Is gene therapy the answer?

Gene therapy uses genetic material, either DNA or RNA, to treat or prevent the progression of a disease. It often involves the introduction of genetic material into a person’s cells to replace a defective or missing gene.

Although early attempts at gene therapy have been effective in achieving the expression of the therapeutic gene in the target tissue, they have also been accompanied by severe adverse effects.

These side effects included a hyperactive immune response, undesired insertion of the genes in the human genome leading to the activation of cancer-related genes, and even death.

Improvements in delivery systems for the administration of therapeutic genes to the target tissue have led to significant progress in the development of gene therapies, including those for retinal diseases.

The easy accessibility of the retina makes it an ideal target for gene therapy. Moreover, the ocular barriers separate the eyes from the rest of the body, thus limiting the immune response in the eye.

The ocular barriers also prevent the vector from spreading throughout the body and restrict the expression of the therapeutic gene to the eyes.

Gene therapy can help treat inherited retinal diseases that were previously considered to be untreatable. In addition to these retinal diseases caused solely by genetic mutations, gene therapy approaches have shown promise in the treatment of acquired retinal diseases, such as glaucoma, that have a complex origin.

Speaking to Medical News Today, Aaron Brock Roller, MD, a vitreoretinal surgeon at Austin Retina Associates in Texas, pointed out that:

“Retinal diseases are among the most ideal candidates for trials and testing of these new therapies, for numerous reasons. It seems that the development of ophthalmic genetic therapy is quickly becoming the gateway and standard model for the application of these techniques to all areas of medicine.”

The development of gene therapy approaches for retinal diseases requires the consideration of numerous factors associated with the type of delivery system and the route of administration.

The factors influencing these decisions include the size of the therapeutic gene, the target retinal cell type, the immune response elicited by the delivery vehicle, and cost-effectiveness.

Gene therapies require the presence of viable target cells for their success, and their efficacy starts to decline with the progressive degeneration of the tissue.

Gene therapy for retinal diseases involves the administration of the genetic material to the retina using a vehicle or vector. Vectors are genetically modified agents used to deliver genetic material, such as a functional copy of the mutated gene, to the cell.

While viruses are the most commonly used vectors for retinal diseases, nonviral gene transfer methods are also being investigated.

Genetically modified viruses, including adenoviruses, lentivirus, and adeno-associated viruses, have been used as vectors for retinal diseases.

These viral vectors can be genetically modified to selectively express the therapeutic gene in the target tissues or reduce the immune response to the virus.

Adenoviruses were previously used as vectors, but these viruses elicit a strong immune response, resulting in their clearance. This had led to the use of adeno-associated viruses instead of adenoviruses.

Adeno-associated viruses are small, nonpathogenic viruses that carry a low risk of integration into the human genome. As a result, these viral vectors have a low likelihood of disrupting the expression of important genes in the body, including those associated with cancer.

Since retinal cells do not divide, adeno-associated viruses can stably express the gene of interest for a long duration without integrating into the genome. In addition, adeno-associated viruses produce a mild immune response, ensuring stable gene expression.

A major drawback of using adeno-associated virus vectors is their ability to carry a limited amount of genetic material. One of the approaches to circumvent this limitation is to use dual adeno-associated viral vectors that carry the genetic material divided into two fragments.

Another option is the use of lentiviruses that can carry more genetic material than adeno-associated viruses. Lentiviral vectors can integrate into the host genome and, thus, ensure durable gene expression.

However, integration of lentiviruses in the genome can pose safety risks. Lentiviruses are also not very effective in delivering genetic material to photoreceptors, which are most frequently affected by inherited retinal diseases.

Nonviral delivery systems for gene therapy can allow the administration of larger DNA molecules and are more amenable to cost-effective, large-scale manufacturing than viral vectors.

Different types of nanoparticles and liposomes, which are vesicles made of lipids, are some promising nonviral delivery systems for gene therapy.

Genetic material can also be administered directly by transiently making the cell membrane permeable using ultrasound or electric pulses.

A major drawback of nonviral delivery systems is that the expression of the introduced genetic material is often short-lived due to its degradation by enzymes.

Subretinal injection and intravitreal injection are the predominant modes of administering viral vectors to the retina.

Subretinal injections deliver the vector to photoreceptors and retinal pigment epithelium, two common targets for treating retinal diseases. The retina is composed of layers of cells, with the retinal pigment epithelium forming the outermost layer.

