Although genetic factors contribute to almost half of all cases of deafness, treatment options for genetic deafness are limited1,2,3,4,5. We developed a genome-editing approach to target a dominantly inherited form of genetic deafness. Here we show that cationic lipid-mediated in vivo delivery of Cas9–guide RNA complexes can ameliorate hearing loss in a mouse model of human genetic deafness. We designed and validated, both in vitro and in primary fibroblasts, genome editing agents that preferentially disrupt the dominant deafness-associated allele in the Tmc1 (transmembrane channel-like gene family 1) Beethoven (Bth) mouse model, even though the mutant Tmc1Bth allele differs from the wild-type allele at only a single base pair. Injection of Cas9–guide RNA–lipid complexes targeting the Tmc1Bth allele into the cochlea of neonatal Tmc1Bth/+ mice substantially reduced progressive hearing loss. We observed higher hair cell survival rates and lower auditory brainstem response thresholds in injected ears than in uninjected ears or ears injected with control complexes that targeted an unrelated gene. Enhanced acoustic startle responses were observed among injected compared to uninjected Tmc1Bth/+ mice. These findings suggest that protein–RNA complex delivery of target gene-disrupting agents in vivo is a potential strategy for the treatment of some types of autosomal-dominant hearing loss.
Sequence Read Archive
This work was supported by DARPA HR0011-17-2-0049 (to D.R.L.), US NIH R01 EB022376 (to D.R.L.), R35 GM118062 (to D.R.L.), R01 DC006908 (to Z.-Y.C), P30 DC05209 (to M.C.L.), R01 DC00138 (to M.C.L.), and R01 DC013521 (to J.R.H.). We are grateful for support from the David-Shulsky Foundation (to Z.-Y.C.), a Frederick and Ines Yeatts Hair Cell Regeneration grant (to Y.T., V.L., M.H., and Y.S.), the Bertarelli Foundation and the Jeff and Kimberly Barber Fund (to J.R.H.), the Broad Institute (to D.R.L and Z.-Y.C.), and the HHMI (to D.R.L.). We thank H. Rees, S. Tsai, M. Packer, K. Zhao and D. Usanov for assistance.
Extended data figures
This file contains a representative example of the gating strategy used in the flow cytometry experiments in Extended Data Figure 1a.
This file contains supplementary methods which includes tables 1 and 2, sequences, notes and figure 1.