Important Step Towards Curing Usher Syndrome

Usher Syndrome type 1F is an uncommon but severe hereditary condition that causes deafness, loss of balance, and blindness.
A team coordinated by Harvard Medical School, Massachusetts Eye and Ear, and The Ohio State University has now taken a vital first step toward developing a gene treatment for the illness.

The study, which was conducted in mice, was published in Nature Communications on April 26.

The researchers created a “mini-gene”—a reduced form of a gene—to replace the faulty gene in Usher 1F. The mutation prevents inner ear hair cells from generating a crucial protein involved in sound transmission. The mini-gene enhanced the synthesis of the missing protein in mice, allowing hair cells to sense sound and restore hearing.

Because vision loss in Usher 1F involves a slightly different form of the same protein, the researchers say the same approach may be useful for preventing blindness.

“Patients with Usher 1F are born with profound hearing loss and progressive vision loss, and so far we have been able to offer very few solutions to these families,” said co-senior author Artur Indzhykulian, HMS assistant professor of otolaryngology–head and neck surgery at Mass Eye and Ear.

The researchers plan to continue testing the mini-gene in other animal models, and eventually, hope to test it in humans.

“It’s completely devastating to be born deaf and then lose your vision, so we hope that this mini-gene can eventually be turned into a treatment for this disease,” said co-senior author David Corey, the Bertarelli Professor of Translational Medical Science in the Blavatnik Institute at HMS.

Children with Usher Syndrome are usually born deaf or with significantly impaired hearing, lack balance, and gradually lose eyesight as the retina deteriorates. Blindness is common by maturity.

These issues develop as a result of a mutation that interferes with the creation of protocadherin-15, a protein that has slightly different versions in the ear and eye and is required for cells in the auditory and visual systems to operate properly.

The Corey lab has long been interested in the role of protocadherin-15 in the inner ear. They were particularly interested in how the protein aids sensory receptors called hair cells in the ear in converting vibrations from the environment into electrical signals that the brain interprets as sound.

Corey’s group previously discovered how protocadherin-15 collaborates with another protein, cadherin 23, to form filaments in hair cells that physically pull open ion channels as the bundles vibrate, allowing electrical current to enter the cells. In the absence of this protein, electrical current cannot reach hair cells, vibration cannot be converted to electricity, and the brain cannot sense sound.

Corey became interested in developing a gene treatment for Usher Syndrome Type 1 as a result of this work. The therapy would include inserting DNA coding for protocadherin-15 into a cell, allowing the cell to begin producing the protein.

However, protocadherin-15’s DNA is too large for the normal viral capsule employed to transfer genetic material into a cell due to its size.

As a result, the researchers decided to try another approach: reducing the DNA to make a mini-gene that still codes for functioning protein but is small enough to fit within the viral capsule.

The first stage included meticulously mapping all 25,000 atoms in the exterior structure of inner-ear protocadherin-15, which was done by co-senior author Marcos Sotomayor, a former HMS research fellow who is now an associate professor of chemistry and biochemistry at The Ohio State University.

Sotomayor discovered that the protein is made up of atoms arranged in what appears to be 11 links in a chain using a combination of X-ray crystallography and cryo-electron microscopy.

Sotomayor created eight alternative variants of protocadherin-15, each with a different number of missing links to reduce the protein’s size. The shortened protein structures were then reverse-engineered into DNA blueprints that the researchers could test as mini-genes.

“The knowledge we gained by studying the structure of protocadherin-15 in excruciating detail allowed us to more quickly design shorter versions of the protein for gene therapy,” Sotomayor explained.

Indzhykulian examined the eight mini-genes in a dish on inner ear cells. He confirmed that truncated forms of protocadherin-15 derived from mini-gene DNA connect to cadherin 23, the protein partner of protocadherin-15 in hair cells.
The scientists then chose three mini-genes that were small enough to fit within the viral capsule.
Maryna Ivanchenko, an instructor in neurobiology at HMS, conducted extensive testing on the three mini-genes in the ears of mice that had been genetically engineered to stop making protocadherin-15. In the end, only one mini-gene worked.
The gene successfully induced hair cells to produce a miniature version of protocadherin-15, which binds to cadherin-23 and forms the filaments required to open ion channels. Vibrations were successfully transformed into electrical impulses by the hair cells.

“We were all pleasantly surprised,” Corey said. “We thought it would take years of optimizing and trying things and tweaking the protein structure, but this one version pretty much worked.”

“The results were thrilling for us,” Ivanchenko added. “The most exciting aspect of our findings was that mice that had been completely deaf could now hear almost as well as normal mice.”

While the mini-gene successfully repaired deafness in the Usher Syndrome Type 1 mouse model, the researchers are even more excited about its potential to treat blindness associated with the illness.

The authors believe that because children with Usher Syndrome Type 1 are born severely deaf and may lack hair cells in their inner ear, the mini-gene is unlikely to enhance their hearing. Furthermore, many of these youngsters are eligible for cochlear implants, which allow them to hear.

The researchers emphasized that blindness is a different situation because children with Usher 1F are born with normal vision. They believe that if the mini-gene could create the version of protocadherin-15 that is absent in the retina, it could prevent eyesight loss.

Why test the mini-gene in the mouse’s inner ear first?

The researchers stated that this was primarily for logistical reasons. In mice, a lack of protocadherin-15 causes only modest eyesight loss that proceeds slowly. This implies that testing the mini-genes in mouse models would take years, and it would be difficult to determine how effectively they operated. The mice, on the other hand, were born profoundly deaf, thus the researchers saw clear benefits within a few weeks.

“The whole project was designed to study the ear with the idea that something that works in the ear can later be applied to the eye, as an article of faith,” Corey said. “While the best test system is the mouse inner ear, the immediate goal is a treatment for blindness.”

The Corey lab is now testing the mini-gene in zebrafish eyes—a better model because these fish experience more severe and rapid vision loss than mice when protocadherin-15 isn’t produced in the retina.

If the mini-gene works in the zebrafish retina, the researchers will move to test the approach in primates and, eventually, in humans.

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