对脊髓小脑共济失调2型有治疗潜在意义的修饰基因的定位克隆及功能验证

Spinocerebellar ataxia type 2 (SCA2) is a neurological movement disorder caused by the atrophy of cerebellum or spinal cord. This disease was caused by mutations in ATXN2 gene and it is an autosomal dominant disease. There are still not effective treatments to control the symptoms of SCAs. We found a big pedigree of spinocerebellar ataxia type 2 (SCA2), and there are two kinds of symptoms in patients of this family: the Parkinson Disease (PD) symptoms and the SCAs symptoms. The cerebellums of Parkinsonism patients were protected, and the symptoms of PD patients can be controlled by L-Dopa treatment. We proposed that a mutation in a modify gene which may play a role of epistasis could change the SCA2 patients’ symptoms to Parkinsonism and protected their cerebellums. Therefore, if the modify gene can be positional cloned, there is possible to find the treatment methods to SCA2 according the mechanism of this modify gene change phenotype. The finding of mechanism of modify gene also can be used as a reference for treating some genetic neurological diseases which have similar pathogenesis with SCA2, such as Huntington’s disease，DRPLA and SCA1, 3, 6, 7, 17. For finding the modify gene, we will perform whole genome sequencing to the 8 patients in this pedigree. Following the principle that the patients of Parkinsonism phenotype should carry the mutation of modify gene and the patients of SCA2 should not, we will located the modify gene and find the mutation. Depend on the previous works, we already have found a possible modify gene HEXB. The HEXB gene was used as an example to perform functional verification experiment and discuss protective effect of modify gene on nerve cells in patients with SCA2. We want the example can increase the feasibility of the project.

脊髓小脑共济失调2型（SCA2）是一由小脑或脊髓萎缩而导致的运动性神经疾病，其由基因ATXN2突变所致并成常染色体显性遗传。现今仍无有效的治疗手段。本研究组发现了一个SCA2的家系，其患者表现出两种症状：帕金森病症状及普通SCA2症状，帕金森表型病人小脑得到了保护，并对多巴治疗敏感。本研究组认为一个起上位效应的“修饰基因”的突变可能造成了帕金森表型和小脑的保护。如能克隆此基因就有可能依其机理治疗该疾病。并且对一些病理相似的神经遗传病的治疗也非常有借鉴意义，如Huntington’s disease，DRPLA和SCA1, 3, 6, 7, 17。本研究组拟对家系中病人进行全基因组测序，结合家系遗传模式首先定位克隆该修饰基因并进行功能验证。基于前期的工作，本研究发现了一个可能的候选修饰基因HEXB，并以此基因为例子设计了功能验证实验讨论其对SCA2病人神经细胞的保护作用，希望增加项目可行性。

本研究组拟在一个患病成员表现有两种不同表型的脊髓小脑共济失调2型（SCA2）的家系中，寻找导致表型改变小脑受到保护的修饰基因。在对家系中8个患者进行全基因组测序并分析后，挑选出了83个候选修饰基因，并进行了功能候选的表达分析。但未能发现挑选出的12个基因明显与小脑保护相关。在2018年底，项目组对家系的持续观察发现家系中出现新的患者，并且第一次观察的某些重要表型有变化。因此在2019年初，项目组重新进行了家系调查，外周血采集及细致的医学检验。我们对22个家系中的成员均进行了全外显子组测序，在新信息的支持下，结合多个医院临床医生的意见，最终确定了5个候选修饰基因。同时对6个病人的进行全基因表达谱分析，发现了两个患者表型间主要的差异存在于细胞因子相互作用受体和癌症相关通路中的100多个基因上。阐明这些基因表达差异与5个候选修饰基因的关系，可能最终验证本项目的假说。我们有信心利用基金剩余款项完成这一验证。项目组成功建立条件表达的人源化ATXN2（SCA2致病基因）突变型和野生型小鼠模型，此模型对本项目最终猜想的验证及后续研究均有重要意义。在进行这个家系研究的同时，项目组也收集了一些其它神经萎缩及肌肉退行性病变的家系，希望从不同角度进行观察，获得一些启发。在对收集的200多个肌张力障碍核心家系中发现一个从未被报道过的突变，此新发突变（NM_000113.2:c.*302T>A，rs563498119）能通过改变TOR1A基因的表达而导致肌张力障碍。与此位点结合的hsa-miR-494能有效的抑制野生型TOR1A基因的表达。同时也发现了另一个位于TOR1A基因外显子的新发突变（NM_000113.2: c.860C>A, p. Ser287Tyr, rs766483672）其可以改变torsinA蛋白的二聚化从而危害细胞的核膜，与疾病的发生密切相关。这些发现对肌张力障碍的诊断和治疗均非常有意义。