleared the vector of any role). A uniQure spokesperson said that detailed analysis of the patient’s liver tissue revealed a precancerous state due to several pre-existing conditions.

At the University of Pennsylvania, Denise Sabatino and colleagues published a paper in Nature Biotechnology showing potential liver problems that had emerged in dogs nearly a decade after they were treated with an AAV gene therapy for canine hemophilia A. In February, the team at bluebird bio announced two cases of acute myeloid leukemia, as well as one incidence of myelodysplastic syndrome (a diagnosis later revised to transfusion-dependent anemia), in participants in its trial of LentiGlobin gene therapy for sickle-cell disease. And in late April 2021, Adverum Biotechnologies announced that a participant in its trial for diabetic macular edema had developed vision loss in the dosed eye.

Unexpected integration

Despite these setbacks, and despite COVID-19, the number of new gene-therapy trials has sped up in the past year. Sands wonders if this is wise.

After Gelsinger’s death, gene-therapy experts began to rely on AAVs as the vector of choice. This small virus seemed like the workhorse the field was looking for. But even as AAV was being hailed as the savior of gene therapy, Sands had noticed some long-term safety issues.

Sands found a hepatocellular carcinoma in several neonatal mice treated with an AAV-based gene therapy, used to treat a metabolic disorder. After Sands published his findings in 2001, however, his conclusions were questioned because previous trials had dosed older mice with AAVs without an apparent problem.

“There were a lot of companies that were springing up and there were a lot of people’s reputations on the line,” Sands says. “Everyone said these vectors were benign and here I come along and say wait a minute. There was enormous pushback.”

Six years of arduous bench work finally yielded the specific sites where the AAV vector had integrated itself into the mouse genome. Sands and colleagues discovered that the viruses had inserted themselves into a 6-kilobase region on chromosome 12, altering the expression of host genes and leading to tumorigenesis. To Sands, this highlights the need for long-term safety studies on AAV-based gene therapy so the field can move forward safely.

“I am a huge proponent of AAV therapy. But my concern has always been that if [integration] is a problem for human gene therapy, it should have been studied in a systematic way for the past 20 years,” he says. “If we don’t understand how integration is happening, we can’t re-engineer the AAVs to make them safer.”

Sabatino agrees. Her work found that the AAV vectors given to treat canine hemophilia sometimes integrated into the dogs’ liver cells, creating clonal expansions that have the potential to become cancerous. But these problems take years to develop in humans. Problems like these are not picked up by tests in non-human primates, Sabatino says, because these animals develop an immune response to the human gene therapy. This means their bodies fight off the virus and its accompanying genetic payload, so monitoring generally lasts for only a few months.

“This is the value of long-term animal models for follow-up. You can’t track a monkey for several months and expect to find a clonal expansion,” she says.

The FDA’s Center for Biologics Evaluation and Research , which oversees gene-therapy trials and approvals, is aware of the potential for long-term issues with gene therapy, Marks says. The FDA is now recommending 15 years of follow-up in its draft gene-therapy guidance for industry.

“We have to have safety of these products, and that’s why we are here at FDA, to make sure that this is done safely,” Marks says.

Alison Bateman-House , a bioethicist at New York University, agrees that long-term animal studies are needed. Without knowing the full magnitude of potential risk, Bateman-House says, patients cannot provide true informed consent for clinical trials.

“We’re really flying blind with this,” she says.

Where do you want it?

Sands and Sabatino remain optimistic about the promise of AAVs and other gene therapies. To Paulk, the challenge will be designing better recombinant vectors with improved tropism and decreased immunogenicity so that patients need less vector and have a reduced chance of a severe immune reaction.

At Affinia Therapeutics, based in Waltham, Massachusetts, Chief Scientific Officer Charles Albright says that the company is working to engineer precisely these types of improved vectors. One vector has 32 times more RNA expression, which allows use of a reduced dose; another vector was tweaked both to steer it away from the liver and again to steer it towards muscle.

“This is very advantageous,” says Albright. “To detarget the liver and increase uptake into muscle—this specific combination will be very powerful if we can translate it to non-human primates.”

Ongoing safety concerns about AAV-based gene therapies have led some scientists rethink how gene therapy is delivered.

In November 2020, Intellia Therapeutics began a phase 1 trial of the world’s first systemically administered CRISPR–Cas9 therapy, for hereditary transthyretin amyloidosis with polyneuropathy. Instead of packaging the gene-editing system in a viral vector, Intellia is using a lipid nanoparticle, somewhat akin to the method used in the mRNA vaccines against SARS-CoV-2 developed by Moderna and BioNTech–Pfizer. Preclinical research showed that the CRISPR–Cas9 therapy knocked down more than 97% of the misfolded transthyretin protein. The advantage to this method, according to Hamilton, is that patients can receive multiple doses over time, since their immune systems will not form antibodies against the vector. In late June 2021, Intellia published the first results of the trial , which showed a dose-dependent drop in serum transthyretin levels.

Hamilton is working to create a different type of CRISPR–Cas9 system that delivers the gene-editing system as a ribonucleoprotein complex that can enter the targeted cell, edit the genome, and then degrade. Not only does this transient dosing strategy prevent the development of antibodies against a viral vector, its short-lived time in the cell also means the immune system will not raise antibodies against Cas9, Hamilton says.

Ex vivo CRISPR has its own disadvantages, because it requires the removal of hematopoietic stem cells for editing in the lab, which are then returned as a stem-cell transplant. Trial participants must undergo treatment with a myeloablative agent such as busulfan, which destroys their own stem cells, requiring weeks of hospitalization. Ex vivo CRISPR gene editing is therefore not an option for low resource settings.

“Low-income countries in sub-Saharan Africa tend to lack the significant infrastructure required to broadly provide bone marrow transplants, let alone gene therapy, which requires the cells to be removed, shipped, genetically modified, and returned to the patient,” says Betsy Foss-Campbell, Director of Policy and Advocacy at the American Society of Gene and Cell Therapy.

Equity of therapy

With an eye toward addressing this challenge, the Bill & Melinda Gates Foundation partnered with Novartis in February 2021 to bring a single-administration, in vivo gene therapy for sickle-cell disease to Africa, where most of the people with this condition live. This therapy will need to be delivered directly to the patient, without the need to modify cells in the lab, and without the need for lengthy and expensive hospitalization. If everything goes to plan, says Susan Stevenson, Executive Director at the Novartis Institute for Biomedical Research, a sickle-cell-disease gene therapy may be ready for human trials in a little over 5 years.

“It’s a great model to follow. Take the treatment where the majority of the patients are,” she says.

From his lab at Pontificia Universidad Javeriana in Bogotá, Carlos Javier Alméciga Díaz has been following advances in gene therapy with interest. He studies a group of hereditary lysosomal disorders that are fatal in childhood. Colombia has the world’s highest rates of a mucopolysaccharide storage disease known as Morquio A syndrome. Díaz says that although Morquio A syndrome is the perfect candidate for gene therapy, getting interest from the pharmaceutical industry and securing funding remains a challenge , because the condition is vanishingly rare elsewhere in the world. What the field needs, he says, are more local solutions, developed for specific populations affected by specific diseases.

“This is not like a regular drug. This is not like an aspirin or a paracetamol that you can give to the patient and say, okay, take two pills every few hours. This is a different treatment,” Díaz says.

Far from being hamstrung by recent setbacks and challenges, the gene-therapy field remains hopeful and optimistic. There are substantial challenges in terms of improving delivery and affordability, as well as monitoring long-term safety, but to the scientists, they do not seem insurmountable.

“Even with all the disappointment and pain, it’s just an incredibly exciting time in this field,” Testa says.

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Addressing the dark matter of gene therapy: technical and ethical barriers to clinical application

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• Volume 141 , pages 1175–1193, ( 2022 )

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• Landon J. Getz   ORCID: orcid.org/0000-0002-3841-1062 2   na1 ,
• Thibaut Peterlini 3 , 4 ,
• Jean-Yves Masson   ORCID: orcid.org/0000-0002-4403-7169 3 , 4 &
• Graham Dellaire   ORCID: orcid.org/0000-0002-3466-6316 1 , 2 , 5  

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Gene therapies for genetic diseases have been sought for decades, and the relatively recent development of the CRISPR/Cas9 gene-editing system has encouraged a new wave of interest in the field. There have nonetheless been significant setbacks to gene therapy, including unintended biological consequences, ethical scandals, and death. The major focus of research has been on technological problems such as delivery, potential immune responses, and both on and off-target effects in an effort to avoid negative clinical outcomes. While the field has concentrated on how we can better achieve gene therapies and gene editing techniques, there has been less focus on when and why we should use such technology. Here we combine discussion of both the technical and ethical barriers to the widespread clinical application of gene therapy and gene editing, providing a resource for gene therapy experts and novices alike. We discuss ethical problems and solutions, using cystic fibrosis and beta-thalassemia as case studies where gene therapy might be suitable, and provide examples of situations where human germline gene editing may be ethically permissible. Using such examples, we propose criteria to guide researchers and clinicians in deciding whether or not to pursue gene therapy as a treatment. Finally, we summarize how current progress in the field adheres to principles of biomedical ethics and highlight how this approach might fall short of ethical rigour using examples in the bioethics literature. Ultimately by addressing both the technical and ethical aspects of gene therapy and editing, new frameworks can be developed for the fair application of these potentially life-saving treatments.

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Acknowledgements

We gratefully acknowledge feedback during the preparation of this review from Sabateeshan Mathavarajah and Dr. Françoise Baylis.

K.K. is a trainee of the Cancer Research Training Program of the Beatrice Hunter Cancer Research Institute, with funds provided by the Dalhousie Medical Research Foundation (DMRF) C. MacDougall Cancer Research Studentship, as well as being supported by a Genomics in Medicine Graduate Studentship from the Dalhousie Faculty of Medicine –DMRF and a Nova Scotia Scholar Award. L.J.G. is funded by a Vanier Canadian Graduate Scholarship from the Natural Science and Engineering Research Council of Canada (NSERC) as well as a Killam Pre-Doctoral Scholarship from the Killam Trusts. J-Y.M. is a Canada Research Chair in DNA repair and Cancer Therapeutics. This work is also supported by a Canadian Institutes of Health Research (CIHR) Project Grant (PJT-156017) to J-Y.M. and G.D.

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Kateryna Kratzer and Landon J. Getz contributed equally to the work.

Authors and Affiliations

Department of Pathology, Faculty of Medicine, Dalhousie University, PO BOX 15000, Halifax, NS, B3H 4R2, Canada

Kateryna Kratzer & Graham Dellaire

Department of Microbiology and Immunology, Faculty of Medicine, Dalhousie University, PO BOX 15000, Halifax, NS, B3H 4R2, Canada

Landon J. Getz & Graham Dellaire

Genome Stability Laboratory, Oncology Division, CHU de Québec Research Centre, Quebec, Canada

Thibaut Peterlini & Jean-Yves Masson

Department of Molecular Biology, Medical Biochemistry and Pathology, Laval University Cancer Research Center, 9 McMahon, Quebec, G1R 3S3, Canada

Department of Biochemistry and Molecular Biology, Faculty of Medicine, Dalhousie University, Halifax, NS, Canada

Graham Dellaire

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KK and GD conceived of the review. KK, LG and GD generated Figs. 1 and 3 , and J-YM and TP generated Figs. 2 and 4 . All authors contributed to the writing and edited of the manuscript.

