Postnatal Slc26a4 gene therapy improves hearing and structural integrity in a hereditary hearing loss model
Yi-Hsiu Tsai, Peng-Yu Wu, Yu-Chi Chuang, Chun-Ying Huang, Hiroki Takeda, Hiroshi Hibino, Chen-Chi Wu, Yen-Fu Cheng
Yi-Hsiu Tsai, Peng-Yu Wu, Yu-Chi Chuang, Chun-Ying Huang, Hiroki Takeda, Hiroshi Hibino, Chen-Chi Wu, Yen-Fu Cheng
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Research Article Genetics Otology

Postnatal Slc26a4 gene therapy improves hearing and structural integrity in a hereditary hearing loss model

Abstract

Mutations in SLC26A4 are the second most common cause of hereditary hearing loss (HL) in many Asian countries, leading to DFNB4, a condition characterized by progressive HL and inner ear malformations. While gene therapy holds great potential, its postnatal application has remained unexplored because of the lack of suitable animal models and the challenges of prenatal intervention. To our knowledge, this study represents the first preclinical investigation of postnatal gene therapy for DFNB4 using a clinically relevant Slc26a4-mutant mouse model that closely replicates human auditory phenotypes. Utilizing the synthetic AAV.Anc80L65 vector, we achieved robust SLC26A4 delivery to critical cochlear regions, including the endolymphatic sac and cochlear lateral wall. Comprehensive phenotypic analyses revealed a critical therapeutic window spanning the neonatal and juvenile stages, within which AAV.Anc80L65-mediated SLC26A4 delivery significantly improved hearing, as evidenced by lower auditory brainstem response thresholds. Moreover, the therapy preserved hair cells, reduced endolymphatic sac enlargement, partially restored the endocochlear potential, and mitigated inner ear structural degeneration. These therapeutic effects persisted into adulthood, highlighting the long-term efficacy of postnatal gene therapy. Together, these findings establish a critical therapeutic window for DFNB4 and demonstrate the feasibility of targeting the endolymphatic sac and cochlear lateral wall for effective intervention.

Authors

Yi-Hsiu Tsai, Peng-Yu Wu, Yu-Chi Chuang, Chun-Ying Huang, Hiroki Takeda, Hiroshi Hibino, Chen-Chi Wu, Yen-Fu Cheng

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Figure 1

Phenotypic characterization of auditory function and cochlear anatomical anomalies in L236P-mutant mice.

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Phenotypic characterization of auditory function and cochlear anatomical...
(A) L236P-mutant mice exhibited varying degrees of HL at P21, categorized into 3 groups: <60 dB (red, n = 23), 60–90 dB (green, n = 26), and >90 dB (blue, n = 10). WT mice served as controls (n = 16). (B) Representative images of endolymphatic sacs from WT and L236P mice across hearing groups at P21 (sacs are outlined by white dashed lines). Scale bars: 1 mm. (C) Quantification of the endolymphatic sac area at P21. Data are presented as the mean ± SD (n = 5–9 mice per group). ***P < 0.001, by 1-way ANOVA multiple comparisons with Tukey’s test). (D) Endolymphatic sac area at subsequent time points. Data represent the mean ± SD (n = 5–9 mice per group). (E) Representative images of marginal cells in the stria vascularis at P21. Scale bars: 50 μm. (F) Quantification of the surface area of marginal cells at P21. Data are presented as the mean ± SD (n = 45–105 cells per group). *P = 0.02 and ***P < 0.001, by 1-way ANOVA with Tukey’s test. (G) EP values for WT and L236P mice stratified by hearing thresholds: < 60 dB (n = 13); 60–90 dB (n = 20); and higher than 90 dB (n = 19). Data are presented as the mean ± SD. **P = 0.001 and *** P < 0.001, by Kruskal-Wallis test with Dunn’s test. (H) Hair cell morphology in the cochlear basal turn. Myosin VIIA (gray), phalloidin (red), and DAPI (blue) stainings are shown. Scale bars: 100 μm. (I and J) Quantification of basal IHCs and OHCs. Data are presented as the mean ± SD (n = 6). *P < 0.05, ** P < 0.01, and ***P < 0.001, by 2-way ANOVA with Dunnett’s test.