The retinal pigment epithelium lies underneath the layer of photoreceptor cells and is involved in supporting the latter. The photoreceptors receive light and convert it into electrical signals that are transmitted to the brain via the optic nerve.

The subretinal mode of injection involves the injection of the vector in the subretinal space between the layer of photoreceptors and the retinal pigment epithelium. Delivery of vectors via the subretinal route is less likely to produce an inflammatory response because the space is anatomically closed and protected from immune responses.

Intravitreal injection involves the delivery of the vector to the vitreous cavity, which is the large gel-filled space between the lens and the surface of the retina.

Larger concentrations of the vector are required to produce the desired effect due to the diffusion of the vector in the vitreous body. Intravitreal injections are used to deliver vectors to the retinal ganglion cells that receive input from the photoreceptors.

Suprachoroidal injections are a newer approach that involves the administration of the vector in the space between the choroid and sclera, the two layers that lie under the retina. The choroid is the layer of connective tissue and blood vessels between the retina and the sclera, whereas the sclera is the protective outermost layer of the eye.

Suprachoroidal injections can deliver the vector to the retina and the retinal pigment epithelium and are much easier to perform than subretinal injections.

Although evidence from human studies suggests that suprachoroidal injections are safe, this method of vector delivery carries a higher risk of the vector entering the circulation, resulting in undesired gene expression in other tissues.

Inherited retinal diseases are a group of rare eye diseases caused by genetic mutations. Mutations in more than 300 genes characterized so far are known to play a causal role in inherited retinal diseases.

Gene therapy approaches offer hope for the treatment of these diseases that doctors previously held to be untreatable.

Animal models have allowed researchers to understand the mechanistic role of the genes involved in inherited retinal diseases, subsequently leading to the development of gene therapies for these conditions.

The development of gene therapies for inherited retinal diseases has also been facilitated by improvements in the design of vectors.

Inherited retinal diseases can be caused by either recessive or dominant genes. In the case of recessive genes, mutations in both copies of the genes are necessary to cause the disease.

Gene therapy for diseases caused by a recessive gene involves the introduction of a functional copy of the mutated gene.

Unlike diseases caused by recessive mutations, mutations in a single copy of the dominant gene are sufficient to produce the disease.

Disease caused by a dominant gene requires the silencing of the mutated gene to suppress its expression or correction of the mutation using CRISPR gene editing.

LCA actually refers to a group of diseases that cause severe visual impairment in infancy or early childhood. It is caused by a mutation in one of the over 20 genes involved in the function or development of retinal cells.

Early efforts at the treatment of retinal diseases with gene therapy were directed at a form of LCA caused by the RPE65 gene mutation. The RPE65 gene encodes an enzyme in the retinal pigment epithelium that facilitates the conversion of light into electrical signals in photoreceptors.

These studies included a phase III clinical trial showing that introducing a functional RPE65 gene using an adeno-associated virus led to sustained improvements in visual function at four years in RPE65-associated LCA patients.

Around 200 clinical trials are currently underway to examine the safety and effectiveness of gene therapy for other inherited retinal diseases.

Notable recent developments include two small phase 1/2 clinical trials — published in The Lancet and Nature Communications, respectively — reporting improvements in visual function in individuals with inherited retinal diseases caused by recessive mutations in the GUCY2D and RLBP1 genes.

The GUCY2D gene encodes the retinal guanylyl cyclase 1 (RETGC-1) protein, which is involved in mediating the ability of photoreceptors to adjust to a dark environment following exposure to bright light.

Specifically, the rods, a type of specialised photoreceptor cell, are affected by the GUCY2D mutation, resulting in impaired vision in the dark.

The GUCY2D phase 1/2 study showed that the subretinal delivery of a functional copy of GUCY2D using an adeno-associated viral vector resulted in improvements in visual function, including a 100-fold improvement in visual sensitivity in a low-light environment.

Significant improvements were visible as early as 4 weeks after the treatment and persisted until at least a year.

The RLBP1 gene encodes a protein that helps the eyes to adapt to changes in light intensities, and mutations in both copies of RLBP1 lead to night blindness and a slower adjustment of eyesight to darkness after exposure to bright light.

