Validation of ubiquilin for Huntingtons disease

描述(申请人提供)：亨廷顿病(HD)是已知的最具破坏性的人类疾病之一，在美国大约有12万人受到影响。尽管进行了广泛的研究，但目前还没有有效的治疗方法。我们之前在HD的成纤维细胞、神经元和线虫模型上进行的研究表明，过表达泛素(Ubqln)可以降低含有扩展多谷氨酰胺重复序列的突变亨廷顿蛋白(HTT)的毒性。结果提示，调节ubqln的表达可能是治疗HD的一个有吸引力的靶点。然而，在这样的考虑之前，一个必要和重要的额外条件是验证ubqln的过度表达是否可以减少该病小鼠模型中的HD样症状。为了进行这样的测试，我们现在已经获得了在整个大脑中过表达人类ubqln-1的转基因小鼠，包括纹状体和海马体。根据寿命和行为分析，这些小鼠是完全正常的。在最初的测试中，我们将我们的ubqln-1转基因小鼠与HD的R6/2模型进行了杂交，发现ubqln-1的过度表达几乎完全恢复了在疾病晚期出现的R6/2动物大脑中ubqln水平的逐渐下降。有趣的是，ubqln-1过表达减少了HTT蛋白的聚集，并使R6/2小鼠的存活率提高了20%。尽管有这些改变，我们在小鼠的行为缺陷方面没有发现明显的差异，这可能归因于有毒的R6/2突变的HTT外显子-1转基因片段的高表达和外显性。重要的是，我们必须用更好的高清模型重复测试。两个这样的模型是YAC128和CAG敲入小鼠，这两个模型都在更生理的水平上表达突变的全长HTT蛋白。这些高清机型在较长的时间内具有更弱的HD症状，应该更适合我们的测试。因此，在目标1中，我们将过表达人泛素-1的转基因小鼠与YAC128转基因小鼠杂交，以评估泛素-1的过度表达是否延缓和/或阻止这种HD模型的神经病理和疾病症状的发展过程。目标2是目标1的重复，但不同之处在于我们将使用HD的CAG200敲入小鼠模型而不是YAC128小鼠。我们将用从小鼠分离的培养神经元的体外研究来补充这些动物研究，这将为泛素在调节HTT毒性中的作用模式提供机制。这项研究的结果将为ubqln过度表达是否可以延缓或预防HD症状提供一个关键的测试。如果得到证实，这一结果可能会对HD产生很大影响，因为它将表明，努力寻找调节泛素表达的因素可以成功地用于HD的治疗。我们描述了如何做到这一点的下一个合乎逻辑的步骤。

DESCRIPTION (provided by applicant): Huntington's disease (HD) is one of the most devastating human diseases known, affecting approximately 120,000 people in the USA. Despite extensive research there is currently no effective treatment for the disease. Our previous studies conducted in fibroblast, neuronal and C. elegans models of HD have shown that overexpression of ubiquilin (ubqln) reduces toxicity of mutant huntingtin (htt) proteins containing expanded polyglutamine repeats. The results suggest that modulation of ubqln expression might be an attractive target for treating HD. However, before such consideration, a necessary and important perquisite is to validate whether overexpression of ubqln can reduce HD-like symptoms in mouse models of the disease. To conduct such tests we have now obtained transgenic mice that overexpress human ubqln-1 throughout the brain, including the striatum and hippocampus. The mice are completely normal based on lifespan and behavioral assays. In an initial test we crossed our ubqln-1 transgenic mice with the R6/2 model of HD and found ubqln-1 overexpression almost completely restores a progressive decline in ubqln levels seen in the brains of R6/2 animals during late stages of the disease. Interestingly, ubqln-1 overexpression reduced htt protein aggregation and increased survival of R6/2 mice by 20%. Despite these alterations we found no significant difference in behavioral defects in the mice, which might be attributed to high expression and penetrance of the toxic R6/2 mutant htt exon- 1 transgenic fragment. It is important that we repeat the test with better models of HD. Two such models are the YAC128 and CAG knock-in mice, both of which express mutant full-length htt proteins at more physiological levels. These HD models have more attenuated HD symptoms over an extended period and should be better suited for our test. Therefore, in Aim 1 we will cross transgenic mice that overexpress human ubiquilin-1 with the YAC128 transgenic mice to evaluate whether ubiquilin-1 overexpression delays and/or prevents the time-course of development of neuropathology and disease symptoms in this model of HD. Aim 2 is a repetition of Aim 1, but differs in that we will use the CAG200 knock-in mouse model of HD instead of the YAC128 mice. We will complement these animal studies with in vitro studies using cultured neurons isolated from the mice that should provide mechanistic into the mode of ubiquilin action in regulating htt toxicity. The results from this study will provide a critical tet of whether ubqln overexpression can delay or prevent HD symptoms. If validated the results could have high impact for HD as it would suggest efforts to find factors that modulate ubiquilin expression could be successfully used for treatment of HD. We describe the next logical steps of how this could be done.