A new method of delivering genes to mice has restored their hearing, sometimes to normal levels. The same method could be used to deliver other large genes.
Hearing loss has been associated with mutations in more than 100 genes, but STRC, the second most frequent genetic cause, is responsible for up to 16 percent of hereditary hearing loss.
A groundbreaking gene therapy technique created at Boston Children’s Hospital successfully replaced the mutant protein stereocilin in the inner ear of mice, reversing severe hearing loss and restoring hearing to normal levels in some cases.
The findings were published in the journal Science Advances.
According to Jeffrey Holt, PhD, a specialist in the departments of Otolaryngology and Neurology at Boston Children’s Hospital and the study’s main investigator, the procedure could be utilized in other circumstances when the therapeutic gene is very large.
The scientists will now try the approach in human inner ear cells in a culture obtained from patients with STRC hearing loss to see if it works with the human stereocilin gene. Holt plans to seek the FDA approval to test gene therapy in humans if it restores hearing function at the tissue level.
Sensory hair cells of the inner ear must make contact with the tectorial membrane of the ear, which vibrates in response to sound, and then transform these vibrations into signals conveyed to the brain in order for sounds to be heard. The stereocilin protein works as a scaffold, allowing the microvilli of hair cells to stand up in a neat bundle so that their tips can touch the membrane.
“If stereocilin is mutated, you don’t have that contact, so the hair cells are not stimulated properly,” said Holt.
“But importantly, the hair cells still remain functional, so they are receptive to the gene therapy. We think this will provide a broad window of opportunity for treatment – from babies to adults with hearing loss.”
The researchers developed a synthetic adeno-associated virus (AAV) that targets hair cells to deliver a healthy stereocilin gene.
“The challenge we faced was that the gene for stereocilin is too big to fit into the gene therapy vector,” Holt said.
“The gene is about 6,200 DNA base pairs long, but the AAV only has a capacity of 4,700 base pairs.”
The study’s first author, Olga Shubina-Oleinik, PhD, came up with a solution: she split the mouse Strc gene in half and put the two halves in two different AAVs. She then employed a procedure known as protein recombination, which involves two halves of a protein finding one other and joining together. But it didn’t work in this case.
“We then realized that the beginning of the protein has a short stretch of amino acids that acts like an ‘address,’ directing the protein to its proper place in the cell,” Shubina-Oleinik said.
“When we split the protein in half, we realized that one half had the signal, but the other half did not, so the halves might not end up in the same location.”
When the signal was applied to both half of the protein, the two halves joined together successfully. The researchers discovered that full-length stereocilin protein was restored in the animals, as well as normal-appearing hair bundles that could contact the tectorial membrane.
The researchers utilized two types of hearing tests: one that was similar to baby hearing tests and another that employed electrodes on the scalp to detect auditory brainstem responses to a variety of sound frequencies and intensities. The mice were shown to be significantly more sensitive to subtle noises and to have improved cochlear amplification, which is the ability to amplify mild sounds, reduce the reaction to loud sounds, and differentiate between sounds of different frequencies with more precision. In some mice, the hearing was restored to normal levels.
“The results were remarkable and are the first example of hearing restoration using dual-vector gene therapy to target sensory outer hair cells,” said Shubina-Oleinik.
A patent application for gene treatment technique has been filed by Shubina-Oleinik and Holt. According to co-author Eliot Shearer, MD, PhD, a clinician-scientist in otolaryngology at Boston Children’s Hospital, about 100,000 people in the United States and 2.3 million people globally have STRC mutations and could benefit from this treatment.
“It turns out that STRC gene variations are more common than we thought, which makes gene therapy for this disorder so important,” added Shearer, who worked with the Children’s Rare Disease Cohort Initiative to screen a large genomic data set for STRC mutations.
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