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异质性神经系统疾病。虽然超过40个基因的突变与遗传性共济失调有关，基因检测已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来源不明。不明原因的共济失调的个体家族通常太小，无法通过传统的定位克隆方法进行基因鉴定;对这些病例中共济失调病因的缺乏了解继续阻碍准确诊断和有效和个性化治疗的新靶点的鉴定。最近，大规模并行测序允许有效发现基因组中的几乎所有变体，或至少其编码部分。然而，由于每个人都携带数千种新的变异，仅测序无法立即识别致病突变。在这里，我们假设，遗传连锁，测序为基础的变异发现，基因表达分析，而不是足够的信息单独提供补充信息，并允许识别新的共济失调基因在家庭中至少有两个隐性或三个显性受影响的成员。我们已经招募并开始分析12个这样的谱系，并开发了收集和分析相关基因组数据的专业知识。我们建议研究22个家系使用一个集成的管道，结合连锁分析，基因表达谱，下一代测序和基因网络分析。由于不同的共济失调家族可能携带不同的变异，这些技术都不足以单独使用，但它们的组合将提供高度的信息以确定候选人。将检测候选突变是否存在于对照样本中，并在不相关病例中筛选候选基因的其他突变。通过使用这样一个综合的方法，我们最近已经确定了一个听神经病基因，并发现了一个可信的新的候选基因显性中枢核肌病。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元细胞进行快速功能随访。这是通过将来自选定家族的活检培养物的成纤维细胞重编程为神经元，并测试潜在突变对mRNA剪接和量、对相关细胞类型内的其他基因以及对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鉴定，这是以前不可能使用纯遗传定位克隆策略。这些发现将使更好的诊断和预后，这将立即帮助受影响的家庭。它可能会导致个性化治疗，并产生新的假设来研究更常见的共济失调的散发形式。鉴定经验证的和功能特征化的分子病变有望促进知情的药物开发。我们的经验，这种方法也将建立一个有用的范例，为其他罕见的孟德尔疾病。 公共卫生相关性： 许多形式的共济失调的原因仍然是未知的，即使在家庭中的遗传明确指出了遗传原因。最近的基因组工具和下一代测序将使我们能够确定几个新的共济失调基因的小家庭。发现这些新基因不仅可以帮助受影响的家庭进行诊断，计划生育和预后，还可以帮助开发全面的共济失调基因网络，从而实现未来的个性化诊断，预后和治疗。

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. PUBLIC HEALTH RELEVANCE: The cause of many forms of ataxia is still unknown, even when inheritance in families clearly points to a genetic cause. Recent genomic tools and next generation sequencing will allow us to identify several novel ataxia genes in small families. Finding these new genes will not only help the affected families with diagnosis, family planning, and prognosis, but also aid in the development of comprehensive ataxia networks of genes that will lead to future personalized diagnosis, prognosis and treatment.