Pathogenetics of the FOX transcription factor gene cluster on 16q24.1

描述（由申请方提供）：肺泡毛细血管发育不良伴肺静脉错位（ACD/MPV）是一种致死性新生儿弥漫性肺部发育障碍，通常与涉及心脏、胃肠道和泌尿生殖系统的多种先天性异常相关。患有ACD/MPV的婴儿在出生后的最初几天内会出现严重的呼吸窘迫伴肺动脉高压，尽管进行了重症监护，但通常很快就会死亡。最近，我们发现由于转录因子Forkhead Box F1（FOXF 1）在16q24.1上的点突变或基因组缺失导致的单倍不足导致ACD/MPV和广泛的先天性畸形。此外，我们已经确定了两个不同的FOXF 1上游的微缺失，暗示在疾病的发病机制的位置效应。在FOXF 1微缺失中遇到的多效性效应，如左心发育不良综合征和胃肠道闭锁，可能是由于相邻基因FOXC 2和FOXL 1的单倍不足，这两个基因都是16q24.1 FOX簇的一部分。杂合子Foxf 1小鼠死于肺出血，伴有肺泡化和血管发生沿着的严重缺陷以及其他器官异常，尽管它们并不完全重现人类的ACD/MPV。Foxf 1基因在发育过程中的表达表明了一种有趣的基因调控模式。我们假设，这种复杂的调节Foxf 1可能是由于位置效应和基因组印记在组织和时间特异性的方式; ACD/MPV也可以引起其他基因或FOXF 1调控元件的破坏;和肺缺陷的Foxf 1小鼠可以预防围产期通过增加剂量的Foxf 1蛋白在毛细血管内皮细胞和周围间充质。我们设计了三个目标来验证这些假设。在目标1中，我们将从两个方面剖析FOXF 1的基因调控。首先，将使用报告基因测定、ChIP芯片和染色质构象捕获（3C）技术鉴定和检测可能对FOX基因簇表达或FOXF 1特异性表达重要的调控元件。其次，我们将分析Foxf 1在小鼠中的分离和等位基因特异性表达。在目标2中，我们将利用目标1中研究获得的知识，筛选FOXF 1上游或下游调控元件和ACD/MPV候选基因中的点突变和拷贝数变异。最后，在目标3中，我们将通过使用基于腺病毒载体的Foxf 1基因转移在外周小鼠肺中探索治疗选择。ACD/MPV是一种致死性疾病，目前尚无可用的治疗方法。我们相信，使用病毒载体的基因治疗方法可能有可能通过逆转致命性毛细血管缺陷的异常形成来纠正ACD/MPV患者的致死性表型。此外，从风险：获益的角度来看，与该实验性治疗相关的风险可能是合理的，并且有可能在医院环境中转化。 公共卫生相关性：我们将阐明发病机制，并确定其他致病基因负责新生儿弥漫性发育障碍的肺，肺泡毛细血管发育不良与肺静脉错位（ACD/MPV），通常与多种先天性畸形，涉及心脏，胃肠道和泌尿生殖系统。这种致命的疾病目前尚无有效的治疗方法。我们认为，在小鼠中使用病毒载体的基因治疗方法可能有可能通过逆转致死性毛细血管缺陷的异常形成来纠正ACD/MPV患者的致死性表型。

DESCRIPTION (provided by applicant): Alveolar capillary dysplasia with misalignment of pulmonary veins (ACD/MPV) is a lethal neonatal diffuse developmental disorder of the lungs, which is commonly associated with multiple congenital anomalies involving the cardiac, gastrointestinal, and genitourinary systems. Infants affected with ACD/MPV develop severe respiratory distress with pulmonary hypertension within the first few days of life and despite intensive care they usually die soon thereafter. Recently, we have found that haploinsufficiency due to point mutations or genomic deletions of the transcription factor Forkhead Box F1 (FOXF1) on 16q24.1 results in ACD/MPV and a broad spectrum of congenital malformations. In addition, we have identified two distinct microdeletions upstream of FOXF1, implicating a position effect in the pathogenesis of the disease. Pleiotropic effects encountered in FOXF1 microdeletions, such as hypoplastic left heart syndrome and gastrointestinal atresias, may be due to haploinsufficiency for the neighboring genes, FOXC2 and FOXL1, both part of the FOX cluster at 16q24.1. Heterozygous Foxf1 mice die from pulmonary hemorrhage with severe defects in lung alveolarization and vasculogenesis along with other organ anomalies, although they do not completely recapitulate ACD/MPV in humans. The expression of the Foxf1 gene during development suggests an intriguing pattern of gene regulation. We hypothesize that this complex regulation of Foxf1 may be due to both position effects and genomic imprinting in a tissue- and time-specific manner; ACD/MPV can also be caused by disruption of other gene(s) or FOXF1 regulatory elements; and the lung defect in Foxf1 mice can be prevented perinatally by increasing the dosage of the Foxf1 protein in the capillary endothelium and surrounding mesenchyme. We have designed three aims to test these hypotheses. In aim 1, we will dissect gene regulation of FOXF1 in two ways. First, regulatory elements that may be important to the expression of the FOX gene cluster or to FOXF1 specific expression will be identified and tested using reporter assays, ChIP-on-chip, and chromatin conformation capture (3C) techniques. Second, we will analyze the segregation and allele-specific expression of Foxf1 in mice. In aim 2, we will use the knowledge gained from our studies in aim 1 to screen for point mutations and copy-number variations in the regulatory elements identified upstream or downstream to FOXF1 and in the ACD/MPV candidate genes. Finally, in aim 3, we will explore therapeutic options by use of adenoviral vector-based Foxf1 gene transfer in peripheral murine lungs. ACD/MPV is a lethal disorder and there is no available treatment at the present time. We believe that a gene therapy approach using viral vectors may have the potential to correct the lethal phenotype of ACD/MPV patients by reversing the abnormal formation of the lethal capillary defect. Moreover, the risks related to this experimental therapy may be justified from a risk: benefit standpoint and have potential to be translated in the hospital setting. PUBLIC HEALTH RELEVANCE: We will unravel the pathogenesis and identify other causative gene(s) responsible for a neonatal diffuse developmental disorder of the lungs, Alveolar capillary dysplasia with misalignment of pulmonary veins (ACD/MPV), commonly associated with multiple congenital malformations involving the cardiac, gastrointestinal, and genitourinary systems. This lethal disease has no available treatment at the present time. We believe that a gene therapy approach using viral vectors in mice may have the potential to correct the lethal phenotype of ACD/MPV patients by reversing the abnormal formation of the lethal capillary defect.