Researchers Test Gene Augmentation Therapy in Patient-Derived Retinal Organoids to Find Eye Disease Cure

Cone-rod homeobox (CRX) is a transcription factor gene essential for the differentiation of photoreceptor cells and is therefore crucial for the early development of the eye. Mutations in the CRX gene lead to Leber congenital amaurosis (LCA), a rare disease that primarily affects the retina.

Leber congenital amaurosis (LCA) is an umbrella term for a group of early-onset retinal dystrophies resulting in vision loss during infancy. Mutations in around 25 genes have been found to cause LCA, and a vast majority of them are essential for photoreceptor development and function. The CRX protein regulates the expression of genes involved in the production of opsins (light-sensitive pigments) by the retina’s photoreceptors. Defective CRX protein leads to photoreceptor cell death and blindness.

A team of researchers from the National Eye Institute has now developed a promising gene therapy for this condition. By establishing an in vitro model of CRX-LCA in retinal organoids using patient-derived induced pluripotent stem cells (iPSCs), they investigated the effects of an Adeno-associated virus (AAV)-mediated CRX gene augmentation therapy. Results show that the therapy partially restored photoreceptor phenotype and expression of phototransduction-related genes.

“Our treatment approach, which adds more copies of the normal gene, could potentially treat autosomal-dominant LCA caused by a variety of mutations,” said Anand Swaroop, Ph.D., Chief of the NEI Neurobiology, Neurodegeneration and Repair Laboratory and senior author of the study.

Research Strategy and Findings

Although animal models of CRX retinopathies are available, the developmental process varies, and the molecular profiles of the human retina cannot be replicated completely. Therefore, the team constructed an in vitro model of the disease in patient-specific genetic background.

First, iPSCs were derived from LCA patients with two different CRX mutations and were differentiated into 3D retinal organoids. The in vitro model of CRX-LCA in retinal organoids exhibited the disease phenotype by showing defective photoreceptor maturation and diminished expression of visual opsins. Next, the correct CRX transgene driven by a human CRX promoter was delivered using an AAV vector. The AAV-mediated CRX gene augmentation therapy partially restored photoreceptor phenotype and both rod and cone gene expression.

“The fact that this strategy worked for both CRX mutations was pretty exciting,” said Swaroop. “Gene augmentation may be a viable therapy for LCA caused by other autosomal-dominant mutations.”

“This proof-of-concept gene therapy study is the first step toward a potential treatment for a rare form of LCA,” said Brian Brooks, M.D., NEI clinical director and co-author on the study. “It’s a great example of bench-to-bedside science when researchers in basic and clinical science collaborate.”

Since gene augmentation increases the presence of the correct gene in the target individual, it can be used to deal with both a shortage of the desired protein and a relative overabundance of the mutant protein. Because of this approach, three of the following questions can be answered.

1. Competition of mutant protein with normal CRX for target binding sites

2. Prolonged occupancy of mutant protein at binding sites preventing the correct form from associating with the target DNA

3. Disruption of stoichiometric interactions with other retinal transcription factors, such as NRL

The experimental strategy used in this research could be applicable in developing treatments for rare and dominant inherited diseases of the retina and other parts of the central nervous system. Besides, the ability to modulate protein production allows disease treatments to be customized for patient needs.

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References

1. Gene therapy of dominant CRX-Leber congenital amaurosis using patient stem cell-derived retinal organoids: Stem Cell Reports
2. https://www.nih.gov/news-events/news-releases/researchers-use-patients-cells-test-gene-therapy-rare-eye-disease

Author

Eduardo Longoria