Somatic mutation detection in brain AVM by massively high-throughput sequencing

描述（由申请人提供）：脑血管畸形影响超过300万美国人，并导致出血性中风和严重的神经系统残疾或死亡。脑动静脉畸形（AVMs）是两种最常见的血管畸形之一，是一种复杂的血管病变，由一团血管组成，这些血管将血液从动脉循环分流到静脉循环，而没有真正的毛细血管床介入。脑动静脉畸形的患病率为10 - 18 / 10万成年人，是年轻人出血性中风的重要原因。该病变的局部性和散发性和孤立性表明，有害的体细胞突变是脑AVM病因的基础。对于在家族性综合征HHT中发生的AVM，已经提出了两击假说，其中在种系中发生杂合突变的同一基因中的第二次体细胞突变或“击中”是AVM病变形成的必要条件。在散发性脑动静脉畸形中，首先发生的生殖系损伤可能是新生的；然后是第二次，躯体攻击，或者可能有两次躯体攻击。AVM病变复杂且异质性强；体细胞突变可能只存在于一小部分病变细胞中。因此，直到最近，检测avm的体细胞突变一直很困难。新一代大规模并行测序技术的出现为脑动静脉畸形和其他复杂病变的体细胞突变检测提供了一种新颖、高灵敏度和高通量的方法。为了验证体细胞突变是脑AVM病因学基础的假设，无论是遗传性（HHT）还是散发性形式的疾病，我们将实施大规模平行下一代测序技术，以检测从AVM病变组织提取的DNA中的体细胞突变。我们将使用该技术对来自100例散发性脑AVM患者和6例HHT患者的AVM病变组织中至少40个候选基因（包括HHT中杂合突变的3个基因）的250千碱基DNA进行重测序。验证用于血管病变体细胞突变检测的大规模高通量测序将是适用于各种血管和肿瘤病变的重要进展。体细胞突变假说的证明将是理解脑AVM病因机制的一个重要里程碑。从长远来看，我们将研究已发现的体细胞突变与该病临床病程之间的关系，以弥合理解基本生物学机制和AVM临床管理风险分层之间的鸿沟。

DESCRIPTION (provided by applicant): Vascular malformations of the brain affect over 3 million Americans, and cause hemorrhagic stroke and serious neurological disability or death in a significant proportion of affected individuals. Brain arteriovenous malformations (AVMs), one of the two most common types of vascular malformation, are complex vascular lesions, comprised of a tangle of blood vessels that shunts blood from the arterial to the venous circulation without an intervening true capillary bed. Brain AVMs have a population prevalence of 10 to 18 per 100,000 adults, and are an important cause of hemorrhagic stroke in young adults. The localized and often sporadic and solitary nature of the lesion suggests the hypothesis that deleterious somatic mutations underlie brain AVM etiology. For AVMs occurring in the familial syndrome HHT, the two- hit hypothesis has been proposed, where a second, somatic mutation or "hit" in the same gene that is heterozygously mutated in the germline is required for the AVM lesion to form. In sporadic brain AVMs, the first, germline, hit may be de novo; followed by a second, somatic hit, or there may be two somatic hits. AVM lesions are complex and heterogeneous; somatic mutations may only be present in a fraction of lesion cells. Therefore, until recently, detection of somatic mutations in AVMs has been difficult. The recent advent of massively-parallel next generation sequencing technology now suggests a novel, highly sensitive and high- throughput approach to somatic mutation detection in brain AVMs and other complex lesions. To test the hypothesis that somatic mutations underlie brain AVM etiology, both in the inherited (HHT) and the sporadic form of the disease, we will implement massively-parallel next generation sequencing technology to detect somatic mutations in DNA extracted from AVM lesion tissue. We will use this technology to resequence 250 kilobases of DNA from at least 40 candidate genes, including the 3 genes heterozygously mutated in HHT, in AVM lesion tissue from 100 sporadic brain AVM patients and 6 HHT patients. Validation of massively high throughput sequencing for somatic mutation detection in vascular lesions will be an important advance applicable to a variety of vascular and neoplastic lesions. Demonstration of the somatic mutation hypothesis will represent a significant milestone in understanding the mechanisms of brain AVM etiology. In the long term, we will study the relationship between somatic mutations identified and the clinical course of the disease, with a long-term goal of bridging the divide between understanding basic biological mechanisms and risk stratification for AVM clinical management. PUBLIC HEALTH RELEVANCE: Brain arteriovenous malformations (AVMs) are an important cause of hemorrhagic stroke in young adults, resulting in a considerable public health burden encompassing high healthcare costs and lost productivity. The causes of brain AVMs are poorly understood. We will investigate whether brain AVMs, both sporadic and those occurring in the familial disease Hereditary Hemorrhagic Telangiectasia, are caused by somatic mutations in the cells of the AVM lesion. This work will enhance understanding of the molecular mechanisms of AVM formation and hemorrhage, with the long-term goal of improving clinical management and developing new treatment approaches to lessen the negative burden AVMs on health.