Resolving Uncertainty in Alagille Syndrome Diagnostics
解决阿拉吉尔综合征诊断中的不确定性
基本信息
- 批准号:10734881
- 负责人:
- 金额:$ 58.15万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2023
- 资助国家:美国
- 起止时间:2023-07-01 至 2028-04-30
- 项目状态:未结题
- 来源:
- 关键词:AccountingAffectAlagille SyndromeAtlasesBenignBiological AssayCalibrationCell LineCellsChildhoodClassificationClinVarClinicalComplexDNADataDetectionDevelopmentDiagnosisDiagnosticDiseaseEtiologyEvaluationExonsEyeFaceFaciesFormalinFutureGene ExpressionGene Expression ProfileGene Expression ProfilingGenesGeneticGenetic DiseasesGenetic VariationGenomic medicineGenomicsGenotypeHeartHigh PrevalenceIndividualKidneyKnowledgeLibrariesLigandsLiverLiver diseasesManaged CareMeasuresMembraneMendelian disorderMethodologyMethodsMissense MutationModelingMolecularMutationNotch Signaling PathwayNucleotidesParaffin EmbeddingPathogenicityPatient CarePatientsPersonsPhenotypePositioning AttributePropertyProtein TruncationProteinsProteomicsReceptor SignalingReporterReportingResolutionSamplingSeriesSeverity of illnessSignal TransductionSiteSystemTechniquesTestingTherapeuticTissue SampleUncertaintyValidationVariantWorkautosomeclinical diagnosticsdesigndisorder controlexperimental studyglycosylationhigh throughput screeningimprovedinnovationliver transplantationloss of functionmultiplex assaymutantnotch proteinprotein functionreceptorsample fixationspine bone structuretooltranscriptomicsvariant of unknown significance
项目摘要
PROJECT SUMMARY/ABSTRACT
Uncertainty in genomic diagnostics creates a barrier to realizing the full potential of genomic medicine.
Uncertainty is evident in: 1) our inability to determine if some DNA variants are pathogenic or benign and 2) our
inability to predict to what extent a person with a disease-causing variant will be affected, due to variable
expressivity. This proposal will study both phenomena for the autosomal dominant disorder Alagille syndrome,
caused by mutations in one of two genes in the Notch signaling pathway, the ligand JAGGED1 (JAG1) or the
receptor, NOTCH2. Alagille syndrome is characterized by pediatric liver, heart, vertebral, renal, ocular, and
facial anomalies with highly variable expressivity. The mechanism of disease for JAG1-related Alagille
syndrome is haploinsufficiency whereas the mechanism for NOTCH2-related Alagille syndrome is less clear,
with fewer reported variants, less functional characterization, and a higher prevalence of missense variants
(>50%). Missense variants are difficult to classify, often requiring functional validation to support or reject
pathogenicity. In Alagille syndrome, functional characterization has been carried out for only 19/125 reported
missense mutations, thus, despite a high detection rate, the diagnostic rate is lower due to this uncertainty.
We propose to resolve uncertainty in the diagnostics of Alagille syndrome using assays designed to
characterize the pathogenicity of JAG1 and NOTCH2 missense variants and analysis of gene expression data
from patient liver samples to identify gene expression signatures that can be used for genotype-phenotype
evaluations. In Aim 1, we will design a Site Saturation Variant Library of all possible nucleotide permutations at
each nucleotide position across a region with high missense variant uncertainty in the JAG1 C-terminus and
test this library by developing a Multiplexed Assay for Variant Effects (MAVEs) that will measure cellular
localization of JAG1 as a readout of protein function. In Aim 2, we will use FFPE liver tissue samples to
analyze gene expression differences between Alagille syndrome patients and controls, as well as between
Alagille syndrome patients with mild versus severe liver disease. In Aim 3, we will study the molecular basis of
NOTCH2 variants through functional, expression, and enzymatic assays using mutant cell lines. We
hypothesize that these proposed assays will identify a high-throughput method to test missense pathogenicity
(Aim 1), identify gene expression differences between Alagille syndrome patients and controls as well as gene
expression signatures that are different between Alagille syndrome patients with mild versus severe liver
disease (Aim 2), and determine the mechanism by which NOTCH2 variants cause Alagille syndrome through
functional analysis (Aim 3). Ultimately, these data will improve variant analysis for Alagille syndrome,
improve our understanding of the molecular basis of liver disease in Alagille syndrome, and establish
a framework for scalable classification of missense variants, delivering diagnostic information that can
directly help clinicians.