The phase 1/2 trial involving individuals with RLBP1-associated retinal dystrophy showed that subretinal administration of a functional RLBP1 copy using an adeno-associated viral vector resulted in faster adaptation to darkness and patient-reported improvements in the ability to perform daily activities.

Adverse effects reported in clinical trials for inherited retinal diseases are typically associated with the surgical procedure. The gene therapy may be associated with dose-dependent inflammation, which can be treated with corticosteroids.

These promising phase 1/2 studies for GUCY2D- and RLBP1-associated retinal dystrophy pave the way for larger phase 3 studies.

Unlike inherited retinal diseases that have a low prevalence, acquired retinal diseases affect a large portion of the population. For instance, an estimated 200 million individuals have age-related macular degeneration across the globe.

Treatment of acquired retinal diseases often involves targeting genes in pathways common to and disrupted in several diseases. This contrasts with gene therapy for inherited retinal diseases, which generally targets disease-specific mutant genes.

In other words, the same gene therapy could be used to target a shared dysfunctional pathway observed in multiple conditions. For instance, both wet age-related macular degeneration and diabetic retinopathy-related complications involve the abnormal growth of blood vessels in the retina.

The vascular endothelial growth factor (VEGF) protein is involved in mediating the excessive growth of blood vessels, and anti-VEGF injections that target this protein are effective in the management of age-related macular degeneration and diabetic retinopathy.

However, these anti-VEGF injections need to be administered frequently by a trained healthcare professional and are costly. Gene therapy approaches have the potential to overcome some of these issues by providing more durable suppression of the VEGF protein.

Several different gene therapy approaches that suppress genes in the VEGF pathway are currently being investigated in preclinical and clinical studies.

These efforts include using an adeno-associated virus carrying a gene encoding a fragment of an anti-VEGF antibody, gene silencing, and gene editing using the CRISPR system.

Since treatments already exist for acquired retinal diseases, gene therapy approaches need to show superior benefits to these treatments to gain regulatory approval.

Several studies have examined the potential of gene therapy for glaucoma, which is the leading cause of irreversible blindness in individuals over 60 years old.

Current treatments for glaucoma involve daily eye drops or surgical approaches that can potentially fail over time. Gene therapy approaches for glaucoma could provide a durable alternative, potentially requiring only a single intervention.

Sophia Millington-Ward, PhD, a research fellow at Trinity College Dublin in Ireland, explained in a press release that glaucoma “is a multifactorial condition with many different risk factors, which adds to the complexity of treating it.”

”Current glaucoma treatments focus on the use of topical eye drops, surgery, or laser therapy. However, the outcomes are variable, with some patients not responding and/or suffering serious side effects,” she pointed out.

Glaucoma is characterized by visual impairment due to the loss of retinal ganglion cells whose nerve fibers form the optic nerve. Retinal ganglion cells lie on the inner surface of the retina and are a part of the network of cells that receive electrical signals from photoreceptor cells.

Excessive pressure exerted due to the accumulation of aqueous fluid, known as intraocular pressure, is a major risk factor for glaucoma. Intraocular pressure is thought to compress the nerve fibers of the retinal ganglion cells, disrupting the transport of various proteins within the retinal ganglion cells and, subsequently, causing their death.

Two major gene therapy approaches are being actively investigated for the treatment of glaucoma. One of the approaches is aimed at reducing the abnormal pressure in the eye.

For instance, the disruption of the aquaporin 1 gene, a gene involved in the production of aqueous fluid, in a mouse model of glaucoma using the CRISPR-Cas9 platform was effective in lowering intraocular pressure and mitigating the loss of retinal ganglion cells.

The other approach involves modulating the expression of genes underlying the loss of retinal ganglion cells. Some of the factors associated with the loss of retinal ganglion cells include the activation of pathways involved in cell death, disruption of energy metabolism, decreased levels of neurotrophic factors such as brain-derived neurotrophic factor, and oxidative stress.

Several studies in rodent models of glaucoma using gene therapies targeting the aforementioned risk factors have been effective in preventing the loss of retinal ganglion cells while preserving their function.

For instance, a recent study — published in the International Journal of Molecular Sciences — which was conducted using a mouse model of glaucoma showed that the delivery of a gene involved in energy metabolism prevented the loss of retinal ganglion cells and improved visual function.

