Ataxia gene identification by integrated genomic analysis

描述(由申请人提供)： 共济失调是由环境和遗传因素引起的一组不同类型的神经系统疾病。虽然40多个基因的突变与遗传性共济失调有关，基因检测已经成为越来越常规的临床实践，但大多数散发性和许多遗传性病例仍然原因不明。患有不明原因共济失调的个体家庭往往太小，无法通过传统的位置克隆方法进行基因鉴定；在这些病例中，缺乏对共济失调病因的了解，继续阻碍准确诊断和确定有效和个性化治疗的新靶点。最近，大规模并行测序可以有效地发现基因组中几乎所有的变异，或者至少是它的编码部分。然而，由于每个人携带数以千计的新变种，仅靠测序不能立即识别致病突变。在这里，我们假设，遗传连锁、基于测序的变异发现和基因表达分析，虽然分别不能提供足够的信息，但可以提供补充信息，并允许在至少有两个隐性或三个显性患病成员的家庭中识别新的共济失调基因。我们已经招募了12个这样的家系并开始分析，并开发了收集和分析相关基因组数据的专业知识。我们建议使用一个结合了连锁分析、基因表达谱分析、下一代测序和基因网络分析的综合流水线来研究22个家系。由于不同的共济失调家系可能携带不同的变异，单靠这些技术是不够的，但它们的结合将为准确定位候选人提供高度的信息。候选突变将在对照样本中进行检测，候选基因将在不相关的病例中筛选出额外的突变。通过使用这种综合方法，我们最近发现了一个听神经病基因，并发现了一个可信的新的候选基因，用于显性中枢性核肌病。我们也有几个共济失调家系的初步数据。除了对互补的基因组数据集进行全球分析外，我们还致力于使用适当的神经细胞进行快速功能跟踪。这是通过将选定家族的活组织培养物中的成纤维细胞重新编程为神经元，并测试潜在突变对剪接和mRNA数量、相关细胞类型内其他基因和细胞表型的影响来实现的。我们预测，这条管道将导致新的罕见共济失调基因突变的识别，这是以前使用纯基因定位克隆策略所不可能的。这些发现将使更好的诊断和预后成为可能，这将立即帮助受影响的家庭。它可能导致个性化治疗，并产生新的假说来研究更常见的散发性共济失调。确认有效的和具有功能特征的分子损伤有望促进知情的药物开发。我们使用这种方法的经验也将为其他罕见的孟德尔疾病建立一个有用的范例。

DESCRIPTION (provided by applicant): Ataxias are a heterogeneous group of neurological disorders caused by environmental as well as genetic factors. While mutations in more than 40 genes have been implicated in hereditary ataxia, and genetic testing has become an increasingly routine clinical practice, most sporadic and many inherited cases are still of unknown origin. Individual families with unexplained ataxia are often too small to allow gene identification by traditional positional cloning approaches; and the lack of understanding of ataxia etiology in these cases continues to hamper accurate diagnosis and identification of novel targets for effective and personalized therapy. Recently, massively parallel sequencing allows efficient discovery of nearly all variants in a genome, or at least its coding portion. However, as each person carries thousands of novel variants, sequencing alone does not permit immediate identification of causative mutations. Here, we hypothesize that genetic linkage, sequencing-based variant discovery, and gene expression analysis, while not sufficiently informative separately, provide complementary information and allow identification of novel ataxia genes in families with a minimum of two recessive or three dominantly affected members. We already have recruited and started to analyze 12 such pedigrees, and have developed expertise to collect and analyze the relevant genomic data. We propose to study 22 pedigrees using an integrated pipeline that combines linkage analysis, gene expression profiling, next-generation sequencing, and gene network analysis. Since different ataxia families likely carry distinct variants, none of these techniques alone will be sufficient, but their combination will be highly informative to pinpoint candidates. Candidate mutations will be tested for absence in control samples, and candidate genes will be screened for additional mutations in unrelated cases. By using such an integrated approach we have recently identified an auditory neuropathy gene and have found a credible novel candidate gene for dominant central nuclear myopathy. We also have promising preliminary data for several ataxia pedigrees. In addition to global analyses of complementary genomic datasets, we are committed to rapid functional follow-up using appropriate neuronal cells. This is achieved by reprogramming fibroblasts from biopsy cultures from selected families into neurons, and testing the effect of potential mutations on splicing and amount of mRNA, on other genes within the relevant cell type, and on cellular phenotypes. We predict that this pipeline will lead to the identification of new rare ataxia gene mutations that were not previously possible using purely genetic positional cloning strategies. The discoveries will enable better diagnosis and prognosis which will immediately help the affected families. It may lead to personalized treatment & generate new hypotheses to study the more common sporadic forms of ataxia. The identification of validated & functionally characterized molecular lesions is expected to facilitat informed drug development. Our experience with this approach will also establish a useful paradigm for other rare Mendelian disorders.