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Kratzer, K., Getz, L.J., Peterlini, T. et al. Addressing the dark matter of gene therapy: technical and ethical barriers to clinical application. Hum Genet 141 , 1175–1193 (2022). https://doi.org/10.1007/s00439-021-02272-5

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How gene editing therapies could go beyond rare diseases.

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Rahman Oladigbolu, a 52-year-old Harvard-educated film maker with sickle cell disease, poses for a ... [+] photo. Born in Nigeria, he is very interested in Casgevy, the CRISPR-based sickle cell treatment that was recently approved by the FDA. He has had both his knees, shoulders and hips replaced because of SCD. (Photo by Jonathan Wiggs/The Boston Globe via Getty Images)

Earlier this year, a group of scientists in the Netherlands used the gene editing tool CRISPR to eliminate HIV from immune cells in the lab, an eye-catching approach that is forming the basis of potentially curative therapies for the disease.

HIV impacts an estimated 39 million people around the world and while it is no longer a death sentence thanks to antiretroviral drugs, there is still no recognized cure. Research groups around the world believe that gene editing - which seeks to disable the virus through cutting large swaths of its genetic code - could finally offer a solution.

However, translating promising results in cells or animals to humans is always a big leap, and earlier this month it was revealed that a pioneering clinical trial run by San Francisco-based biotech Excision BioTherapeutics had ended in relative failure. The company released interim data from five HIV patients who had participated in its Phase 1 trial, showing that while using gene editing to make cuts in the HIV genome seemed to be relatively safe, it did not lead to meaningful suppression of the virus.

The company is planning to refine the approach in a future clinical trial using a different method of delivering CRISPR to patient immune cells. Elena Herrera-Carrillo, an assistant professor at the University of Amsterdam who led the Dutch HIV study, is still hopeful gene editing will provide a pathway to a cure.

One day, she believes, CRISPR could offer a new way of targeting a range of chronic viral infections, not just HIV. “With its precise gene-editing capabilities, CRISPR can potential target and disrupt viral genomes, both DNA and RNA, offering new avenues for the treatment and prevention of infectious diseases,” she says. “It has been utilized to target the SARS-CoV-2 genome, and it could be employed to combat hepatitis B.”

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The Excision BioTherapeutics trial is a particularly big step because until now, the majority of CRISPR applications have been targeted at rare diseases. In December, CRISPR Therapeutics and Vertex Pharmaceuticals made history when their gene editing therapy Casgevy was approved by the Food and Drug Administration as a treatment for the blood disorders beta thalassemia and sickle cell disease. While this marked the first time that a CRISPR-based therapy was greenlighted by regulators in the U.S., there are thought to be only 16,000 patients who are eligible to receive the treatment.

Yet with the next generation of gene editing tools also on the horizon, many patients could benefit in the long run from this nascent field. Metagenomi, a biotech based in Emeryville, California that received an investment from Leaps in 2020 and went public earlier this year, has a series of preclinical programs targeting diseases ranging from familial and spontaneous ALS to cardiovascular disease and cystic fibrosis. (I remain a board member at Metagenomi.)

“We’ve just seen this initial approval, and some of the other first generation gene editing companies are doing exciting work, such as Intellia Therapeutics and Verve Therapeutics, but it’s just getting started,” says Brian C. Thomas, a former UC Berkeley academic who is now CEO and Founder of Metagenomi. “In the coming years it’s not just going to be about one single gene editing tool, but a whole variety of tools which can be used to interact with the human genome to address a broad range of diseases. Being able to match the right tool to a given disease target is essential to this.”

Beyond Cas9

In recent years, scientists have used CRISPR-Cas9 to create new disease models to study neurodegenerative conditions such as Parkinson’s and Huntington’s , as well to discover gene targets that are key to cancer growth and metastasis.

Various biotech companies and academic researchers are now looking into the next generation of gene editing tools which may be safer and easier to deliver into the body. In the last four years, various genome engineering studies have investigated the possible applications of Cas11, Cas12 and even Cas13 enzymes, and companies such as Caszyme, which was co-founded by Professor Virginijus Šikšnys, one of the original pioneers of CRISPR, are actively pursuing these new molecular tools.

“We’re working on developing safer and smaller Cas nucleases that would be more compatible with diverse delivery technologies,” says Monika Paule, CEO of Caszyme.

At Metagenomi, Thomas predicts that the second generation of gene editing therapies will use CRISPR as a framework for other functions such as CRISPR-associated transposases (CASTs) which allow scientists to effectively cut-and-paste a large piece of DNA at a particular site in the genome.

He suggests that this could ultimately provide a single way of tackling complex diseases such as the many different forms of muscular dystrophy, which can be caused by hundreds of possible variants in the dystrophin gene.

“Because there are so many different variants, a monumental number of therapies would be required to treat all patients that suffer from that disease using current gene editing technologies,” he says. “But with a CAST system, you might be able to go in and address all underlying mutations with a single treatment by replacing the same chunk of problematic DNA in every patient.”

The Delivery Challenge

While CASTs are still a long way from the clinic, newer gene editing nucleases may help solve some of the delivery challenges that have contributed to the high cost of gene editing therapies along with existing safety concerns. Many of these novel approaches were the subject of considerable excitement at the recent American Society of Gene & Cell Therapy (ASGCT) annual meeting in Baltimore, which brought together some of the leading experts in the field.

Right now, patients who receive the Casgevy therapy, which costs more than $1 million, need to have stem cells extracted from their bone marrow, which are then modified outside their body, and reinfused back into the bone marrow, where they produce new blood cells that reduce their symptoms. However, more tolerable methods of delivering CRISPR into the body may be lipid nanoparticles or modified lentiviruses.

Thomas says CRISPR therapies delivered via lipid nanoparticles will soon allow scientists to target metabolic disorders that occur in the liver. He is even more excited by the potential offered by nucleases that are smaller and more versatile than Cas9, which could make it possible to penetrate further into the body. It is believed that such features will facilitate delivery of genome editing tools to previously inaccessible tissue types and organ systems.

“It’s very straightforward to take a lipid nanoparticle and get that into the liver,” he says. “But you can’t use that method for neurodegenerative diseases because it won’t pass the blood-brain barrier. Instead, you can use small viruses such as adeno-associated viruses (AAVs). They are limited in what they can carry but that’s where these smaller nucleases, which are a third of the size of Cas9, become really exciting. You can comfortably get them packaged into an AAV and start thinking about jumping outside the liver and treating these other diseases.”

As more clinical trials of CRISPR-based therapies start to happen and more receive regulatory approval, Thomas predicts that we will learn a tremendous amount about how to make these treatments effective and accessible for as many patients as possible.

The era of one-and-done gene editing therapies is just beginning. Once the science matures, I predict it will be nothing short of transformational.

Thank you to David Cox for additional research and reporting on this article.

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New gene therapy strategies for cancer treatment: a review of recent patents

Affiliation.

• 1 Department of Health Science, University of Jaén, Jaén; 23071, Spain.
• PMID: 22339358
• DOI: 10.2174/157489212801820093

Cancer is the second leading cause of death in the Western world. The limited successes of available treatments for cancer mean that new strategies need to be developed. The possibility of modifying the cancer cell with the introduction of genetic material opens the way to a new approach based on gene therapy. There are still many technical difficulties to be overcome, but recent advances in the molecular and cellular biology of gene transfer have made it likely that gene therapy will soon start to play an increasing role in clinical practice, particularly in the treatment of cancer. Gene therapy will probably be the therapeutic option in cases in which conventional treatments such as surgery, radiotherapy and chemotherapy have failed. The development of modified vectors, and an improved understanding of interactions between the vector and the human host, are generating inventions that are being protected by patents due to the considerable interest of industry for their possible commercialization. We review the latest strategies, patented and/or under clinical trial, in cancer gene therapy. These include patents that cover the use of modified vectors to increase the security and specificity, recombining adenovirus that leads to loss or gain of gene function, activation of the patient's own immune cells to eliminate cancer cells by expression of molecules that enhance immune responses, silencing genes related to the development of drug resistance in patients, inhibition of angiogenesis of solid tumors by targeting the tumor vasculature, and the development of enzymes that destroy viral or cancerous genetic material.

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• Research Support, Non-U.S. Gov't
• Gene Silencing / physiology
• Genes, Transgenic, Suicide / genetics
• Genes, Transgenic, Suicide / physiology
• Genetic Therapy / legislation & jurisprudence*
• Genetic Therapy / methods
• Genetic Therapy / trends*
• Genetic Vectors / genetics
• Genetic Vectors / physiology
• Immunomodulation / genetics
• Immunomodulation / physiology
• Models, Biological
• Neoplasms / therapy*
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Safety and Effectiveness of Gene Therapy

Andrew P. Byrnes, Ph.D.

Office of Tissues and Advanced Therapies Division of Cellular and Gene Therapies Gene Transfer and Immunogenicity Branch

[email protected]

Chief, Gene Transfer and Immunogenicity Branch

BS, MS, Yale University, Department of Molecular Biophysics and Biochemistry

PhD, Oxford University, Department of Human Anatomy

Postdoctoral Fellow, Johns Hopkins University, School of Hygiene and Public Health

General Overview

Gene therapy holds great promise for treating cancer, inherited disorders, and other diseases. Gene therapy uses carriers called 'vectors' to deliver genes to tissues where they are needed. Researchers are currently investigating the safety and effectiveness of a variety of different gene therapy vectors in hundreds of clinical trials in the US.

We are studying one type of commonly-used gene therapy vector that is made from a disabled cold virus -- the adenovirus vector. While adenovirus vectors are very efficient at delivering genes, adenovirus vectors are not always easy to target to the correct tissue. In addition, adenovirus vectors can cause toxic effects that limit the amount of vector that doctors can give to patients. We are particularly interested in how to safely deliver large amounts of adenovirus vectors intravenously, with the goal of specifically targeting tumors and other tissues.

New adenovirus gene therapy vectors are tested in animals before human clinical trials begin, and it is important for both researchers and the FDA to know how well these animal studies can predict safety. Thus, another of our major goals is to develop animal models that reliably predict the safety and effectiveness of adenovirus vectors in humans.

Our studies will help us to understand the mechanisms for adenovirus vector targeting and toxicity, and the relevance of animal models to human outcomes. This new knowledge will enable researchers to design safer and more effective gene therapy vectors.

Scientific Overview

Adenovirus (Ad) vectors have shown considerable promise in animal models and are currently being used in numerous clinical trials, especially for the therapy of cancer. We are interested in improving the safety and efficacy of Ad vectors, especially when administered through the vascular system. Certain properties of Ad vectors make them hazardous to administer intravenously in large doses, and our laboratory is trying to understand and fix this problem.

One of our major areas of interest is the innate immune response to Ad vectors. These rapid responses can cause serious toxicity and may severely limit the doses of Ad vectors that are safe to use. In addition, we are also studying how cells in the liver such as Kupffer cells and hepatocytes recognize Ad, since the liver is the major site at which Ad vectors are cleared from the circulation. A better understanding of these mechanisms will help us to develop strategies to improve vector efficacy and reduce toxicity. We will also gain a better understanding of the advantages and disadvantages of using different animal species to predict the behavior of Ad vectors in humans, which is particularly relevant to the regulatory work of the FDA.

Recent work from our lab and others has shown that Ad vectors are heavily influenced by plasma proteins that rapidly opsonize the vectors after intravenous injection. We found that natural IgM antibodies bind to Ad vectors, activate complement, and reduce liver transduction. Intriguingly, the Ad hexon protein specifically binds to coagulation factor X (FX), and we found that recruitment of FX by Ad vectors protects them against neutralization by complement. These findings show that Ad vectors recruit a number of plasma proteins that interact in complex ways with each other and with cells, and that these host proteins ultimately help to determine whether the vector successfully reaches its target.

