Cochlear gene therapy with ancestral AAV in adult mice: complete transduction of inner hair cells without cochlear dysfunction

Abstract

The use of viral vectors for inner ear gene therapy is receiving increased attention for treatment of genetic hearing disorders. Most animal studies to date have injected viral suspensions into neonatal ears, via the round window membrane. Achieving transduction of hair cells, or sensory neurons, throughout the cochlea has proven difficult, and no studies have been able to efficiently transduce sensory cells in adult ears while maintaining normal cochlear function. Here, we show, for the first time, successful transduction of all inner hair cells and the majority of outer hair cells in an adult cochlea via virus injection into the posterior semicircular canal. We used a “designer” AAV, AAV2/Anc80L65, in which the main capsid proteins approximate the ancestral sequence state of AAV1, 2, 8, and 9. Our injections also transduced ~10% of spiral ganglion cells and a much larger fraction of their satellite cells. In the vestibular sensory epithelia, the virus transduced large numbers of hair cells and virtually all the supporting cells, along with close to half of the vestibular ganglion cells. We conclude that this viral vector and this delivery route hold great promise for gene therapy applications in both cochlear and vestibular sense organs.

A dual‐AAV approach restores fast exocytosis and partially rescues auditory function in deaf otoferlin knock‐out mice

“…Nevertheless, improving the large transgene reconstitution efficacy by simply increasing the amount of available vector genomes does not improve protein expression levels as shown for dual‐AAV vectors in the retina (Carvalho et al , ). Since the IHC transduction rates of our single AAV2/6 eGFP‐expressing virus reached 99%, it is unlikely that other AAV serotypes such as AAV2/Anc80 (Landegger et al , ; Suzuki et al , ) will increase the amount of fully functional otoferlin protein levels in IHCs.…”

Selective targeting of unipolar brush cell subtypes by cerebellar mossy fibers

“…Semicircular canal injection was done following (Suzuki et al, 2017) using AAV2-retro-CAG-GFP (1.0E13 GC/ml,Janelia Farm), AAV2-retro-CAG-tdTomato (7.0E12 GC/ml, Addgene) or AAV-PHP.S-CAG-tdTomato (1.7E13, Addgene) (Chan et al, 2017) under isoflurane anesthesia. Briefly, a small hole we bored into the posterior semicircular canal using a 27 ga needle.…”

A CEP290 C-Terminal Domain Complements the Mutant CEP290 of Rd16 Mice In Trans and Rescues Retinal Degeneration

“…However, two of the nine treated mice exhibited a remarkably lower threshold at 32 kHz, suggesting that the treatment improved their sensitivity to higher frequency sounds (Figure S6A). Although functional rescue was not statistically significant due to the inherent variability of the treatment to P2 mice (Isgrig et al, 2017; Suzuki et al, 2017), immunofluorescence analysis revealed more inner hair cells in treated cochlea than those of untreated cochlea (Figures S6B–S6D). The hair cells at the cochlear base are tonotopically tuned for high-frequency sounds (Davis and Silverman, 1970).…”

“…To test whether the myo-tail vector could complement the mutation in cochlear hair cells, which are responsible for auditory sensation, the vector was injected into the posterior semicircular canal at P2 (Isgrig et al, 2017; Suzuki et al, 2017), and the mice were examined for auditory brainstem response (ABR) at 3 months of age. The mice displayed more severe auditory deficits when responding to higher frequency sound, because a significantly elevated ABR threshold was detected in all untreated mice when a 32-kHz stimulus was given (Figure S6A).…”