Project 1: Molecular Impact of VWF on Clinical VWD

VWD的齐默尔曼生物学计划(ZPB-VWD)中的项目1侧重于分子影响 VWF对临床VWD的影响。它整合了现有的两个队列-1)以前的追溯PPG队列 在Zimmerman计划的VWD分子和临床生物学诊断和2)展望 R01队列新兵。在目标1中，实验室研究是对1型、2型和3型VWD受试者进行纵向研究 以及VWF水平降低的组，我们统称为低von Willebrand因子，包括 那些以前可能被标记为“轻度VWD”的患者，VWF&lt；正常，但很少或没有出血。 这些可能被认为是出血的危险因素，但不是疾病。我们不仅要研究 他们的实验室表型随时间的变化，但也研究他们和中期出血评估 评分(IBAT)半定量地评估“新的临床出血量”，因为蝙蝠评估出血 但分数已经饱和，无法量化临时出血。此外，目标1将量化评估 与新的基质蛋白(如肌球蛋白)或新的功能性受体(如血小板糖蛋白)的潜在异常结合 IIB-IIIa(2b3a)。在Aim 2中，将研究类型2功能变体以确定诊断(类型)的保真度 2A、2B、2M和2N)在局部表型分配到集中分配的表型分配之间。大鼠的动物模型 已经/将开发2型变种，并使用出血和血栓形成测试(Tail)进行比较 出血、模板出血、静脉流量和激光损伤评估。目标3将在我们的 患有1或3型VWD或LVWF的合并队列，并且在这些现有队列中添加类似的现有 来自金斯敦和都柏林的已定义队列，但没有VWF突变，并且阵列比较结果为阴性 基因组杂交(ACGH)或多重连接依赖的探针扩增(MLPA)。在我们的 队列，我们至少有3名3型VWD受试者没有VWF序列变异，aCGH阴性，并且 没有抗VWF抗体。此外，我们还有40个类型1 VWD和180个LVWF主题样本，其中包括 NSV和aCGH阴性。因此，我们将首先对这些类型1和3的受试者进行全基因组研究 使用核心B中的NGS测序进行测序(WGS)。当识别出单个候选基因时， 将对受影响的家庭成员(AFM)和未受影响的家庭成员(UFM)进行排序，以确定 与表型连锁。然后，候选人SV将被外包到项目1-4，取决于它们是否 1)VWF内含子改变(项目1)；2)涉及碳水化合物或碳水化合物受体(项目2)；3) VWF编码区以外的机制(项目3)；或4)暗示表观遗传机制(项目4)。 对于可能被识别的常见或科学上独特的原因，这些动物模型将在 C核心，并在各个项目中进行了研究。通过项目1，我们希望对疾病有深入的了解 引起VWD的机制及其通过发育的动物对实验室和临床表型的影响 模型，并协助确定1型和3型VWD的外源原因。

Project 1 in the Zimmerman Program for the Biology of VWD (ZPB-VWD) is focused on the Molecular Impact of VWF on Clinical VWD. It consolidates the two existing cohorts – 1) retrospective PPG cohort previously diagnosed in the Zimmerman Program for the Molecular and Clinical Biology of VWD and 2) the prospective R01 cohort recruits. In Aim 1 laboratory studies are done longitudinally on the Type 1, 2, and 3 VWD subjects as well as the group with reduced VWF levels that we collectively call Low von Willebrand Factor that includes those that might have been previously labeled as “Mild VWD” with VWF <normal but little or no bleeding. These may be considered as having a risk factor for bleeding but not a disease. Not only will we study the changes with time of their laboratory phenotype but also study them with and interim Bleeding Assessment Score (iBAT) to semiquantitatively assess the amount of “new clinical bleeding” since BATs assess bleeding but the score saturates and cannot quantify interim bleeding. Additionally, Aim 1 will quantifiably evaluate potential abnormal binding to new matrix proteins (e.g. myosin) or new functional receptors (e.g. platelet GP IIb-IIIa (2b3a). In Aim 2 type 2 functional variants will be studied to determine the fidelity of diagnosis (types 2A, 2B, 2M, and 2N) between phenotypic assignments locally to those centrally assigned. Animal models of type 2 variants have been/will be developed and compared using bleeding and thrombosis testing (tail bleeding, template bleeding, VenaFlux, and Laser-injury assessment. Aim 3 will include subjects in our combined cohort who have type 1 or 3 VWD or LVWF and to these existing cohorts add existing, similarly defined, cohort from Kingston and Dublin but have no VWF mutation and are negative by Array Comparative Genome Hybridization (aCGH) or Multiplex Ligation-dependent Probe Amplification (MLPA). Within our cohort, we have at least 3 type 3 VWD subjects with no VWF sequence variant, are negative by aCGH, and have no anti-VWF antibodies. In addition we have >40 type 1 VWD and >180 LVWF subject samples with NSV and negative aCGH. Therefore we will first study these type 1 and 3 subjects by whole Genome Sequencing (WGS) using NGS sequencing in Core B. When individual candidate genes are identified, sequencing will be performed on the affected (AFM) and unaffected family members (UFM) to ascertain linkage with phenotype. Candidate SV will then be farmed out to Projects 1-4, based upon whether they are 1) VWF intronic changes (Project 1); 2) involve carbohydrates or carbohydrate receptors (Project 2); 3) mechanisms outside the VWF coding region (Project 3); or 4) suggest an epigenetic mechanism (Project 4). For frequent or scientifically unique causes that might be identified, animal models of these will be produced in Core C and studied in the various projects. Through Project 1 we expect to gain insight into the disease mechanisms causing VWD and how they affect laboratory and clinical phenotypes through developed animal models, and to assist with determining extragenic causes of both type 1 and type 3 VWD.