In the long run, a better fundamental understanding of Ad vector biology will facilitate the design of safer Ad vectors that are easier to target. Better animal models will be important for testing novel vectors for safety and efficacy.

Publications

• PLoS Pathog 2022 Sep 26;18(9):e1010859 Binding of adenovirus species C hexon to prothrombin and the influence of hexon on vector properties in vitro and in vivo. Tian J, Xu Z, Moitra R, Palmer DJ, Ng P, Byrnes AP
• FEBS Lett 2019 Dec;593(24):3449-60 Interaction of adenovirus with antibodies, complement and coagulation factors. Allen RJ, Byrnes AP
• PLoS One 2018 Feb 5;13(2):e0192353 Hexons from adenovirus serotypes 5 and 48 differentially protect adenovirus vectors from neutralization by mouse and human serum. Harmon AW, Moitra R, Xu Z, Byrnes AP
• Methods Mol Biol 2017;1643:187-96 Evaluating the impact of natural IgM on adenovirus Type 5 gene therapy vectors. Xu Z, Tian J, Harmon AW, Byrnes AP
• J Control Release 2016 Aug 10;235:379-92 Substitution of blood coagulation factor X-binding to Ad5 by position-specific PEGylation: Ppeventing vector clearance and preserving infectivity. Krutzke L, Prill JM, Engler T, Schmidt CQ, Xu Z, Byrnes AP, Simmet T, Kreppel F
• J Virol 2015 Mar;89(6):3412-6 Impact of natural IgM concentration on gene therapy with adenovirus type 5 vectors. Qiu Q, Xu Z, Tian J, Moitra R, Gunti S, Notkins AL, Byrnes AP

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Gene Therapy for Genetic Syndromes: Understanding the Current State to Guide Future Care

Marian l. henderson.

1 The Department of Biology, Calvin University, Grand Rapids, MI 49546, USA; moc.kooltuo@0002ramedneh

2 Department of Pediatrics and Human Development, College of Human Medicine, Michigan State University, Grand Rapids, MI 48824, USA; ude.usm@cajabeiz (J.K.Z.); ude.usm@oaixil (X.L.); ude.usm@179bpmac (D.B.C.); ude.usm@4343lliw (M.R.W.); ude.usm@2nadtgov (D.L.V.); [email protected] (C.P.B.); [email protected] (S.R.); [email protected] (N.L.H.)

Jacob K. Zieba

Xiaopeng li, daniel b. campbell, michael r. williams, daniel l. vogt, caleb p. bupp.

3 Medical Genetics, Corewell Health, Grand Rapids, MI 49503, USA

Yvonne M. Edgerly

4 Office of Research, Corewell Health, Grand Rapids, MI 49503, USA; [email protected]

Surender Rajasekaran

5 Pediatric Intensive Care Unit, Helen DeVos Children’s Hospital, Corewell Health, Grand Rapids, MI 49503, USA

Nicholas L. Hartog

6 Allergy & Immunology, Corewell Health, Grand Rapids, MI 49503, USA

Jeremy W. Prokop

Jena m. krueger.

7 Department of Neurology, Helen DeVos Children’s Hospital, Corewell Health, Grand Rapids, MI 49503, USA

Associated Data

Not applicable.

Gene therapy holds promise as a life-changing option for individuals with genetic variants that give rise to disease. FDA-approved gene therapies for Spinal Muscular Atrophy (SMA), cerebral adrenoleukodystrophy, β-Thalassemia, hemophilia A/B, retinal dystrophy, and Duchenne Muscular Dystrophy have generated buzz around the ability to change the course of genetic syndromes. However, this excitement risks over-expansion into areas of genetic disease that may not fit the current state of gene therapy. While in situ (targeted to an area) and ex vivo (removal of cells, delivery, and administration of cells) approaches show promise, they have a limited target ability. Broader in vivo gene therapy trials have shown various continued challenges, including immune response, use of immune suppressants correlating to secondary infections, unknown outcomes of overexpression, and challenges in driving tissue-specific corrections. Viral delivery systems can be associated with adverse outcomes such as hepatotoxicity and lethality if uncontrolled. In some cases, these risks are far outweighed by the potentially lethal syndromes for which these systems are being developed. Therefore, it is critical to evaluate the field of genetic diseases to perform cost–benefit analyses for gene therapy. In this work, we present the current state while setting forth tools and resources to guide informed directions to avoid foreseeable issues in gene therapy that could prevent the field from continued success.

1. Introduction

With the discoveries that DNA codes for genes and that a DNA sequence can have variants that increase disease susceptibility, a future was envisioned in which modifying genetic material to reduce disease risk/progression is achievable. Multiple possibilities arose to modify genetic material ( Figure 1 ) [ 1 , 2 ], including taking cells out of the body to correct genetics followed by delivery back to the individual (ex vivo gene therapy), packaging material to make the changes systemically (in vivo gene therapy), or targeting a tissue or cell to be edited (in situ gene therapy). Gene therapy consists of packaging nucleic acids (plasmid, DNA, RNA, antisense oligonucleotides) or gene editing machinery such as clustered regularly interspaced short palindromic repeats—CRISPR- and CRISPR-associated protein 9 (Cas9)—with guide RNA within a particle, often formed by an attenuated virus or nanoparticle, and delivering it to a cell or tissue to modulate a desired gene [ 3 , 4 , 5 , 6 , 7 ]. While animal models showed incredible promise for gene therapy in the 1970s and 1980s, there were early signs of safety risks posed by delivering biomaterials to humans [ 8 ].

Schematic of three gene therapy approaches: in vivo, ex vivo, and in situ. Generated with BioRender.

One of the first human gene therapy clinical trials, completed in 1990 by Rosenberg et al., involved the transfer of tumor-infiltrating lymphocytes modified with a neomycin resistance gene via a retroviral vector to patients with advanced melanoma [ 9 ]. The success of this trial provided proof of concept for the clinical application of gene therapy. With that promise of gene therapy, it is rather surprising to follow the complex multiple-decade history of gene therapy setbacks and complications [ 1 ]. However, the excitement associated with gene therapy has finally translated into clinical utility within the past few years, with the FDA and other world regulators approving their use, opening the door for correcting or replacing broader disease genetics [ 2 ].

Within rare diseases, genomic sequencing has increased to identify pathogenic variants [ 5 , 6 ], which yields an increasing hope of gene therapy to correct the variants. Rare diseases account for USD 997 billion in healthcare costs annually, impacting 15.5 million people within the U.S. [ 10 ]. Internationally, the frequency of rare diseases is uncertain due to limitations in diagnosis, but estimates are greater than 100 million individuals. While each rare disease occurs in less than 200,000 individuals (United States) and in 1/2000 births (European Union) [ 11 ], more than 5000 unique, rare diseases add up to a considerable fraction of healthcare costs internationally [ 12 ]. As international sequencing initiatives have expanded, so has the number of diagnosed individuals for each rare disease, largely contributed to the sharing of flagged genomic variants across borders [ 13 , 14 , 15 ]. The International Rare Diseases Research Consortium (IRDiRC), founded in 2011, has set forth a critical mission of expanding therapeutics for international usage through integrating international efforts into funding within each country or foundation [ 16 , 17 ]. This international partnership highlights the growing efforts to expand access across borders, which is critical to growing the number of patients with each rare disease to grow the demand and offset drug development costs [ 18 ]. The international efforts must continue to translate the United States and European union clinical trials into cross-border initiatives to increase clinical trial implementation for rare diseases [ 19 ].

As diagnoses of rare diseases have improved with the implementation of genome sequencing [ 20 , 21 , 22 ], the knowledge of the exact variant for each individual yields details of how to best treat each case [ 23 , 24 , 25 ]. If a variant results in loss of function of a protein, it is possible to replace that protein with a functional gene (gene delivery) or remove the cell, followed by CRISPR editing. If a variant causes a gain of function, one can reduce the function using antisense oligonucleotides. Thus, rare diseases are one of the areas where gene therapy holds incredible promise. However, a balance must be maintained between evaluating gene therapy benefits and safety risks to have a sustainable gene therapy ecosystem moving forward. Within this review article, we address the field’s current state in rare diseases and provide insights and guidance to advance the clinical use of gene therapy sustainably and safely. The article consists of an analysis of gene therapy based on publications, funding, status of clinical trials, and approved clinical usages while expanding considerations for additional rare disease genes, immune modulation, cost of therapy, and the need for increased transparency. At the end, the work is concluded through a discussion of the current and future ethical considerations for gene therapy advancement.

2. Past and Current Work in Gene Therapy

2.1. publications.

The advancements and applications of gene therapy can be reflected in yearly publications ( Figure 2 ). Publications mentioning “gene therapy” date back to the 1970s (1922 total papers) but expanded rapidly in the 1990s (76,314 papers) to the 2000s (317,383 papers) and 2010s (637,126 papers). The number of papers per year seems to have stabilized at the beginning of the 2020s, with 2020 having 88,853 papers, 2021 having 98,207 papers, and 2022 having 99,992 papers. In 2022, the gene therapy papers reflected diverse topics based on a Web of Science analysis. These include general fields like genetic heredity, biochemistry, and pharmacology. More specialized fields such as oncology, immunology, and neurosciences rank the highest in 2022 publications ( Figure 2 ). There are a total of 802,029 papers for “gene therapy” and “Genetic Heredity” over all years, with 25,280 of those articles also containing “Rare Disease.” A similar search within PubMed for “gene therapy” and “rare disease” returns 16,032 papers.

Publications on “gene therapy.” The first panel shows the number of publications found on Web of Science per year for the search “gene therapy,” with every five years labeled in black. The number of publications in 2022 is in red. The second panel shows the breakdown of the top 20 research areas of the 2022 papers. The analysis was performed on 18 April 2023.

Literature analysis provides valuable insights, especially those of nucleotide delivery systems for studying animal modeling of rare diseases. In the 2000s, a strategy known as morpholino oligonucleotides was widely used in research to knockdown genes in animal models [ 26 ]. Building on the toxic nature of oligonucleotides in developmental studies [ 27 ], morpholinos were developed to inhibit gene translation using chemical alterations of the oligonucleotide that allow for complementation with the transcript to prevent ribosome engagement [ 28 ]. In 2000, these morpholinos were shown to be functional in the knockdown of zebrafish genes during development, mimicking rare disease phenotypes [ 29 ]. This novel animal modeling tool progressed with hundreds of papers defining knockdown to phenotype correlations for rare genetic disorders [ 30 ]. However, in 2007, the same group that had presented the promise of zebrafish morpholinos showed that the system also regulated the tumor protein p53 (TP53, coded by the p53 gene) cascade and induced phenotypes independent of the targeted morpholino [ 31 ], a finding also shown through small interfering RNA (siRNA) [ 32 ] and phosphorothioate-linked DNA [ 33 ]. While there are off-target oligonucleotide functions in gene regulation, the tools continue to be used through understanding mechanisms and the growth of control datasets [ 34 , 35 ]. For example, our group has shown morpholino use in zebrafish followed by human mRNA recovery allows for definitive outcomes of human genotype-to-phenotype insights and gene therapy modeling for kidney disease [ 36 ]. While these techniques are being phased out with newer CRISPR-based animal modeling [ 37 ], they still provide a valuable lesson in considering off-target impacts for delivering nucleic acids. These findings highlight the persistent need for refined knowledge of how foreign nucleotides can impact cellular processes to better predict unexpected, off-target outcomes.

