Treatment of monogenic and digenic dominant genetic hearing loss by CRISPR-Cas9 ribonucleoprotein delivery in vivo

Abstract Mutations in Atp2b2, an outer hair cell (OHC) gene, cause dominant hearing loss (HL) in humans. Using a mouse model Atp2b2Obl+, with a dominant HL mutation (Oblivion), we show that liposome-mediated in vivo delivery of CRISPR-Cas9 ribonucleoprotein (RNP) complexes leads to specific editing of the Obl allele. Large deletions encompassing the Obl locus and indels were identified as the result of editing. In vivo genome editing promotes OHC survival and restores OHC function, leading to robust hearing rescue. We further show in a double-dominant mutant mouse model, in which the Tmc1 Beethoven mutation and the Atp2b2 Oblivion mutation cause di-genic genetic HL, RNP delivery targeting both mutations lead to hearing rescue. These findings suggest that liposome-RNP delivery can be used as a strategy to rescue hearing with dominant mutations in OHC genes and with di-genic mutations in the auditory hair cells, expanding therapeutics of gene editing to treat HL.