项目概要/摘要
基因组诊断的不确定性为实现基因组医学的全部潜力造成了障碍。
不确定性明显表现在:1)我们无法确定某些 DNA 变异是致病性还是良性的;2)我们的
由于变量的存在,无法预测携带致病变异的人会受到多大程度的影响
表现力。该提案将研究常染色体显性遗传疾病 Alagille 综合征的这两种现象,
由 Notch 信号通路中两个基因之一的突变引起,即配体 JAGGED1 (JAG1) 或
受体,NOTCH2。 Alagille 综合征的特点是小儿肝脏、心脏、椎骨、肾脏、眼部和
表现力高度可变的面部异常。 JAG1相关Alagille的发病机制
综合征是单倍体不足,而 NOTCH2 相关的 Alagille 综合征的机制尚不清楚,
报告的变异较少,功能表征较少,错义变异的发生率较高
(>50%)。错义变体很难分类,通常需要功能验证来支持或拒绝
致病性。在 Alagille 综合征中,仅对 19/125 报道的患者进行了功能表征
因此,尽管错义突变的检出率很高,但由于这种不确定性,诊断率较低。
我们建议使用旨在解决 Alagille 综合征诊断的不确定性
表征 JAG1 和 NOTCH2 错义变异的致病性并分析基因表达数据
从患者肝脏样本中识别可用于基因型-表型的基因表达特征
评价。在目标 1 中,我们将设计一个位点饱和变体库,其中包含所有可能的核苷酸排列
JAG1 C 末端具有高错义变异不确定性的区域中的每个核苷酸位置和
通过开发变异效应多重检测 (MAVE) 来测试该库,该检测将测量细胞
JAG1 的定位作为蛋白质功能的读数。在目标 2 中,我们将使用 FFPE 肝组织样本
分析 Alagille 综合征患者和对照之间以及之间的基因表达差异
患有轻度与重度肝病的阿拉吉尔综合征患者。在目标 3 中,我们将研究其分子基础
使用突变细胞系通过功能、表达和酶测定来检测 NOTCH2 变体。我们
假设这些提议的测定将确定一种高通量方法来测试错义致病性
(目标 1),识别 Alagille 综合征患者和对照之间的基因表达差异以及基因
轻度和重度肝脏 Alagille 综合征患者之间的表达特征不同
疾病(目标 2),并通过以下方式确定 NOTCH2 变异导致 Alagille 综合征的机制
泛函分析(目标 3)。最终,这些数据将改善阿拉吉尔综合征的变异分析,
提高我们对 Alagille 综合征肝病分子基础的理解,并建立
错义变异可扩展分类的框架,提供可以
直接帮助临床医生。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Nancy Bettina Spinner其他文献
Nancy Bettina Spinner的其他文献
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{{ truncateString('Nancy Bettina Spinner', 18)}}的其他基金
Training Program in the Genetic Basis of Pediatric Gastrointestinal Disorders
儿科胃肠道疾病遗传基础培训计划
- 批准号:
8666845 - 财政年份:2014
- 资助金额:
$ 58.15万 - 项目类别:
Training Program in the Genetic Basis of Pediatric Gastrointestinal Disorders
儿科胃肠道疾病遗传基础培训计划
- 批准号:
8883521 - 财政年份:2014
- 资助金额:
$ 58.15万 - 项目类别:
Training Program in the Genetic Basis of Pediatric Gastrointestinal Disorders
儿科胃肠道疾病遗传基础培训计划
- 批准号:
8666845 - 财政年份:2014
- 资助金额:
$ 58.15万 - 项目类别:
Genetic Modifiers of Liver Disease Severity in Alagille Syndrome
阿拉吉尔综合征肝病严重程度的基因修饰
- 批准号:
7661203 - 财政年份:2009
- 资助金额:
$ 58.15万 - 项目类别:
Genetic Modifiers of Liver Disease Severity in Alagille Syndrome
阿拉吉尔综合征肝病严重程度的基因修饰
- 批准号:
8306850 - 财政年份:2009
- 资助金额:
$ 58.15万 - 项目类别:
Genetic Modifiers of Liver Disease Severity in Alagille Syndrome
阿拉吉尔综合征肝病严重程度的基因修饰
- 批准号:
8090799 - 财政年份:2009
- 资助金额:
$ 58.15万 - 项目类别:
Genetic Modifiers of Liver Disease Severity in Alagille Syndrome
阿拉吉尔综合征肝病严重程度的基因修饰
- 批准号:
7883529 - 财政年份:2009
- 资助金额:
$ 58.15万 - 项目类别:
Genetic Modifiers of Liver Disease Severity in Alagille Syndrome
阿拉吉尔综合征肝病严重程度的基因修饰
- 批准号:
8502652 - 财政年份:2009
- 资助金额:
$ 58.15万 - 项目类别:
Genetic Modifiers of Liver Disease Severity in Alagille Syndrome
阿拉吉尔综合征肝病严重程度的基因修饰
- 批准号:
8097573 - 财政年份:2009
- 资助金额:
$ 58.15万 - 项目类别:
NOTCH SIGNALING PATHWAY LIGANDS IN CARDIOVASCULAR DISEASE
心血管疾病中的 NOTCH 信号通路配体
- 批准号:
6565108 - 财政年份:2002
- 资助金额:
$ 58.15万 - 项目类别:
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