Fetal Pharmacotherapy for Congenital Deafness

描述（由申请人提供）：双侧永久性先天性听力损失估计影响全球每1000名活产婴儿中有1-3名新生儿。遗传基础在大约50%的个体中是因果关系。先天性耳聋或听力受损儿童的治疗选择在范围和疗效上是有限的，通常涉及助听器或人工耳蜗植入。两种干预都不能完全恢复原生听力的丰富性。该建议的概念基础是确定胎儿治疗方法，在发育中的内耳感觉上皮发生明显致病性变化之前纠正基因表达或功能缺陷。我们使用小鼠作为模型系统，因为存在不断扩大的自然和诱导基因突变阵列，这些突变可作为人类内耳功能障碍的准确范例。我们的技术方法依赖于实验胚胎学、外科手术、成像、显微注射和分子技术，这些技术允许在子宫内进入发育中的小鼠内耳，并对产生听觉和前庭感觉结构的前体细胞进行遗传操作。在本研究中，我们的目标是：1)恢复突触传递缺陷小鼠模型的听觉功能，2)恢复Usher综合征小鼠模型的听觉和前庭功能。编码泡状谷氨酸转运蛋白-3 （VGLUT3）的人类SLC17A8基因的错义突变与常染色体显性进行性高频非综合征性耳聋有关。VGLUT3选择性定位于耳蜗内毛细胞，并将兴奋性氨基酸神经递质谷氨酸装载到突触囊泡中。VGLUT3基因敲除小鼠由于内毛细胞不能在传入神经末梢释放谷氨酸而先天性失聪。

DESCRIPTION (provided by applicant): Bilateral permanent congenital hearing loss is estimated to affect 1-3 neonates per 1,000 live births worldwide. A genetic basis is causal in approximately 50% of the individuals. Treatment options for congenitally deaf or hearing impaired children are limited in scope and efficacy and typically involve hearing aids or cochlear implants. Neither intervention fully restores the richness of native hearing. The conceptual basis of this proposal is to define fetal therapies that correct defects in gene expression or function prior to the onset of overt pathogenic changes in the sensory epithelia of the developing inner ear. We use the mouse as a model system because an ever-expanding array of natural and induced genetic mutations exists that serve as accurate paradigms for human inner ear dysfunction. Our technical approach relies on experimental embryology, a palette of surgical, imaging, microinjection and molecular techniques that permit access to the developing mouse inner ear in utero and enable genetic manipulation of precursor cells that give rise to the auditory and vestibular sensory structures. In this proposal, we aim: 1) to restore auditory function in a mouse model defective in synaptic transmission~ and 2) to restore auditory and vestibular function in a mouse model of Usher syndrome. A missense mutation in the human SLC17A8 gene that encodes vesicular glutamate transporter-3 (VGLUT3) has been associated with an autosomal-dominant form of progressive, high-frequency nonsyndromic deafness. VGLUT3 is selectively localized to inner hair cells in the cochlea and loads the excitatory amino acid neurotransmitter glutamate into synaptic vesicles. The VGLUT3 knockout mouse is born deaf due to the inability of inner hair cells to release glutamate at the afferent nerve terminals. We hypothesize that virus-mediated gene transfer of VGLUT3 to otic epithelial precursors will force VGLUT3 expression in inner hair cells and restore synaptic transmission and hearing in the VGLUT3 knockout mouse. Usher syndrome is the leading genetic cause of combined deafness and blindness and is associated with 13 loci and 10 genes in humans. Three distinct clinical forms are differentiated by the time of onset and severity of auditory, vestibular, and visual dysfunction. Usher syndrome type 1 is the most severe with bilateral profound hearing loss and balance difficulty at birth and retinitis pigmentosa present by early adolescence. The USH1 gene encodes the protein harmonin that is expressed in the sensory hair cell bundle and the ribbon synapse. A single base pair mutation in the harmonin gene creates a cryptic splice site resulting in a frameshift and truncated harmonin protein. The USH1C mouse mutant is born deaf, uncoordinated, and develops vision impairment. We hypothesize that fetal administration of an antisense oligonucleotide will correct harmonin messenger RNA splicing and restore auditory and vestibular function in the USH1C mouse mutant. We are optimistic that the proposed studies will establish the first fetal therapies for congenital deafness and vestibular dysfunction and will help inform translation of these approaches to the clinic.