Patients’ Cells Offer Insight into New Gene Therapy Approach to Rare Eye Disease

Daniel S. Levine

At the end of 2017, the U.S. Food and Drug Administration approved Luxturna, the first directly administered gene therapy approved in the United States that targets a disease caused by mutations in a specific gene.

Luxturna treats a rare form of inherited vision loss known as Leber congenital amaurosis (LCA). It is approved to treat patients with RPE65 mutations that lead to vision loss and may cause complete blindness in certain patients. The RPE65 gene provides instructions for making an enzyme that is essential for normal vision.

While Luxturna represented a significant breakthrough, it works on only a small subset of patients—about 1,000 to 2,000 patients in the United States—with LCA.

LCA, though, can have many underlying genetic causes. In fact, there are at least 25 genes, many of which are critical for the development or function of photoreceptors, that have been identified as capable of causing LCA. Most patients with LCA have autosomal recessive forms of the disease. In such cases, mouse models of the disease have served as useful tools for understanding how the disease arises and the development of therapies. But existing animal models have limited utility. Today, no treatments are available for other forms of LCA, particularly dominant forms of the condition.

Now, researchers at the National Institutes of Health’s National Eye Institute have developed a gene therapy strategy they say has the potential to treat a dominant form of LCA and other retinopathies cause by mutations of the CRX gene. The CRX gene encodes the CRX protein, which signals photoreceptor cells in the retina—rods and cones—to produce light-sensitive pigments known as opsins. In the absence of functional CRX protein, photoreceptors lose their ability to detect light and eventually die.

The researchers created an in vitro model of CRX-LCA by creating retinal organoids—derived from two patients with different forms of CRX-LCA—to create living models of the disease for proof-of-concept, a strategy they say “should be applicable in developing effective treatments for rare, and even dominant, inherited diseases of the retina and other parts of the central nervous system.” Their findings appeared at the end of January in the journal Stem Cell Reports.

Autosomal-dominant LCA, like other dominant disorders, can be challenging to treat with gene therapies because providing normal copies of the gene often fails to restore function. That’s because people with these types of conditions have one normal copy of the gene, but its function can be impaired by the mutant version of the protein, which interferes with the normal protein. Adding more of the protein can enhance the disease in unpredictable ways.

“Gene therapy has been used generally to correct loss of function disease. That means if a gene is inactive, you put the gene back, and it can reverse the phenotype, which is what happened with Luxturna, or RPE 65 gene therapy,” said Anand Swaroop, chief of the NEI Neurobiology, Neurodegeneration and Repair Laboratory and senior author of the study. “In a dominant disease, what happens is that you generally have a gain of function. The normal allele is not able to give you the correct function because the bad, or the dominant allele, either suppresses that function by squelching that particular protein, or it creates a new function that suppresses the function of the normal allele so that the normal gene cannot do its job.”

To treat the CRX-LCA, the researchers used a gene augmentation strategy. Rather than replace the faulty gene as is done with traditional gene therapies, they sought to counteract the effects of the dominant allele by adding additional copies of the functional gene.

“We are not replacing that. We are bombarding it with a lot of normal gene,” said Swaroop, “Not too much, but enough that it can suppress whatever bad effects of dominant gene were there and give some normal activity for the patient to see.”

What enabled the researchers to pursue their strategy was the ability to take skin cells from patients with the condition to make induced pluripotent stem cells and create retinal organoids—three-dimensional, living tissue models of the disease.

“It’s a viable and useful approach for evaluating different treatment paradigms. It doesn’t have to be just gene therapy, but it can be small molecules or an antisense oligonucleotide approach, or whatever you want to try,” said Swaroop, who noted with mouse models of disease researchers are often limited to a single model. “They are human systems, not mouse. You can test many of them. In the near future, it should be very easy to test these treatments for multiple patients.”

Because it was critical to control how much CRX gene would be expressed by the recipient photoreceptors, the researchers packaged a CRX promoter—a neighboring DNA sequence that controls how and when the gene is expressed—within the vector as part of the therapy. 

The work showed that the augmentation strategy restored some CRX protein function in the organoids grown from the patients’ cells and successfully drove expression of opsins in photoreceptors.

While Swaroop is encouraged by the proof-of-concept of the gene therapy strategy, he’s hoping a foundation or biotech company will license the technology to carry the gene therapy forward through further translational studies and clinical development.

“We were able to take care of a dominant disease. From a scientific perspective that was quite exciting. We were not sure whether we would be successful without removing the dominant allele,” said Swaroop. “These kids go blind very early in their life or they lose vision. We hope somebody will take over and do some trials and get it to the children.”

Image: Fluorescently labeled organoid showing columnal cells in outer ring. Anand Swaroop, NEI

Thanks to Pfizer, Inc., Bluebird, and Novartis Gene Therapies for their support of this article, part of our Platforms of Hope: Advances in Gene Therapy and Gene Editing series.