2.2. Funding

Similar to publications, funding can establish the trajectory of the gene therapy field. The top funder of worldwide science, the National Institutes of Health (NIH), is experiencing rapid funding growth in “gene therapy,” based on an analysis of NIH reporter. Beginning in 2016, funding mentioning “gene therapy” could be found in the project terms of NIH grants ( Figure 3 ). In 2018, the term could be found in project abstracts, and in 2019 within project titles, with a fast elevation to the USD 8.279 billion in total funding for 2022. The 2022 levels of NIH funding broken down by institutes show the top to be the National Cancer Institute (NCI, USD 1.8 billion), followed by the National Institute of Allergy and Infectious Diseases (NIAID, USD 1.5 billion), National Heart Lung and Blood Institute (NHLBI, USD 885 million), and the National Institute of Aging (NIA, USD 669 million).

NIH funding mentioning “Gene Therapy.” The first panel shows the funding (in millions of USD) per year by the National Institutes of Health (NIH) mentioning the term “gene therapy” in various annotation bins (mentioned in project: gray—title, yellow—abstract, cyan—terms, red—any of the three). The total annotated funding in 2022 is in red text. The second panel shows the breakdown of the top NIH institutes of the 2022 NIH funding for “Gene Therapy.” Abbreviations: NCI—National Cancer Institute, NIAID—National Institute of Allergy and Infectious Diseases, NHLBI—National Heart, Lung, and Blood Institute, NIA—National Institute on Aging, NINDS—National Institute of Neurological Disorders and Stroke, NIDDK—National Institute of Diabetes and Digestive and Kidney Diseases, NIGMS—National Institute of General Medical Sciences, NEI—National Eye Institute, NIDA—National Institute on Drug Abuse, NICHD—Eunice Kennedy Shriver National Institute of Child Health and Human Development. The analysis was performed on 1 May 2023 using NIH reporter.

The top ten highest funded awards from NIH represent a diversity of institutes and initiatives ( Table 1 ). Many of these awards were for mRNA vaccine programs and testing sites (1ZIATR000437, 1U19AI171421, 1U19AI171443, 1U19AI171110, 1U19AI171954, 1U19AI171292, 1U19AI171403), which primarily reflects the SARS-CoV-2 pandemic response. This mRNA vaccine expansion is likely the most significant factor in the rapid funding investments for gene therapy in 2022. A few of these large projects also reflect the growth of gene therapy within oncology (75N91019D00024–0-759102200019–1, 1U24CA224319) and neurodegeneration (5U01AG059798, 1UF1NS131791, 5R01AG068319, 5U19NS120384).

Top ten highest NIH-funded projects mentioning “gene therapy”. The analysis was performed on 1 May 2023 using NIH reporter.

Further refining NIH investments using a co-search with “rare disease” identified 787 funded awards ( Figure 4 ) with 728 unique project numbers totaling USD 526,396,101. Of these awards, 276 are traditional R01 NIH research awards, summing USD 155,491,503 in research. Additional funding for gene therapy comes from intramural awards (ZIA, 76 awards, USD 109,812,041), contract awards (U54, 63 awards, USD 36,059,350; U01, 47 awards, USD 48,207,117), and small research pilot grants (R21, 57 awards, USD 13,314,891). There is a surprisingly low number amongst these awards of trainee funding, such as K08 clinician scientist awards (24 awards for USD 3,829,993), K23 patient-oriented training (12 awards, USD 2,235,855), F30/F31 predoctoral awards (18 awards, USD 755,681), and F32 postdoctoral awards (3 awards, USD 235,260). As gene therapy is one of the most promising clinical tools, there seems to be a need for elevating targeted training awards.

Word usage for “gene therapy” and “rare disease” within all NIH-funded projects. The analysis was performed using WordClouds.com. The first panel shows words enriched within the 787 funded project titles. The second panel shows words enriched from their public health relevance statements. The analysis was performed on 1 May 2023 using NIH reporter.

Based on the titles and the public health relevance statements of “gene therapy” and “rare disease” funded grants, there is a diverse clinical perspective ( Figure 4 ). The mention of genes within the abstracts of the projects also reflects this diverse perspective. From the list of genes, funding is in the areas of neuroscience ( TSC , MTOR , CLN1 , CMT1A , NF1 ), neurodegeneration ( APOE , TAU , TREM2 ), cancer ( RUNX1 , P53 , MDM2 , KRAS ), and cystic fibrosis ( CFTR ). As rare diseases are dispersed between the NIH units, with no primary home that focuses on all rare diseases as a single pathology, it is unsurprising that the funding is spread across different institutes. As gene therapy grows in development, it is critical to consider new cross-NIH initiatives focusing on funding gene therapy advancements, especially those outside of oncology or vaccine designs. As the new ARPA-H (Advanced Research Projects Agency for Health) is established in the United States, gene therapy will likely be a significant component of agency design.

2.3. Clinical Trials

The translational advancement of gene therapy is reflected through clinical trials and approved therapies. Since 1990, the field has proliferated, with over 2000 completed clinical trials registered on ClinicalTrials.gov ( Figure 5 , as of 18 April 2023). The first registered trial returned for “gene therapy” (ClinicalTrials.gov Identifier: {"type":"clinical-trial","attrs":{"text":"NCT00001166","term_id":"NCT00001166"}} NCT00001166 ) was initiated in 1978, an observational trial of “Gyrate Atrophy of the Choroid and Retina” that focused on genetic determination of disease. In the 1990s, trials began expanding, with fast growth in the 2010s to the 2022 level of 397 trials initiated ( Figure 5 A). Of the studies, 37% of the total results have been marked as completed, 21% are recruiting, 9% are active but not recruiting, 9% have been terminated, and 5% are not yet recruiting ( Figure 5 B).

Analysis of ClinicalTrials.gov for “gene therapy.” All analyses were performed on 18 April 2023 using the ClinicalTrial.gov site. ( A ) Number of trials started each year, with the 2022 number in red. ( B ) Breakdown of trial status. Groups below 2% are not shown. ( C ) Breakdown of completed trials for FDA phase and age group inclusion. ( D ) Breakdown of the delivery system used, with a call out of adeno-associated virus subtypes shown to the right.

Among the completed studies, cancer was the primary target of most trials, as determined by an analysis of the disease categories provided by ClinicalTrials.gov. Monogenic disorders follow. The FDA breaks human clinical trials into four phases [ 38 ]. Phase I trials aim to answer whether a treatment may be given safely, assessing for toxicity in a small population. Phase II trials determine if the treatment is effective at different dosages, such as if adequate protein expression is obtained following gene therapy. Phase III trials compare the new treatment to those already utilized or a placebo, often in a larger population, where it is determined if it will be sent for federal approval as a new therapy. Once approved, interventions are continually monitored for adverse events in phase IV. Rare disease trials, particularly those utilizing gene therapy, often combine phases I and II and utilize a stepwise approach.

Among the clinical trials returned when searching for “gene therapy” and marked as complete, most fall under phase I or II trials ( Figure 5 C). In addition, most of these were only tested in adults (18 years and older). Viral vectors are the most utilized delivery system in gene therapy clinical trials, the most common being adenoviruses, retroviruses, lentiviruses, and adeno-associated viruses ( Figure 5 D). Of the adeno-associated viruses, AAV2 and AAV5 were the most selected for use. Plasmid DNA delivery, lipofection, and RNA transfer are the most utilized among nonviral vectors.

As “gene therapy” returns trial data irrelevant to interventions, we further filtered genetic diseases with intervention therapies ( Table 2 ). Multiple disorders have completed phase III trials, including cystic fibrosis, hemophilia B, retinal dystrophy, cerebral adrenoleukodystrophy, Spinal Muscular Atrophy (SMA), and β-Thalassemia. It should be noted that enrollment numbers are minimal for many rare diseases due to the low frequency of disorders within the population. This makes it challenging to build placebo control systems and generate sufficient data for FDA approval processes. These issues suggest the need for thoughtful reconsiderations in gene therapy authorization processes in the future [ 39 ] in addition to international cooperation efforts.

Top genetic diseases with interventional “gene therapy” clinical trials. A “-“ is used in the FDA-authorized treatment column when no treatments are authorized. The “*” indicates drugs that are not gene therapy.

Table 3 shows a curated list of phase III trials with gene therapy for rare diseases. Among these nine completed studies, four were for SMA using Onasemnogene Abeparvovec (also known as Zolgensma) for different inclusion criteria ( {"type":"clinical-trial","attrs":{"text":"NCT03306277","term_id":"NCT03306277"}} NCT03306277 , {"type":"clinical-trial","attrs":{"text":"NCT03461289","term_id":"NCT03461289"}} NCT03461289 , {"type":"clinical-trial","attrs":{"text":"NCT03505099","term_id":"NCT03505099"}} NCT03505099 , {"type":"clinical-trial","attrs":{"text":"NCT03837184","term_id":"NCT03837184"}} NCT03837184 ). SMA is characterized by an autosomal recessive dysfunction to exons 7 and 8 of the SMN1 gene, resulting in progressive spinal cord motor neuron degeneration and muscle atrophy [ 40 ]. Type 1 SMA decreases muscle tone so severely that children are never able to sit independently. Without intervention, type 1 SMA patients die of respiratory failure prior to their second birthday. The known genetic mechanisms and the progressive debilitating phenotype have resulted in SMA inclusion in many newborn screenings for early detection before the phenotype manifests [ 41 ], making it a compelling target for gene therapy intervention. {"type":"clinical-trial","attrs":{"text":"NCT03306277","term_id":"NCT03306277"}} NCT03306277 , known as STR1VE, was the first completed gene therapy phase III study, showing in 22 participants that a single AAV9 cDNA intravenous delivery of the SMN1 gene (Zolgensma) could prevent the phenotype of SMA type 1 [ 42 ]. Of the 22 participants, 3 were withdrawn, with 1 due to an unrelated death and 1 due to an adverse event. Of the patients enrolled, they had an average age of 3.7 months at gene delivery, with half identifying as white and 12 as female. All patients with therapy showed marked clinical improvement and achieved independent sitting at 18 months. Of the 22 individuals, 4 showed signs of respiratory distress, 1 with signs of secondary sepsis, and 2 with hepatic elevated enzymes. Presymptomatic genetically screened SMN1 variant-positive individuals were assessed for earlier delivery of this therapy ( {"type":"clinical-trial","attrs":{"text":"NCT03505099","term_id":"NCT03505099"}} NCT03505099 ), where all 14 patients had marked clinical improvements [ 43 ].

Curated phase III intervention studies for genetic syndromes.