Retinal ganglion cells derived from individuals with glaucoma show an increase in oxidative stress and dysfunction of mitochondria, the cell organelles involved in energy production.

In this study, the researchers examined the therapeutic potential of the NDI1 gene that encodes an NADH dehydrogenase, a protein that forms part of a larger enzyme complex involved in energy generation in the mitochondria.

Delivery of the NDI1 gene to retinal cells obtained from humans with glaucoma resulted in lower mitochondrial dysfunction. Furthermore, the administration of the NDI1 gene to the retina of a mouse model of glaucoma prevented the loss of retinal ganglion cells and improved visual function.

“Developing broadly applicable gene therapies for large numbers of patients is particularly important, given high development costs associated with each therapy – and here we have highlighted this therapy has real potential for boosting mitochondrial function in glaucoma.”

Despite the progress in the development of gene therapies for retinal diseases, there are concerns about their efficacy and safety.

Although the durable expression of the therapeutic gene over several years is advantageous, gene therapy could lead to undesired expression of genes after the cessation of disease symptoms and produce adverse effects. For instance, prolonged suppression of VEGF in individuals with age-related macular degeneration may cause adverse effects.

Another major challenge is the large-scale manufacture of vectors for gene therapy. There are concerns about the low yield of vectors, the presence of contaminants that can elicit an immune response, and maintaining purity, efficacy, and safety across batches.

Once approved, limited accessibility and high costs could also hinder the potential application of gene therapy approaches for retinal diseases. Currently, gene therapy for retinal diseases affecting photoreceptors and retinal pigment epithelium generally requires administration of the vector using subretinal injections.

Unlike intravitreal injections that can be conducted in the office, subretinal injections need to be administered in a surgical setting. Subretinal injections are also associated with a higher risk of surgical complications.

In addition, gene therapy approaches for the treatment of inherited retinal diseases require the accurate identification of the genetic mutation underlying the condition.

Mutations in numerous genes can result in identical disease-associated changes in the retina and symptoms, making a genetic diagnosis necessary. Obtaining a genetic diagnosis may not be accessible or affordable in developing countries.

Genes underlying several inherited retinal diseases are yet to be identified and characterized. Developing gene therapies targeting each specific mutation underlying rare inherited retinal diseases is unlikely to be cost-effective or feasible.

A potential solution for the treatment of rare inherited retinal diseases is to develop gene therapies that target shared pathways in a manner similar to gene therapies for acquired retinal diseases. Such therapies, referred to as modifier gene therapies, can rectify pathways upstream and downstream of the administered gene, ideally leading to the resolution of disease symptoms.

Roller also told MNT that “medication cost is an area that may be a challenge in the immediate future given the tremendous costs of drug development and clinical trial financing, but the accessibility of these treatments is sure to improve rapidly as clinical efficacy is proven and production methods become standardized.”

Another issue is the necessity of viable photoreceptor cells for gene therapy to be effective. As a result, gene therapy for both acquired and inherited retinal diseases in the advanced stages is often less effective in restoring visual function.

This is also a reason for the lack of new therapies for inherited retinal disease (IRDs) since the approval of Luxturna. Artur Cideciyan, PhD, a professor at the University of Pennsylvania, told MNT that:

“The common denominator among the great majority of all [inherited retinal diseases] is that they all cause vision loss due to a problem with photoreceptor cells of the retina. However, there can be many differences in details. The IRD that Luxturna was approved for was RPE65-associated Leber congenital amaurosis, which showed evidence that photoreceptor cells were surviving but not functioning.”

“Luxturna provided the missing enzyme, and like a switch, photoreceptors got turned on and started functioning within days. Nearly all other gene therapies evaluated to date were in other genetic forms of IRDs where vision loss is due to degeneration of photoreceptors. Thus, treatments need to show over the long term that the slow progression of the photoreceptors can be arrested. And that has been more challenging,” he further detailed.

To circumvent this issue of degeneration of photoreceptors, researchers have been harnessing optogenetics, a method involving the administration of a gene called opsin into the other remaining cells that receive signals from photoreceptors. The integration of opsins into these second or third-order cells that communicate with photoreceptors can make them light-sensitive.

With the large number of ongoing clinical trials, more gene therapies for retinal diseases are expected to be approved in the near future. Cideciyan noted that “additional gene therapies for other rare causes of vision loss are likely to be approved for the public over the next 5 years.”