Congenital hearing loss refers to impaired auditory function that occurs due to genetic causes. GJB2 is the gene responsible for approximately half of all cases of hereditary hearing loss. Connexin 26 (CX26), which is encoded by GJB2, helps in the formation of intercellular gap junctions—channels that allow for the movement of ions and chemical messenger molecules between adjacent cells, where it regulates auditory function.
GJB2 mutations often lead to fragmentation of gap junctions and gap junction plaques (GJPs) which are composed of CX26. While the inheritance of recessive GJB2 mutation containing two copies of the defective gene can be functionally cured via GJB2 gene replacement, GJB2 dominant-negative mutation where the mutant protein inhibits the normal functioning of the wild-type protein necessitates a gene editing approach.
In this light, researchers from Japan have successfully developed a gene therapy to repair R75W, a dominant-negative mutation of GJB2 that causes syndromic hearing loss. The research team included Associate Professor Dr. Kazusaku Kamiya and Assistant Professor Dr. Takao Ukaji in the Department of Otorhinolaryngology, Juntendo University Faculty of Medicine, Japan, and Dr. Osamu Nureki from the Department of Biological Sciences, Graduate School of Science, The University of Tokyo , Japan. Their research findings were published in Volume 10, Issue 5 of JCI Insight on March 10, 2025.
"The overwhelming majority of mutations causing hereditary hearing loss involve the GJB2 gene. However, treatments that can restore hearing in patients with genetic deafness are lacking," shares Dr. Kamiya. "Our research can contribute to the development of gene therapy to tackle the increasing incidence of hereditary hearing loss patients."
First, the researchers developed an AAV (AAV-Sia6e) that can deliver genome editing tools to a wide range of inner ear cells that form gap junctions. AAV is a useful vector for delivering genes; however, the size of the gene that can be carried is limited. Therefore, Dr. Kamiya and his team constructed a base editing tool (SaCas9-NNG-ABE8e) that was miniaturized to a size that can be carried by AAV using SaCas9-NNG, which has a smaller size and higher genome editing efficiency than conventional Cas9. Next, they loaded this base editing tool onto AAV-Sia6e, which has high infection tropism for inner ear cells and developed an all-in-one AAV vector for inner ear genome editing.
Genome editing via the all-in-one AAV vector showed considerable efficiency and specificity. It showed on-target T to C conversion in human cells, expressing the GJB2 R75W mutation, repaired the R75W mutation, and formed a clear GJP. Furthermore, after the base editing procedure, the physiological function of gap junction cell-cell communication was restored.
Finally, to validate their results, the researchers performed AAV-mediated genome editing in a transgenic mouse model with the GJB2 R75W mutation. Inner sulcus cells of the cochlea, which were infected with the all-in-one AAV vector, formed distinct GJPs with pentagonal or hexagonal structures arranged around cells that were similar to those observed in wild-type cells.
"By using an all-in-one AAV vector with high infectivity for the inner ear, we expect to improve the therapeutic effect, simplify the development process, and reduce costs. Furthermore, the ABE-based gene editing approach is expected to be less toxic and safer than the conventional CRISPR-Cas9 technology. The AAV genome editing therapy we developed can also be applied to the treatment of other genes that cause hearing loss," concludes Dr. Kamiya, elaborating on the potential implications of the research.
Taken together, these findings highlight the immense therapeutic potential of the AAV-mediated genome editing approach for treating hereditary hearing loss.
Reference
Authors |
Takao Ukaji1, Daisuke Arai1, Harumi Tsutsumi1, Ryoya Nakagawa2, Fumihiko Matsumoto1, Katsuhisa Ikeda1, Osamu Nureki2, and Kazusaku Kamiya1 |
Title of original paper |
AAV-mediated base editing restores cochlear gap junction in GJB2 dominant-negative mutation-associated syndromic hearing loss model |
Journal |
JCI Insight |
DOI |
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Affiliations |
1Department of Otorhinolaryngology, Juntendo University Faculty of Medicine, Tokyo, Japan 2Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Tokyo, Japan |
About Associate Professor Dr. Kazusaku Kamiya
Dr. Kazusaku Kamiya is an Associate Professor at the Department of Otorhinolaryngology, Juntendo University Faculty of Medicine, Japan. He specializes in ear diseases with expertise in areas such as stem cells, hearing loss, gap junction, mutation, and cell therapy. He is also the Representative Director and CEO of Gap Junction Therapeutics, Inc. Dr. Kamiya has over 50 publications and over 1,080 citations to his credit.