Additional phase III trials have been completed for Choroideremia, cerebral adrenoleukodystrophy, β-Thalassemia, and Leber Hereditary Optic Neuropathy. {"type":"clinical-trial","attrs":{"text":"NCT03496012","term_id":"NCT03496012"}} NCT03496012 showed that a single-dose delivery of an AAV2-encoded REP1 gene targeted to the eye (in situ) with local injections was able to prevent monogenic inherited retinal dystrophies [ 44 ]. {"type":"clinical-trial","attrs":{"text":"NCT01896102","term_id":"NCT01896102"}} NCT01896102 showed the ex vivo delivery of CD34+ stem cells treated with lentiviral encoded ABCD1 to treat males with cerebral adrenoleukodystrophy [ 45 ]. Within that study, there was one reported death, 47% of individuals identified as white, all patients were males, and there were eight events of febrile neutropenia, six with a severe fever, and an extensive list of nonserious adverse events. {"type":"clinical-trial","attrs":{"text":"NCT02906202","term_id":"NCT02906202"}} NCT02906202 and {"type":"clinical-trial","attrs":{"text":"NCT03207009","term_id":"NCT03207009"}} NCT03207009 showed the ex vivo delivery of CD34+ stem cells treated with lentiviral encoded βA-T87Q-Globin gene for β-Thalassemia, with a 91% success rate of individuals showing transfusion independence [ 46 ]. Four individuals had adverse events, including one case of thrombocytopenia. {"type":"clinical-trial","attrs":{"text":"NCT03406104","term_id":"NCT03406104"}} NCT03406104 showed the intravitreal delivery (in situ) of the AAV2-encoded ND4 gene to improve vision in individuals with Leber Hereditary Optic Neuropathy [ 47 ]. In summary, it should be noted that SMA therapy is the only completed phase III trial with in vivo intravenous gene therapy results.

{"type":"clinical-trial","attrs":{"text":"NCT00073463","term_id":"NCT00073463"}} NCT00073463 started in 2003, aiming to test 100 participants age 12 or older for aerosolized AAV-encoded CFTR for the treatment of cystic fibrosis. While the phase I and II studies for this aerosolized therapy showed safety [ 48 , 49 ], the phase III trial showed no improvement in lung function [ 50 ]. The trial was terminated with the last enrolled participant in October 2005.

Below is a description of active trials with posted or published results, focusing on serious adverse responses reported. {"type":"clinical-trial","attrs":{"text":"NCT00999609","term_id":"NCT00999609"}} NCT00999609 used subretinal-injected AAV2-encoded RPE65 to treat retinal dystrophy in 21 patients, where two of the cases showed adverse drug reactions, and one individual showed convulsions [ 51 ]. {"type":"clinical-trial","attrs":{"text":"NCT03370913","term_id":"NCT03370913"}} NCT03370913 , {"type":"clinical-trial","attrs":{"text":"NCT03392974","term_id":"NCT03392974"}} NCT03392974 , and {"type":"clinical-trial","attrs":{"text":"NCT04323098","term_id":"NCT04323098"}} NCT04323098 showed the use of AAV5-encoded Coagulation Factor VIII infusion in 134 males with hemophilia A, where 22 serious adverse events were reported [ 52 ]. {"type":"clinical-trial","attrs":{"text":"NCT03569891","term_id":"NCT03569891"}} NCT03569891 used AAV5-encoded Human Factor IX infusion (Hemgenix, etranacogene dezaparvovec) to treat 67 males with hemophilia B, with five severe events, including acute myocardial infarction, gastrointestinal hemorrhage, pseudarthrosis, and acute kidney injury. In nearly all of the recruiting studies, there is a lack of posted results, meaning until completed, most gene therapy clinical trials lack reported data on adverse events. A commonality of gene therapy studies is the prescreening of antibodies towards the AAV system with no reported issues with immunosuppressive agents.

2.4. Approved Therapies

The FDA classifies gene therapy products in combination with cellular therapies within the Office of Tissues and Advanced Therapies, where there are 32 approved licensed products (as of 2 August 2023), 8 of which are gene therapies.

Two therapies have been authorized for SMA treatment: Spinraza and Zolgensma. Spinraza (Nusinersen, Biogen) is an antisense oligonucleotide that targets the SMN2 gene to alter splicing to recover SMN protein function [ 53 ]. The phase III trial ( {"type":"clinical-trial","attrs":{"text":"NCT02292537","term_id":"NCT02292537"}} NCT02292537 ) for Spinraza showed success in preventing SMA in 84 patients, with severe adverse events similar to sham control [ 54 ]. It should be noted that Spinraza is delivered intrathecally to the cerebral spinal fluid, and one case of post-lumbar puncture syndrome was noted in the clinical trial. Spinraza requires repeat dosing every four months indefinitely to maintain clinical benefits. Spinraza therapy was submitted to the FDA and approved on 23 December 2016 under a fast-track and orphan drug designation. Zolgensma (Onasemnogene Abeparvovec, Novartis Gene Therapies Inc.) was submitted to the FDA on 1 October 2018 and approved on 24 May 2019, creating an intravenous gene therapy for SMA. Zolgensma is a functioning copy of the full human SMN1 gene, which codes for the SMN protein that is lacking in SMA patients. Zolgensma currently requires only one dose.

Elevidys (delandistrogene moxeparvovec-rokl, Sarepta Theraputics, Inc.) was submitted to the FDA on 28 September 2022 and approved on 22 June 2023 for the treatment of Duchenne Muscular Dystrophy. Approval was limited to ambulatory patients aged 4–5 years. Elevidys utilizes an adeno-associated viral vector (AAVrh74) to deliver a portion of the dystrophin gene “microdystrophin.”. Sarepta was approved under accelerated status by demonstrating that patients treated with Elevidys had increased microdystrophin expression. It was noted in a published FDA summary memo that the decision for approval went against the recommendations made by the Clinical, Clinical Pharmacology, and Statistics review teams, who did not feel the data submitted showed a definite clinical benefit. Elevidys was approved with the contingency that further clinical trial data would be submitted.

Hemgenix (etranacogene dezaparvovec-drlb, CSL Behring LLC) was submitted to the FDA on 24 March 2022 and approved on 22 November 2022 for the treatment of hemophilia B. Luxturna (voretigene neparvovec-rzyl, Spark Therapeutics Inc.) was submitted to the FDA on 16 May 2017 and approved on 18 December 2017 for the treatment of biallelic RPE65 mutation-associated retinal dystrophy. Skysona (elivaldogene autotemcel, bluebird bio Inc.) was submitted to the FDA on 18 October 2021 and approved on 16 September 2022 to treat active cerebral adrenoleukodystrophy. Zynteglo (betibeglogene autotemcel, bluebird bio Inc.) was submitted to the FDA on 20 September 2021 and approved on 19 August 2022 to treat ß-Thalassemia. Roctavian (valoctocogene roxaparvovec-rvox) was submitted to the FDA on 23 December 2019 (resubmitted 29 September 2022) and approved on 29 June 2023 to treat severe hemophilia A only in the absence of AAV-5 preexisting antibodies. Vyjuvek (beremagene geperpavec) was submitted to the FDA on 20 June 2022 and approved on 19 May 2023 for the treatment of those >6 months of age with dystrophic epidermolysis bullosa due to COL7A1 variants. It should be noted that Vyjuvek is the first ever approved topical gene therapy and utilizes a herpes simplex virus type 1 (HSV-1) delivery system. HSV-1 is optimal for skin delivery as the virus naturally infects skin cells.

In the case of many of these FDA-approved therapies, their phase III trials continued after their authorizations, with an expectation of progression into phase IV studies.

While gene therapy in cystic fibrosis has had mixed results, it should be noted that small molecule regulators of the CFTR gene have proven that nucleotide delivery is not the only approach to modify gene expression in rare diseases. The FDA approved Elexacaftor–tezacaftor–ivacaftor, also known as triple therapy, which is recommended in patients with at least one copy of Phe508del CFTR variants [ 55 , 56 ]. Cystic fibrosis is an example where strategies outside of gene therapy should be continued in parallel, setting a critical mission that gene therapy trials do not overpower or result in underfunding small-molecule or other therapeutic approaches.

3. Biological Considerations

For effective gene therapy, one must confidently identify a causal gene, package that gene into a delivery system expressing the right amount in the right tissue/cell, and replace or repair the molecular mechanism with a measurable phenotype. This must be achieved while avoiding unforeseen biological challenges of viral vectors and overexpression of mRNA within cells. Below, we provide several areas of consideration for expanding gene therapy into additional clinical genetics.

3.1. Genetic Syndromes

The OMIM database ( https://www.omim.org/ ) [ 57 ] represents a catalog of human genetic conditions. As of April 2023, the database contained >6000 gene-to-disease correlations. These correlations represent 4771 unique human genes on all human chromosomes ( Figure 6 A). Using the UniProt database of protein annotations [ 58 ], it is evident that only a few represent DNA binding factors or have annotated domains like a zinc finger or coiled-coil segment ( Figure 6 B). A significant portion of these proteins are transmembrane, suggesting they localize to the surface of a cell. Many proteins have catalytic activity, binding sites, and active sites. In some rare and genetic diseases, the active site becomes hyperactive, where inhibitors can ameliorate disease. Most diseases manifest from loss-of-function to protein biology and thus need correctors instead of inhibitors.

OMIM genes connecting human genotypes to phenotypes. ( A ) Number of OMIM genes per chromosome. ( B ) The number of OMIM genes with various human UniProt annotations. ( C ) Tissue- or ( D ) single-cell-specific expression annotation from the Human Protein Atlas for each of the OMIM genes. ( E ) The number of OMIM genes with various International Mouse Phenotyping Consortium (IMPC) annotations following knockout and phenotyping. ( F ) Each OMIM gene number of IMPC phenotypes altered in knockout (x-axis) relative to the % of datasets from the Human Protein Atlas where the gene is expressed >1 transcript per million (TPM). ( G ) The amino acid length of each OMIM gene (x-axis) relative to the number of ClinVar annotated pathogenic or likely pathogenic variants. ( H ) The percent of each variant class relative to variant alterations for the ClinVar database.

Using the Human Protein Atlas (HPA) database [ 59 ], it is observed that most of the genes are ubiquitously expressed in human tissues ( Figure 6 C). At the same time, they have more specificity when annotated based on cell types within each tissue ( Figure 6 D). This observation suggests that we should not address tissue specificity for each gene but rather cell type specificity, where emerging tools like single-cell transcriptomics are opening new doors for these insights. Of the OMIM genes, 2398 have been knocked out in a mouse model, can be purchased for lab use, and have undergone extensive phenotypic analysis based on the International Mouse Phenotyping Consortium (IMPC, Figure 6 E) [ 60 ]. A total of 90% (2158/2398) of these genes show at least one observable phenotype altered by removing the gene, many matching the known human conditions, where these animals can serve as a pre-clinical gene therapy testing system.

It should be noted that 341 gene knockouts from the IMPC result in heterogeneous preweaning lethality (incomplete penetrance), and 131 are highly penetrant for lethality. The heterogeneity within phenotypes for genetic diseases represents one of the most considerable challenges in gene therapy; namely, how can one develop clinical trials to know success when phenotypes are not always predictable with our current state of knowledge.

It should be noted that the number of datasets showing gene expression within the HPA has little correlation to the number of altered phenotypes observed in the IMPC (R 2 of 2 × 10 −5 , Figure 6 F). This points to the need for further tools in genotype-to-phenotype predictions that will strengthen our ability to know when and how gene therapies may apply to an individual.

Many gene therapy delivery systems have a limited size of the genetic insert, with most of the OMIM genes within this window ( Figure 6 G). The largest database of human genetics, ClinVar [ 61 ], shows that of these OMIM genes, we have an array of known confident pathogenic variants ( Figure 6 G). While our pathogenic and likely pathogenic variants usually are significant changes to proteins (frameshift and nonsense variants), the current state of research is challenged by missense genetic changes and whether they confidently result in disease states ( Figure 6 H). Gene therapy can only be employed in high-confidence situations. Thus, the million plus variants of uncertain significance (VUSs) in OMIM genes would have a low probability of successful clinical trials, primarily if implemented based on newborn screening. This finding highlights that variant characterization remains a significant challenge in gene therapy expansion for genetic syndromes.

