Transduction of the Mouse Auditory System with AAV

描述（由申请人提供）：超过2800万美国人患有 各种形式的听力损失和3000万以上暴露在危险的 噪音水平。缺乏对多种形式的后天性和后天性疾病的有效治疗， 遗传性听力障碍引起了人们对 新开发的基因传递技术，以恢复正常的耳蜗功能。 为了实现这一目标，许多载体已成功地用于介导 脑内基因递送包括脂质体、疱疹病毒、慢病毒， 腺病毒和腺相关病毒（AAV）。不幸的是，病毒载体（具有 AAV除外）将病毒基因引入转导的细胞，靶向 它们被免疫系统破坏并限制转基因的持续时间 表情此外，这些载体中有许多与 致病条件。相比之下，AAV作为基因特别有吸引力， 因为96%的病毒基因组已经被移除。此外，委员会认为， 它从未与疾病有关，具有广泛的宿主范围， 促进基因的长期表达。重要的是，6种不同的AAV血清型 已经分离出（AAV 1-6），它们各自的衣壳蛋白不同。 因此，这些血清型表现出中度至实质性的差异， 转导效率以及转导的细胞类型。AAV-2已经 用于耳蜗基因传递研究， 耳蜗毛细胞然而，AAV-2需要细胞表面的硫酸乙酰肝素 对于进入靶细胞，重要的是，毛细胞缺乏硫酸乙酰肝素。 因此，毫不奇怪，AAV-2不是成功的载体， 耳蜗内的基因转移。 我们建议系统地评估转基因传递到小鼠耳蜗 和螺旋神经节的研究。我们的目标 是确定每种AAV血清型将抑制的那些细胞群， 最有效。例如，其他AAV血清型如4和5不是 依赖于硫酸乙酰肝素进入靶细胞，因此可能 为基因传递到耳蜗提供了更好的载体。此外，AAV-5 最近显示使用唾液酸作为进入靶细胞的辅助受体 细胞毛细胞被认为在其表面表达唾液酸， 这表明AAV-5可以作为一种有效的基因递送载体， 细胞这些研究的另一个关键组成部分是比较 正常耳蜗和庆大霉素损伤耳蜗之间的转导效率。我们 选择用小鼠来开发这种转基因传递 系统，以便未来的基因转移研究可以利用众多的 小鼠的突变已被证明会影响听觉发育， 功能这些可行性研究代表了我们实验室的新方向 并将形成一个全面的研究计划的发展的基础。 这些未来研究的重点将是将特定的基因传递给 听觉系统，可能发挥作用的功能恢复， 耳蜗，以及耳蜗损伤后的神经保护。

DESCRIPTION (provided by applicant): More than 28 million Americans suffer from various forms of hearing loss and 30 million more are exposed to dangerous levels of noise. The lack of effective treatment for many forms of acquired and inherited hearing disorders has prompted interest in the potential application of newly developed gene delivery techniques to restore normal cochlear function. Towards this goal, a number of vectors have been successfully used to mediate intracochlear gene delivery including liposomes, herpes virus, lentivirus, adenovirus, and adeno-associated virus (AAV). Unfortunately, viral vectors (with the exception of AAV) introduce viral genes into the transduced cells, targeting them for destruction by the immune system and limiting the duration of transgene expression. In addition, many of these vectors have been associated with pathogenic conditions. In contrast, AAV is particularly attractive as a gene delivery system because 96% of its viral genome has been removed. Furthermore, it has never been associated with disease, has a broad host range, and facilitates long term gene expression. Importantly, 6 different AAV serotypes have been isolated (AAV 1-6) which differ in their respective capsid proteins. Consequently these serotypes exhibit moderate to substantial differences in the efficiency of transduction as well as the cell types transduced. AAV-2 has been used in cochlear gene delivery studies where it does not appear to transduce cochlear hair cells. However, AAV-2 requires heparan sulfate on the cell surface for entry into target cells and significantly, hair cells lack heparan sulfate. Therefore, it is not surprising that AAV-2 has not been a successful vector for gene transfer within the cochlea. We propose to systematically evaluate transgene delivery to the mouse cochlea and spiral ganglia using AAV serotypes 1-5 both in vitro and in vivo. Our goal is to determine those cell populations that each AAV serotype will transduce most effectively. For example, other AAV serotypes such as 4 and 5 are not dependent on heparan sulfate for entry into target cells, and therefore may provide a better vehicle for gene delivery to the cochlea. In addition, AAV-5 has recently been shown to use sialic acid as a coreceptor for entry into target cells. Hair cells are thought to express sialic acid on their surface, suggesting that AAV-5 may serve as an efficient gene delivery vehicle to these cells. Another critical component to these studies will be to compare transduction efficiencies between normal and gentamicin-damaged cochleae. We have chosen to use the mouse for the development of this transgene delivery system so that future gene transfer studies can take advantage of the numerous mouse mutations that have been shown to influence auditory development and function. These feasibility studies represent a new direction for our laboratory and will form the basis for the development of a full-scale research program. The focus of these future studies will be on delivering specific genes to the auditory system that potentially play a role in both the functional recovery of the cochlea, as well as neuroprotection following cochlear damage.