Gene therapy for rare blindness
Researchers from the National Eye Institute (US National Institutes of Health, NIH) have developed a gene therapy that eliminates cilia defects in retinal cells in Leber congenital amaurosis, a disease that causes vision loss in early childhood. Using retinal organoids grown from patients' cells, the researchers found that a type of congenital amaurosis caused by a mutation of the NPHP5 gene leads to serious defects in the primary cilia.
Mild deficiency of the NPHP5 gene causes blindness, and with a more pronounced deficiency, many patients also have kidney diseases (Senor-Loken syndrome) along with retinal degeneration.
Leber's congenital amaurosis is a rare genetic disease that leads to degeneration of the light–sensitive cells of the retina of the eye. Leber's amaurosis can be caused by defects in at least 25 different genes. Currently, gene therapy has been approved for only one form of congenital amaurosis, other forms of the disease are not treatable. The type caused by mutations in NPHP5 is relatively rare. It leads to blindness in all cases, and is often accompanied by kidney failure. The research team collected stem cell samples from two patients with NPHP5 deficiency at the NIH Clinical Center. This material was used to create retinal organoids – cultured clusters of tissues that have many structural and functional features of the real retina. Retinal organoids grown from patients' cells are valuable because they accurately mimic the genotype and manifestations of the disease in real patients and provide a close-to-natural environment for testing experimental interventions, including gene therapy. As in patients, photoreceptor defects were found in the retinal organoids, including the loss of the protruding part of the receptor cells.
In a healthy retina, the outer segments of photoreceptors contain the photosensitive protein opsin. When it is exposed to light, the photoreceptor initiates a nerve impulse that enters the brain and provides vision. The outer segment of the photoreceptor is a special type of primary cilia.
It is believed that in a healthy eye, the NPHP5 protein is located in the ciliary gate, a structure at the base of the primary cilia that helps filter proteins entering the cilia. Previous studies in mice have shown a link between NPHP5 and cilia function, but the exact role of this protein in photoreceptor cilia was unknown.
AAV-NPHP5 restores the accumulation of rhodopsin in the outer segments of photoreceptors.
In the present study, reduced levels of NPHP5 protein were found in retinal organoids grown from patients' cells, as well as CEP-290 protein, which interacts with NPHP5 and forms primary ciliary gates. (Mutations in CEP-290 constitute the most common cause of Leber's amaurosis). In addition, the external segments of photoreceptors in the retinal organoids were completely absent, and the opsin protein, which should have been localized on the external segments, was instead found elsewhere in the body of photoreceptor cells.
Treatment of retinal organoids grown from cells taken from patients with Leber's amaurosis and healthy volunteers. Above: retinal organoids (DNA is colored blue, NPHP5 is colored red and rhodopsin is green). Below is an enlarged image of the photoreceptor layer of organoids (rhodopsin is colored green).
When the researchers introduced an adeno-associated viral (AAV) vector containing a working version of the NPHP5 gene, retinal organoids showed significant recovery of the opsin protein concentrated in the right place – in the outer segments. The obtained data also indicate that the functional gene NPHP5 may have stabilized the primary ciliary gate.
Article by K.Kruczek et al. In vitro modeling and rescue of ciliopathy associated with IQCB1/NPHP5 mutations using patient-derived cells is published in the journal Stem Cell Reports.
Aminat Adzhieva, portal "Eternal Youth" http://vechnayamolodost.ru based on the materials of the National Institutes of Health: NIH researchers develop gene therapy for rare ciliopathy.