3.2. Cell and Promoter Specificity

Gene therapy is targeted to cell types based on the vector used to deliver the nucleic acids and sequences that can drive the expression of each gene only within that tissue/cell type, such as a cell-specific promoter element. The control of expression enables each gene to be made into mRNA and protein only in a specific cell type. To minimize the size of expression regulation sequences, promoters rather than enhancers are often used to achieve cell-type specificity [ 62 ]. Since the advent of RNA sequencing, there has been an expansion in defining tissue/cell-specific expression. Still, more recently, with techniques such as single-cell RNA sequencing, we are now resolving specificity in the different functional cell types within each tissue. This specificity of expression is critical to controlling many OMIM genes contributing to developmental pathways. The HPA annotation of cell specificity for 75 different cell types shows 1908 different human genes with highly specific expression within one of the cell types ( Figure 7 ). More work is needed to determine which promoter elements may work, independent of cell-type-specific enhancers, for the desired tissue of an OMIM gene being nominated for gene therapy.

Heatmap of expression for cell-type-specific genes from the Human Protein Atlas. Red indicates the highest expression in the row. Dendrograms show one minus Spearman rank correlation with cell type on top and genes shown to the left.

It should be noted that the developmental trajectory of many genes makes it challenging to identify when gene therapy will be safe and effective. For example, variants that disrupt the complex developmental process of neural crest cells give rise to multiple, diverse peripheral cells [ 63 ] and require more critical reasoning on whether gene therapy can recover the developmental changes. Our work on LRP1-related syndrome [ 64 ] highlights the complex multi-phenotype traits associated with neural crest cells that will be difficult to advance gene therapy approaches within the developmental stages, often active in utero.

3.3. Variant Location within Proteins

As shown in Figure 6 H, many clinically sequenced variants within genes that may benefit from gene therapy fall within sites that are difficult to annotate and thus result in an annotation as a VUS. These are often subtle missense variants within a gene and are the first observance of such variants. Most of these variants have only been identified in a single individual and never observed in the millions of sequenced human genomes completed to date, making it difficult to establish a causal nature of the missense variant [ 22 ]. Thus, it has become common that gene therapies are initiated only in individuals that have either a variant that occurs in multiple individuals (often autosomal recessive conditions) or the variant results in a frameshift or nonsense change that removes large chunks of protein observed to be removed in other patients with the disorder. There is a need for characterizing VUSs rapidly using existing data [ 23 , 64 , 65 , 66 , 67 , 68 ], high-throughput wet lab techniques used in NAA10 characterizations [ 69 ], knowledge from paralog proteins such as the work on SOX transcription factors [ 70 ], or through crowd-sourcing variant lists to identify matching variant locations and phenotypes as was the case for MED13 [ 71 ].

Unique variants within genes with early and penetrant phenotypes matched to other pathogenic cases with similar phenotypes are easier to diagnose and determine a missense variant as pathogenic. This relies on phenotype matching, even if variants are unique to a patient. However, in the case of most progressive disorders (such as neurodegeneration) that are detectable in newborn screening before the phenotype is observed, these missense variants cannot be mapped with confidence, preventing the initiation of gene therapy until a phenotype appears. Therefore, if we anticipate gene therapy to apply to every individual for a gene approved with therapy, we must build more robust tools for interpreting each amino acid within an observed gene.

3.4. Gene Isoforms and Common Variants

Among the OMIM genes, each gene has an average of 6.2 protein-coding isoforms. These isoforms represent changes in splicing or transcriptional start sites that can alter the sequence of each protein. It is important to remember that many genes have different isoforms within different tissues and that human variants can result in altered splicing [ 72 ]. Previously, we showed how variants could alter gene splicing, such as small GTPases [ 73 ], and how alternative transcriptional start sites can change the interpretation of common disease association variants, such as SHROOM3 for chronic kidney disease [ 36 ].

The SMN1 and SMN2 genes each contain multiple spliced isoforms variably expressed in different human datasets based on the GTEx database [ 72 ] ( Figure 8 A). Each of these different isoforms has splice differences that remove one of three exons, resulting in various-sized proteins of each ( Figure 8 B). New genomic tools such as GTEx have built correlations between genomic variants within genomes and expression (eQTLs) or splicing (sQTLs) for each gene. Both the SMN1 and SMN2 genes have eQTLs and sQTLs that modify the genes ( Figure 8 C). More importantly, these variants are found enriched within human populations such as Africans/African Americans and remain understudied. Interestingly, both the sQTLs in SMN1 and SMN2 are found at the C-terminal region of the genes in similar locations ( Figure 8 D).

Isoforms and genetics of SMN1 and SMN2. ( A ) Top three protein-coding isoforms for SMN1 and SMN2 genes. ( B ) Exon map of isoforms within panel ( A ). ( C ) GTEx-measured eQTLs and sQTLs for the SMN1 and SMN2 genes. The significance and the population with the highest frequency of the variants are labeled in red below the violin plots. ( D ) Chromosome 5 map of the top eQTL and sQTL signals for SMN1 and SMN2.

While we highlight the role of variants of SMN1 and SMN2 , many human genes have variants that can modify expression levels or splicing [ 72 ]. However, most of these variants have remained understudied regarding how to incorporate them into gene therapy approaches. This represents a promising area for further exploration as we develop gene therapies for diverse human populations that are increasingly being studied using population-level genomics such as GTEx.

3.5. Risk of Overexpression

In gene therapy, determining and controlling the appropriate protein expression level in cells can be challenging, with uncertain outcomes if the expression is too high. Tools are available to help guide us to potential outcomes of gene overexpression, ranging from additional copies to overexpression in disease states. When determining a gene for therapy, it is critical to observe using data analysis tools if the overexpression could result in any measurable phenotypes. This can include the analysis of ClinGen [ 74 ] to determine if there are any known genetic events within humans for dosage sensitivity, specifically the genetic duplication of the gene that results in a measurable phenotype (triplosensitivity). As noted above, eQTLs can also tell us when subtle variants, often noncoding, can result in population-level increases in gene expression. These eQTL variants can be compared to Genome-Wide Association Studies (GWASs) or Phenome-Wide Association Studies (PheWASs) to find when these variants associated with elevated expression can also overlap with a measurable phenotype, taking care to determine the maximum peak overlap of colocalization of the expression and phenotype of the same variant [ 23 ].

An example of colocalized variants can be seen in the NF1 gene, which is emerging as a new potential gene therapy target for Neurofibromatosis [ 75 ]. The variant chr17_31326275_T_C (rs9894648) is found in diverse populations with significant known NF1 eQTLs over multiple tissues and a colocalized signal for the variant to traits such as sex-hormone-binding globulin protein ( Figure 9 ). This suggests that modulation of NF1 levels in gene therapy could have a resulting perturbation in hormone signaling that could be measured over gene therapy trials to determine if this has clinical utility. We must utilize our massive biological knowledgebases, such as eQTLs and GWASs/PheWASs, to determine non-biased traits that should be measured within clinical trials as a risk of overexpression of a chosen gene.

Representative analysis of a variant colocalized for expression and phenotypes. The first panel shows variant allele frequency data from gnomAD population genomics sequencing. The GTEx eQTL plots show five different tissues with significant eQTLs for the variant within the NF1 gene. The bottom plot shows the Open Target Genetics [ 76 ] data curation for significant traits associated with this variant.

3.6. Delivery Systems

A gene therapy delivery system must reach the targeted cells, evade immune system phagocytosis (depleting therapy), and make a functional protein once in the cell while avoiding lysosomal degradation [ 4 ]. Delivery strategies such as lipid-based systems and nanoparticles have little cell specificity for delivery, while viral strategies have more surface receptor specificity and higher risks of immune activation [ 5 , 77 ]. Non-viral delivery systems have seen a recent boost with use in SARS-CoV-2 and other mRNA vaccines, which has increased the hope of applying them to broader gene therapies [ 78 ]. Newer biological strategies, such as extracellular vesicles, are also emerging as ways to avoid immune activation [ 79 ]. Viral vectors such as adeno-associated viruses (AAVs) have lower immune activation and a limited 4.8 kilobase insert size. In contrast, larger viruses such as herpes simplex virus (HSV) have a larger insert capacity but higher immunogenicity with narrower cell targeting [ 80 ]. As many of these viruses are natural sources of infection, some individuals carry antibodies or T-cells that are responsive during gene therapy and must be monitored [ 81 ]. Substantial ongoing efforts are therefore aimed at reducing the immunogenicity of viral vectors and functionalizing non-viral vectors to enhance cell-type-specific targeting and effects.

Viral delivery systems are often matched to the cell/tissue type of natural infection, opening the door for engineering opportunities to enhance delivery to tissues without an optimal viral system. While there were significant investments in gene therapy approaches for cystic fibrosis, these therapies struggled to find therapeutic benefits due to difficulty in delivery to the progenitor cells of the lung. Hurdles to AAV gene transfer to airway epithelia for cystic fibrosis include (1) by-passing the mucus to reach the cell surface; (2) binding a receptor at the apical cell surface; (3) endocytosis for cell entry; (4) trafficking to the nucleus; (5) conversion of the single-stranded DNA core to double-stranded DNA followed by concatemerization and/or integration; and (6) achieving therapeutic levels of protein expression. As the current small molecule cystic fibrosis drugs are only recommended for individuals with a delta508 variant, gene therapy is still needed to treat individuals of diverse ancestry not having delta508 [ 82 ]. Over the past decade, improvement in the efficiency of AAV targeting of airway epithelia has been achieved by using different serotypes [ 83 , 84 , 85 , 86 , 87 , 88 ], site-directed mutagenesis modifications of viral capsids [ 89 ], and targeted evolution selection [ 90 , 91 ]. Currently, ongoing clinical trials using the AAV vector derived from directed evolutions demonstrate promising safety profiles for treating individuals who are ineligible for or unable to tolerate triple therapy ( {"type":"clinical-trial","attrs":{"text":"NCT05248230","term_id":"NCT05248230"}} NCT05248230 ).

The prevailing hope throughout the gene therapy field is that viral delivery systems studied within each trial will be carried forward into the subsequent development to minimize the risks of gene therapy with delivery system human validation data [ 92 ]. Multiple AAV clinical trials have pointed towards hepatic injury risks [ 93 ], including cytokine/neutrophil-dependent mechanisms [ 94 ]. In animal studies, these risks are contributed to by environmental factors such as obesity and diabetes [ 95 ]. As gene therapy progresses in clinical trials and FDA-approved clinical use, we must document risk factors for adverse outcomes to each vector and determine environmental or genetic factors to help identify risks.

4. Immune Response

Currently, gene therapy is designed to deliver the desired effect in one dose. However, there is a lack of long-term data on the efficacy of these treatments as the FDA approvals have only been in the past few years [ 96 , 97 ]. As more data are obtained about these therapies, redosing may be necessary. The possibility of redosing poses a challenge to gene therapy vectors [ 98 ]. Viral vectors have most of their replication machinery removed to enable them to carry the desired gene. However, the vector still contains surface epitopes that elicit innate and adaptive responses against the virus as the wild-type immune response [ 99 , 100 ]. Usually, producing antibodies or T-cell adaptive responses to viral infections is advantageous to help clear infection and enables future viral detection to provide resistance. However, in the case of viral vectors of gene therapy, it is a significant roadblock, as the antibodies may already be present from similar natural infections, or the first dose may inhibit the efficacy of vector reutilization for future doses of gene therapy [ 101 ].

