Otoferlin (OTOF) gene mutations represent the primary cause of hearing impairment and deafness in auditory neuropathy. The c.2485C>T (p. Q829X) mutation variant is responsible for approximately 3% of recessive prelingual deafness cases within the Spanish population. Previous studies have used two recombinant AAV vectors to overexpress OTOF, albeit with limited efficacy. In this study, we introduce an enhanced mini-dCas13X RNA base editor (emxABE) delivered via an AAV9 variant, achieving nearly 100% transfection efficiency in inner hair cells. This approach is aimed at treating OTOF^Q829X, resulting in an approximately 80% adenosine-to-inosine conversion efficiency in humanized Otof^Q829X/Q829X mice. Following a single scala media injection of emxABE targeting OTOF^Q829X (emxABE-T) administered during the postnatal day 0–3 period in Otof^Q829X/Q829X mice, we observed OTOF expression restoration in nearly 100% of inner hair cells. Moreover, auditory function was significantly improved, reaching similar levels as in wild-type mice. This enhancement persisted for at least 7 months. We also investigated P5–P7 and P30 Otof^Q829X/Q829X mice, achieving auditory function restoration through round window injection of emxABE-T. These findings not only highlight an effective therapeutic strategy for potentially addressing OTOF^Q829X-induced hearing loss but also underscore emxABE as a versatile toolkit for treating other monogenic diseases characterized by premature termination codons.

耳铁蛋白 （OTOF） 基因突变是听觉神经病变中听力障碍和耳聋的主要原因。c.2485C>T （p. Q829X） 突变变体导致西班牙人群中约 3% 的隐性舌前耳聋病例。以前的研究使用两种重组 AAV 载体过表达 OTOF，尽管疗效有限。在这项研究中，我们引入了一种通过 AAV9 变体递送的增强型 mini-dCas13X RNA 碱基编辑器 （emxABE），在内毛细胞中实现了近 100% 的转染效率。这种方法旨在治疗 OTOF^Q829X，导致人源化 Otof^Q829X/Q829X 小鼠的腺苷到肌苷转化效率约为 80%。在 Otof^Q829X/Q829X 小鼠出生后第 0-3 天期间注射靶向 OTOF^Q829X （emxABE-T） 的单次标量培养基注射后，我们观察到近 100% 的内毛细胞中 OTOF 表达恢复。此外，听觉功能得到显著改善，达到与野生型小鼠相似的水平。这种增强持续了至少 7 个月。我们还研究了 P5-P7 和 P30 Otof^Q829X/Q829X 小鼠，通过圆窗注射 emxABE-T 实现听觉功能恢复。这些发现不仅强调了一种可能解决 OTOF^Q829X 诱导的听力损失的有效治疗策略，而且还强调了 emxABE 是治疗其他以过早终止密码子为特征的单基因疾病的多功能工具包。