The presence of viral vector antibodies before treatment threatens the future accessibility of gene therapy and increases the risk of adverse events. In 1999, the University of Pennsylvania conducted a clinical trial for an adenovirus serotype 5 (Ad5)-based gene therapy for a rare metabolic disease known as ornithine transcarbamylase (OTC). One of the participants suffered from lethal systemic inflammation four days post-treatment [ 102 ]. A recent study by Somanathan et al. (2020) presents data suggesting that preexisting Ad5 antibodies may have contributed to the lethal inflammatory response [ 103 ]. Additionally, recent deaths in a pediatric high-dose adeno-associated virus (AAV) gene therapy trial for X-linked myotubular myopathy may have been caused by AAV antibodies and an exaggerated immune response similar to that observed in the OTC trial [ 104 ]. As a result of the risk of exaggerated immune response, made evident by these incidents, individuals with pre-existing immunity to specific viral vectors are to be excluded from viral-based gene therapy clinical trials [ 105 ].

Levels of pre-existing antibodies for AAVs have been noted to be high enough to reduce the patient inclusion population for clinical trials by almost 50% [ 106 ]. The prevalence of these antibodies (seroprevalence) can differ across populations. Some populations have been found to have over 90% pre-existing adenovirus immunity by age 2 [ 107 ]. The high prevalence of pre-existing antibodies can biologically limit the accessibility of gene therapies to specific populations and even perpetuate current racial disparities in healthcare accessibility. A recent study by Khatri et al. (2022) found seroprevalence was higher among U.S. racial minorities, specifically Hispanic and African American individuals [ 108 ]. Therefore, gene therapies utilizing viral vectors may have decreased efficacy in racial minorities.

Zolgensma highlights the gravity of this issue. Zolgensma uses the AAV9 vector. Khatri et al. (2022) found significantly higher AAV9 seroprevalence among black donors than white donors [ 108 ]. However, in their study of the differences in SMN1 allele frequency in North America among different ethnic groups, Hendrickson et al. (2009) found black individuals to have five times the risk of being a carrier for SMA compared to white individuals [ 109 ]. The design of Zolgensma creates the potential for a lack of biological accessibility to one of the populations that could benefit the most from it.

To avoid this issue, gene therapy vectors must be chosen with their target population in mind. The vector utilized should be that which, along with being the most biologically functional and effective to deliver the gene of interest, is accessible to the broadest possible range of populations. Antibody titers can be used to measure pre-existing immunity. Two primary assays have been developed: binding assays that measure the total amount of antibodies (neutralizing and non-neutralizing) and neutralizing assays that only measure neutralizing antibodies [ 105 ]. Continued monitoring of global seroprevalence and continued prescreening of trial participants and potential gene therapy patients will be necessary to address the growing challenge of pre-existing immunity to viral vectors.

Research is needed to understand the immune response to viral vectors further. This enhanced understanding may allow for the targeted modulation of the immune response to improve vector efficacy and allow for possible redosing. Immune system modulation may involve antibody neutralization, as described in a review of recent research by Herzog and Biswas (2020) [ 110 ]. A specific strategy utilizes immunoglobulin-degrading enzymes from Streptococcus that can be administered prior to AAV treatment. The enzymes cut immunoglobulins at a specific site to make them unable to neutralize the vector. This strategy would prevent the development of an immune response, allowing for improved transduction and treatment efficacy [ 111 ].

Using viral vectors mandates the co-administration of steroids to prevent transaminitis, a broad immune modification [ 112 ]. Although initial study protocols suggested treatment for 30 days followed by a 30-day taper, most patients required steroids longer due to persistent transaminitis. Chand et al. summarized the initial studies with Onasemnogene abeparvovec (Zolgensma) for SMA and noted an average steroid usage of 83 days, ranging from 33 to 229 days [ 113 ]. In general, limited use of steroids is safe in infants and children. Steroids are frequently given to neonates with bronchopulmonary dysplasia, infants with infantile spasms, or children with nephrotic syndrome. Common short-term side effects include changes in appetite, mild immunosuppression, and gastrointestinal discomfort. Infants may exhibit changes in hunger or sleep patterns when started on steroids and often have a disrupted vaccination schedule. Stopping steroids after gene transfer becomes more difficult the longer the patient is on the steroids; careful tapering is required to avoid an adrenal crisis. Although common steroids, like prednisone and prednisolone, are relatively affordable compared to gene therapy, the potential side effects from longer-term steroid use could increase the overall cost burden, particularly if hospitalization is required.

5. Cost of Gene Therapy

While gene therapy brings significant benefits to patients, it also comes with incredible costs. Research and development have been estimated to cost between USD 318 million and USD 3 billion per gene therapy development [ 114 ]. Gene therapy for SMA consists of a one-time intravenous dose. The disease’s rarity ensures a small number of patients receive the medication. The limited usage of the drug drives up the cost. More importantly, this suggests a needed international effort to identify all patients with these rare diseases to reduce cost per patient. Zolgensma, a gene therapy for SMA, costs USD 2.1 million for a one-time dose. The approved gene therapy for hemophilia B, Hemgenix, costs USD 3.5 million per treatment, making it the most expensive drug worldwide, highlighting the need to identify more patients with disease or drug competition to reduce pricing. The high cost of these treatments can be absorbed into the payer’s system because the number of patients requiring treatment is relatively low. This may not be feasible when gene therapy is available for more diseases and a broader population of patients. A cost analysis of gene therapy versus other maintenance therapies for SMA shows that gene therapy is more cost-effective than lifelong intermittent doses of maintenance therapy [ 115 ]. This cost-effectiveness is not maintained when SMA patients suffer a relapse [ 116 ]. It is also likely to be less cost-effective in more mild diseases. As more data are obtained, cost-effectiveness may not be maintained.

With effective treatments that are more cost-effective for rare diseases, it will be imperative for payment systems to adapt and accommodate the high cost of the medications. It has been suggested that paying smaller amounts over time instead of one large payment before the administration could be an effective mechanism to share the cost between the payers and pharmaceutical companies [ 112 ]. It would also ensure the payment could be stopped if the therapy ceases to be effective, similar to stopping the medication if it is no longer effective. This model has already been used in national health plans [ 117 ]. Spain and France, for example, will only continue payments for hepatic C treatment if the patient is cured [ 114 ]. Novartis also utilizes this approach with Kymriah, a gene therapy for B-cell acute lymphoblastic leukemia. Novartis has an agreement with hospitals that they do not invoice for Kymriah until a 30-day outcome test is completed. No payment is required if the patient does not respond successfully to the treatment in this period [ 118 ]. This approach limits the financial burden on patients and hospital systems and increases the financial accessibility of these potentially curative treatments.

In the United States, the Orphan Drug Act (ODA) (1983) was developed to provide financial benefits to pharmaceutical companies for the development of drugs for rare diseases affecting fewer than 200,000 people in the U.S. Some of these benefits include market exclusivity, federal grants, and waivers of marketing application user fees [ 119 ]. However, there is a need to incentivize gene therapy development further and reduce the cost of this therapy. These reforms may include implementing a stratified benefit system in which incentives depend on the disease population size and decreasing exclusivity periods to ensure benefits are only utilized for drugs with small patient populations and limited economic potential [ 119 ].

6. Need for Increased Transparency

Gene therapy has a history of false hope and exaggerated hype. In the early 1990s, completing the first gene therapy clinical trial led to a wave of excitement perpetuated by the media. This enthusiasm spread to researchers and the public alike, leading to the initiation of numerous research projects and a push to advance gene therapy clinical trials. However, this excitement and rapid advancement proved to be self-destructive. In 1995, the NIH released a statement criticizing the field of gene therapy for rushed clinical trials, poor experimental design, and lack of rational scientific logic [ 120 ].

This pattern was seen again in 2008, with two reports in the New England Journal of Medicine describing a gene therapy to correct a form of congenital blindness. The media extrapolated the results of these reports to suggest the potential for curing eye conditions of all kinds. These statements were met with backlash from the scientific community, specifically about the pressure put on them to accelerate gene therapies [ 121 ].

These incidents illustrate how the revolutionary potential of gene therapy needs to be paired with humility. Gene therapy has the potential to do a lot of good, but there are risks and uncertainties. Improved multiway communication between all stakeholders—physicians, researchers, policymakers, companies, patients, and the public—about gene therapy’s risks and benefits is necessary. The information conveyed to patients and the general public should be clear, relatable, concise, and reliable. This information may be paired with increased genetic education through genetic counseling for patients and their families, as knowledge of genetics is crucial to understanding gene therapy’s risks and benefits [ 122 ].

The potential for side effects, the possibility that effectiveness may wane, and the plethora of new gene therapy drugs in the pipeline necessitate discussion between researchers, clinicians, and patients. This ongoing discussion will be essential to ensure side effects are noted swiftly, and changes to clinical practice can be made. Currently, rare disease advocacy groups have well-established registries collecting patient data across institutions, including groups serving multiple diagnoses like the Muscular Dystrophy Association and groups specific to one disease process like CureSMA, CureDuchenne, and Parent Project Muscular Dystrophy. These databases have years of patient information and already have the infrastructure to collect information on safety and patient outcomes as gene therapy is used and implemented in the future. These groups serve as a valuable resource for communication between patients, clinicians, and researchers.

Physicians from every specialty should know about the field to effectively communicate relevant information to their patients. Physicians and researchers should work together to ensure access to relevant information about current gene therapy developments to keep patients well informed about the current state of research. However, not all education is top-down. Researchers also need to hear from patients about their concerns and experiences to ensure research efforts align with the needs of the patient population for which they are developing treatments [ 120 ].

The high cost of gene therapy leads to a complicated pay structure. This requires clinicians, payers, and hospital systems to communicate to ensure timely patient drug delivery. Lastly, communication between policymakers, clinicians, patients, payers, and hospital systems must be prioritized to ensure safety and equitable distribution are established.

An increase in information sharing between companies and researchers, specifically about failed clinical trials, is also imperative to the informational accessibility of gene therapy. After a failed phase III clinical trial for gene therapy for epidermolysis bullosa, the company leading the study contacted other companies working on the disease and unpacked their data, presenting what they had learned from the failed study. As a result, one of the companies changed its inclusion endpoints [ 123 ]. This model of accountability and transparency is necessary for the future progression of gene therapy. The success of a gene therapy clinical trial hinges on multiple components, such as the vector selection, the gene delivered, and the promoter utilized. The accessibility of this information is essential to the analysis of both prior and present clinical trials to analyze current trends in trial design and common denominators for observed outcomes.

7. Ethical Considerations for Gene Therapy—Conclusions

The ethics of gene therapy are as multi-faceted as the field of medicine itself. We have laid out the biological, clinical, and public/patient-centric ethical considerations of gene therapy within this article ( Figure 10 ). However, the ethical issues surrounding gene therapy are less about gene therapy itself and more about the medical, cultural, social, and political contexts in which it emerged. We cannot boil down these questions and issues to one-time decisions and solutions, which would disregard the relational and longitudinal nature of ethics [ 124 ]. Addison and Lassen unravel the concept of the ethics of gene therapy clinical trials as follows: “The ethical complexities of gene therapy are not confined to the consent process or the procedure, nor does the ethics review process resolve them. Rather, the treatment unfurls a multitude of ethical dilemmas, which manifest both in discrete moments of choice and the on-going endeavor of how to live well or care well in the aftermath of the event itself”.

Summary of the ethical considerations of gene therapy. This figure was generated with BioRender.

The Hippocratic Oath [ 125 ], often referred to as the basis of ethical medical practice, presents the purpose of medicine as “to do away with suffering of the sick, to lessen the violence of their diseases.” The purpose of medicine and the principle of ethical practice hinge on relieving patient suffering, which, at its core, is patient-centered [ 126 ]. Therefore, the ethical advancement of gene therapy hinges on developing patient-centered solutions to the present and emerging ethical dilemmas and issues faced within this field. With every decision and every advancement, we must remember the patient.

This patient-centered lens can serve as the basis for thinking about the ethics of many gene therapy topics we have discussed. As evident in our analysis of gene therapy clinical trials, gene therapy is still in its early stages of development, with most clinical trials falling into the early phase categories (phases I, I/II, and II). The lack of international partnerships has prevented the scale of gene therapy from matching the rarity of diseases it is being developed to treat, representing a significant ethical consideration for cross-border study designs [ 19 ].

Severe adverse events, even patient deaths, although they are to be actively avoided through proper monitoring and reporting, are not uncommon within early phase trials, especially phase I trials [ 127 ]. In 1999, 153,964 severe adverse events (17,399 of them patient deaths) were reported to the Center for Drug Evaluation and Research of the United States FDA [ 128 ]. That same year, the phase I gene therapy clinical trial for OTC deficiency resulting in death was highly publicized, with 22 New York Times articles [ 129 ]. The media focusses on gene therapy more than other disciplines, leading to an amplified perception of risk. We must be clear about who these risks fall upon. Ultimately, they fall upon the patients—those actively involved in trials, those who will receive these treatments in the future, and those directly and indirectly affected by the outcomes of these discussions and decisions. Therefore, we must actively involve patient populations in the discussions and decision-making processes about the acceptable level of risk.

One option discussed by Pattee in their commentary titled “Protections for Participants in Gene Therapy Trials: A Patient’s Perspective” [ 130 ] is to consult patients who have participated in trials on trial design, development, and direction, such as ensuring the adequacy of informed consent materials and trial logistics. Doing so would increase trial transparency and public trust in gene therapy, even amid complex uncertainties within the field. Pattee also suggests further protecting patients participating in clinical trials through improved public education about clinical trials to clarify information and concerns presented in the media and including disease-specific experts within centralized IRBs to incorporate additional perspectives specific to the patient population during trial design and monitoring [ 130 ].

Accessibility is a crucial factor to be considered in the ethical advancement of gene therapy. Rare diseases affect a small number of individuals in distinct ways. No two patients are identical. Gene therapy reflects the patient population in this way—it is designed to be specialized. The needs are not equal; therefore, treatments cannot be equal. However, treatment equity is still needed, from costs to the type of rare disease to trial access that disproportionately benefits a few [ 118 , 131 ]. To think about equity and accessibility is to consider already present disparities in healthcare systems, patterns we see emerging from early research and clinical trials, and other potential barriers that could threaten the ethical advancement of gene therapy, the safety of patient populations, and the ability of patients to access these potentially curative treatments.

Over 50% of individuals with rare diseases report using their savings to cover medical costs, with one in ten filing for bankruptcy [ 123 ]. The high cost of gene therapies is thus likely to continue overwhelming patients with rare diseases and the funding agencies for medical care, thus limiting personal access. As shown in Table 1 , Table 2 and Table 3 , only a few rare diseases have authorized gene therapies, where the >5000 unique rare diseases represent a significant opportunity to reduce production costs through transparent design that enables the subsequent therapy to be developed at a lower cost. Further expansion of international collaborations will unite rare disease patients to present a more extensive base of therapies. No matter how effective or miraculous, a treatment inaccessible to patients has no real value. Thus, a balance of patient risk, education, and accessibility remains the ethical priority for gene therapy of rare diseases.

Abbreviations

FDA, Food and Drug Administration; SMA, Spinal Muscular Atrophy; DNA, deoxyribonucleic acid; RNA, ribonucleic acid; IRDiRC, International Rare Diseases Research Consortium; CRISPR, clustered regularly interspaced short palindromic repeats; Cas9, CRISPR-associated protein 9; TP53, tumor protein p53; siRNA, small interfering RNA; mRNA, messenger RNA; NIH, National Institutes of Health; NCI, National Cancer Institute; NIAID, National Institute of Allergy and Infectious Diseases; NHLBI, National Health Lung and Blood Institute; NIA, National Health Lung and Blood Institute; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2; ARPA-H, Advanced Research Projects Agency for Health; AAV, adeno-associated virus; OMIM, Online Mendelian Inheritance in Man; HPA, Human Protein Atlas; IMPC, International Mouse Phenotyping Consortium; VUS, variant of uncertain significance; eQTL, expression quantitative trait loci; sQTL, splicing quantitative trait loci; GWAS, Genome-Wide Association Study; HSV, herpes simplex virus; Ad5, adenovirus serotype 5; OTC, ornithine transcarbamylase; ODA, Orphan Drug Act.

Funding Statement

This research was partially supported by Helen DeVos Children’s Hospital (J.M.K.), Michigan State University (X.L., D.B.C., M.R.W., D.L.V. and J.W.P.), and the Gerber Foundation (C.P.B., S.R., and J.W.P.). M.L.H. was an undergraduate fellow supported by the Gerber award.

Key Contribution

The promise of gene therapy is reflected through the FDA approvals for multiple genomic syndromes. This work reflects on the field’s current state while providing topics that must be considered as the field progresses with more clinical usages.

Author Contributions

Conceptualization, M.L.H., J.W.P. and J.M.K.; methodology, M.L.H., J.K.Z. and J.W.P.; formal analysis, M.L.H., J.K.Z., J.W.P. and J.M.K.; writing—original draft preparation, M.L.H., J.K.Z., X.L, D.B.C., M.R.W., D.L.V., C.P.B., Y.M.E., S.R., N.L.H., J.W.P. and J.M.K.; writing—review and editing, M.L.H., J.K.Z., X.L., D.B.C., M.R.W., C.P.B., Y.M.E., S.R., N.L.H., J.W.P. and J.M.K.; supervision, J.W.P. and J.M.K.; project administration, J.W.P. and J.M.K. All authors have read and agreed to the published version of the manuscript.

Institutional Review Board Statement

Informed consent statement, data availability statement, conflicts of interest.

The authors declare no conflicts of interest.

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Gene therapy relieves back pain, repairs damaged disc in mice

Study suggests nanocarriers loaded with dna could replace opioids.

Disc-related back pain may one day meet its therapeutic match: gene therapy delivered by naturally derived nanocarriers that, a new study shows, repairs damaged discs in the spine and lowers pain symptoms in mice. 

Scientists engineered nanocarriers using mouse connective-tissue cells called fibroblasts as a model of skin cells and loaded them with genetic material for a protein key to tissue development. The team injected a solution containing the carriers into damaged discs in mice at the same time the back injury occurred. 

Assessing outcomes over 12 weeks, researchers found through imaging, tissue analysis, and mechanical and behavioral tests that the gene therapy restored structural integrity and function to degenerated discs and reduced signs of back pain in the animals. 

“We have this unique strategy that’s able to both regenerate tissue and inhibit some symptoms of pain,” said co-senior author Devina Purmessur Walter , associate professor of biomedical engineering at The Ohio State University. 

Though there is more to learn, the findings suggest gene therapy could offer an effective and long-lasting alternative to opioids for the management of debilitating back pain. 

“This can be used at the same time as surgery to actually boost healing of the disc itself,” said co-senior author Natalia Higuita-Castro , associate professor of biomedical engineering and neurological surgery at Ohio State. “Your own cells are actually doing the work and going back to a healthy state.” 

The study was published online recently in the journal Biomaterials . 

An estimated 40% of low-back pain cases are attributed to degeneration of the cushiony intervertebral discs that absorb shocks and provide flexibility to the spine, previous research suggests. And while trimming away bulging tissue from a herniated disc during surgery typically reduces pain, it does not repair the disc itself – which continues to degenerate with the passage of time. 

“Once you take a piece away, the tissue decompresses like a flat tire,” Purmessur Walter said. “The disease process continues, and impacts the other discs on either side because you’re losing that pressure that is critical for spinal function. Clinicians don’t have a good way of addressing that.” 

This new study builds upon previous work in Higuita-Castro’s lab, which reported a year ago that nanocarriers called extracellular vesicles loaded with anti-inflammatory cargo curbed tissue injury in damaged mouse lungs . The engineered carriers are replicas of the natural extracellular vesicles that circulate in humans’ bloodstream and biological fluids, carrying messages between cells. 

To create the vesicles, scientists apply an electrical charge to a donor cell to transiently open holes in its membrane, and deliver externally obtained DNA inside that converts to a specific protein, as well as molecules that prompt the manufacture of even more of a functional protein. 

In this study, the cargo consisted of material to produce a “pioneer” transcription factor protein called FOXF1 , which is important in the development and growth of tissues. 

“Our concept is recapitulating development: FOXF1 is expressed during development and in healthy tissue, but it decreases with age,” Purmessur Walter said. “We’re basically trying to trick the cells and give them a boost back to their developmental state when they’re growing and at their healthiest.” 

In experiments, mice with injured discs treated with FOXF1 nanocarriers were compared to injured mice given saline or mock nanocarriers and uninjured mice. 

Compared to controls, the discs in mice receiving gene therapy showed a host of improvements: The tissue plumped back up and became more stable through production of a protein that holds water and other matrix proteins, all helping promote range of motion, load bearing and flexibility in the spine. Behavioral tests showed the therapy decreased symptoms of pain in mice, though these responses differed by sex – males and females showed varying levels of susceptibility to pain based on the types of movement being assessed. 

The findings speak to the value of using universal adult donor cells to create these extracellular vesicle therapies, the researchers said, because they don’t carry the risk of generating an immune response. The gene therapy also, ideally, would function as a one-time treatment – a therapeutic gift that keeps on giving. 

“The idea of cell reprogramming is that you express this transcription factor and the cell is then going to convert to this healthier state and stays committed to that healthier phenotype – and that conversion is not normally transient,” Higuita-Castro said. “So in theory, you would not expect to have to re-dose significantly.” 

There are more experiments to come, testing the effects of other transcription factors that contribute to intervertebral disc development. And because this first study used young adult mice, the team also plans to test the therapy’s effects in older animals that model age-related degeneration and, eventually, in clinical trials for larger animals known to develop back problems. 

Higuita-Castro, director of advanced therapeutics and engineering in the College of Medicine  Davis Heart and Lung Research Institute and a core faculty member of Ohio State’s  Gene Therapy Institute , and Purmessur Walter, an investigator in Ohio State’s Spine Research Institute and director of the Spinal Therapeutics Laboratory in the College of Engineering, are co-principal investigators on National Institutes of Health grants funding this research. 

Additional co-authors include co-first authors Shirley Tang and Ana Salazar-Puerta, Mary Heimann, Kyle Kuchynsky, María Rincon-Benavides, Mia Kordowski, Gilian Gunsch, Lucy Bodine, Khady Diop, Connor Gantt, Safdar Khan, Anna Bratasz, Olga Kokiko-Cochran, Julie Fitzgerald and Benjamin Walter, all of Ohio State; Damien Laudier of Icahn School of Medicine at Mount Sinai; and Judith Hoyland of the University of Manchester. 

Ohio State has filed a patent application on nonviral gene therapy for minimally invasively treating painful musculoskeletal disorders.

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